WO2025003344A1 - Type ii cas proteins and applications thereof - Google Patents

Abstract

Type II Cas proteins, for example Type II Cas proteins referred to as DGHJ, CUAZ, AHWY, CBGI, ASDR, BCZZ, DRCY, DRPY, EAJR, and CFDE Type II Cas proteins; gRNAs for Type II Cas proteins; systems comprising Type II Cas proteins and gRNAs; nucleic acids encoding the Type II Cas proteins, gRNAs and systems; particles comprising the foregoing; pharmaceutical compositions of the foregoing; and uses of the foregoing, for example to alter the genomic DNA of a cell.

Description

TYPE II CAS PROTEINS AND APPLICATIONS THEREOF 1. CROSS-REFERENCE TO RELATED APPLICATIONS [0001] This application claims the priority benefit of U.S. provisional application no.63/510,713, filed June 28, 2023 the contents of which are incorporated herein in their entireties by reference thereto. 2. SEQUENCE LISTING [0002] The instant application contains a Sequence Listing which has been submitted electronically in XML format and is hereby incorporated by reference in its entirety. Said XML Sequence Listing, created on June 21, 2024, is named ALA-012WO_SL.xml and is 689,481 bytes in size. 3. BACKGROUND [0003] CRISPR-Cas genome editing with Type II Cas proteins and associated guide RNAs (gRNAs) is a powerful tool with the potential to treat a variety of genetic diseases. Adeno-associated viral vectors (AAVs) are commonly used to deliver Cas proteins, for example Streptococcus pyogenes Cas9 (SpCas9), and their guide RNAs (gRNAs).The discovery of novel nucleases with new PAM specificities can broaden the range of targetable sites in the cell genome, making genome editing more flexible and efficient. 4. SUMMARY [0004] This disclosure is based, in part, on the discovery of a Type II Cas protein from an unclassified Firmicutes bacterium ^UHIHUUHG^WR^KHUHLQ^DV^³ZLOG-type DGHJ Type II &DV´^, a Type II Cas protein from an unclassified bacterium from the Prochlorococcus genus ^UHIHUUHG^WR^KHUHLQ^DV^³ZLOG-type CUAZ Type II &DV´^, a Type II Cas protein from an unclassified bacterium from the Eggerthellaceae family (referred to KHUHLQ^DV^³ZLOG-type AHWY Type II &DV´), a Type II Cas protein from an unclassified bacterium from the Clostridia class ^UHIHUUHG^WR^KHUHLQ^DV^³ZLOG-type CBGI Type II &DV´), a Type II Cas protein from an unclassified bacterium from the Eggerthellaceae family ^UHIHUUHG^WR^KHUHLQ^DV^³ZLOG-type ASDR Type II &DV´^^^D^7\SH^,,^&DV^SURWHLQ^IURP^an unclassified bacterium from the Clostridium genus (referred to KHUHLQ^DV^³ZLOG-type BCZZ 7\SH^,,^&DV´^^^D^7\SH^,,^&DV^SURWHLQ^IURP^Ellagibacter isourolithinifaciens ^UHIHUUHG^WR^KHUHLQ^DV^³ZLOG-type DRCY 7\SH^,,^&DV´^^^D^7\SH^,,^&DV^SURWHLQ^IURP a Lachnospiraceae bacterium ^UHIHUUHG^WR^KHUHLQ^DV^³ZLOG-type DRPY 7\SH^,,^&DV´^^^D^7\SH^,,^&DV^SURWHLQ^IURP^an unclassified bacterium from the Ruminococcaceae family ^UHIHUUHG^WR^KHUHLQ^DV^³ZLOG-type EAJR Type II &DV´^^^ and a Type II Cas protein from an unclassified Acholeplasmatales bacterium (referred to herein DV^³ZLOG-type CFDE 7\SH^,,^&DV´^. [0005] In one aspect, the disclosure provides Type II Cas proteins whose amino acid sequence comprises an amino acid sequence that is at least 50% identical (e.g., at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 85%, at least 90%, at least 95% identical, or more) to SEQ ID 12^^^^VXFK^SURWHLQV^UHIHUUHG^WR^KHUHLQ^DV^³DGHJ 7\SH^,,^&DV^SURWHLQV´^^^([HPSODU\^DGHJ Type II Cas protein sequences are set forth in SEQ ID NO:1, SEQ ID NO:2, and SEQ ID NO:3. [0006] In one aspect, the disclosure provides Type II Cas proteins whose amino acid sequence comprises an amino acid sequence that is at least 50% identical (e.g., at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 85%, at least 90%, at least 95% identical, or more) to SEQ ID NO:7 ^VXFK^SURWHLQV^UHIHUUHG^WR^KHUHLQ^DV^³CUAZ 7\SH^,,^&DV^SURWHLQV´^^^([HPSODU\^CUAZ Type II Cas protein sequences are set forth in SEQ ID NO:7, SEQ ID NO:8, and SEQ ID NO:9. [0007] In one aspect, the disclosure provides Type II Cas proteins whose amino acid sequence comprises an amino acid sequence that is at least 50% identical (e.g., at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 85%, at least 90%, at least 95% identical, or more) to SEQ ID NO:13 ^VXFK^SURWHLQV^UHIHUUHG^WR^KHUHLQ^DV^³AHWY 7\SH^,,^&DV^SURWHLQV´^^^([HPSODU\^AHWY Type II Cas protein sequences are set forth in SEQ ID NO:13, SEQ ID NO:14, and SEQ ID NO:15. [0008] In one aspect, the disclosure provides Type II Cas proteins whose amino acid sequence comprises an amino acid sequence that is at least 50% identical (e.g., at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 85%, at least 90%, at least 95% identical, or more) to SEQ ID NO:19 ^VXFK^SURWHLQV^UHIHUUHG^WR^KHUHLQ^DV^³CBGI 7\SH^,,^&DV^SURWHLQV´^^^([HPSODU\^CBGI Type II Cas protein sequences are set forth in SEQ ID NO:19, SEQ ID NO:20, and SEQ ID NO:21. [0009] In one aspect, the disclosure provides Type II Cas proteins whose amino acid sequence comprises an amino acid sequence that is at least 50% identical (e.g., at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 85%, at least 90%, at least 95% identical, or more) to SEQ ID NO:25 ^VXFK^SURWHLQV^UHIHUUHG^WR^KHUHLQ^DV^³ASDR 7\SH^,,^&DV^SURWHLQV´^^^([HPSODU\^ASDR Type II Cas protein sequences are set forth in SEQ ID NO:25, SEQ ID NO:26, and SEQ ID NO:27. [0010] In one aspect, the disclosure provides Type II Cas proteins whose amino acid sequence comprises an amino acid sequence that is at least 50% identical (e.g., at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 85%, at least 90%, at least 95% identical, or more) to SEQ ID NO:31 (such proteins referred WR^KHUHLQ^DV^³BCZZ 7\SH^,,^&DV^SURWHLQV´^^^([HPSODU\^BCZZ Type II Cas protein sequences are set forth in SEQ ID NO:31, SEQ ID NO:32, and SEQ ID NO:33. [0011] In one aspect, the disclosure provides Type II Cas proteins whose amino acid sequence comprises an amino acid sequence that is at least 50% identical (e.g., at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 85%, at least 90%, at least 95% identical, or more) to SEQ ID NO:37 ^VXFK^SURWHLQV^UHIHUUHG^WR^KHUHLQ^DV^³DRCY Type II Cas prRWHLQV´^^^([HPSODU\^DRCY Type II Cas protein sequences are set forth in SEQ ID NO:37, SEQ ID NO:38, and SEQ ID NO:39. [0012] In one aspect, the disclosure provides Type II Cas proteins whose amino acid sequence comprises an amino acid sequence that is at least 50% identical (e.g., at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 85%, at least 90%, at least 95% identical, or more) to SEQ ID NO:43 ^VXFK^SURWHLQV^UHIHUUHG^WR^KHUHLQ^DV^³DRPY 7\SH^,,^&DV^SURWHLQV´^^^([HPSODU\^DRPY Type II Cas protein sequences are set forth in SEQ ID NO:43, SEQ ID NO:44, and SEQ ID NO:45. [0013] In one aspect, the disclosure provides Type II Cas proteins whose amino acid sequence comprises an amino acid sequence that is at least 50% identical (e.g., at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 85%, at least 90%, at least 95% identical, or more) to SEQ ID NO:49 ^VXFK^SURWHLQV^UHIHUUHG^WR^KHUHLQ^DV^³EAJR 7\SH^,,^&DV^SURWHLQV´^^^([HPSODU\^EAJR Type II Cas protein sequences are set forth in SEQ ID NO:49, SEQ ID NO:50, and SEQ ID NO:51. [0014] In one aspect, the disclosure provides Type II Cas proteins whose amino acid sequence comprises an amino acid sequence that is at least 50% identical (e.g., at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 85%, at least 90%, at least 95% identical, or more) to SEQ ID NO:55 ^VXFK^SURWHLQV^UHIHUUHG^WR^KHUHLQ^DV^³CFDE 7\SH^,,^&DV^SURWHLQV´^^^([HPSODU\^CFDE Type II Cas protein sequences are set forth in SEQ ID NO:55, SEQ ID NO:56, and SEQ ID NO:57. [0015] In another aspect, the disclosure provides Type II Cas proteins comprising an amino acid sequence having at least 50% (e.g., at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 85%, at least 90%, at least 95%, or more) sequence identity to a RuvC-I domain, RuvC-II domain, RuvC-III domain, BH domain, REC1_1 domain, REC2 domain, REC1_2 domain, HNH domain, Topo domain or CTD of a DGHJ Type II Cas protein, CUAZ Type II Cas protein, AHWY Type II Cas protein, CBGI Type II Cas protein, ASDR Type II Cas protein, BCZZ Type II Cas protein, DRCY Type II Cas protein, DRPY Type II Cas protein, EAJR Type II Cas protein, or CFDE Type II Cas protein. In some embodiments, a Type II Cas protein of the disclosure is a chimeric Type II Cas protein, for example, comprising one or more domains from a DGHJ, CUAZ, AHWY, CBGI, ASDR, BCZZ, DRCY, DRPY, EAJR, and/or CFDE Type II Cas protein(s) and one or more domains from a different Type II Cas protein such as SpCas9. [0016] In some embodiments, the Type II Cas proteins of the disclosure are in the form of a fusion protein, for example, comprising a DGHJ Type II Cas protein, CUAZ Type II Cas protein, AHWY Type II Cas protein, CBGI Type II Cas protein, ASDR Type II Cas protein, BCZZ Type II Cas protein, DRCY Type II Cas protein, DRPY Type II Cas protein, EAJR Type II Cas protein, or CFDE Type II Cas protein sequence fused to one or more additional amino acid sequences, for example, one or more nuclear localization signals and/or one or more tags. Other exemplary fusion partners can enable base editing (e.g., where the fusion partner is nucleoside deaminase) or prime editing (e.g., where the fusion partner is a reverse transcriptase). [0017] Exemplary features of Type II Cas proteins of the disclosure are described in Section 6.2 and specific embodiments 1 to 287 and 736 to 737, infra. [0018] In further aspects, the disclosure provides guide (gRNA) molecules, for example single guide RNAs (sgRNAs). In various embodiments, the disclosure provides gRNAs that can be used with the DGHJ Type II Cas proteins of the disclosure, gRNAs that can be used with the CUAZ Type II Cas proteins of the disclosure, gRNAs that can be used with the AHWY Type II Cas proteins of the disclosure, gRNAs that can be used with the CBGI Type II Cas proteins of the disclosure, gRNAs that can be used with the ASDR Type II Cas proteins of the disclosure, gRNAs that can be used with the BCZZ Type II Cas proteins of the disclosure, gRNAs that can be used with the DRCY Type II Cas proteins of the disclosure, gRNAs that can be used with the DRPY Type II Cas proteins of the disclosure, gRNAs that can be used with the EAJR Type II Cas proteins of the disclosure, and gRNAs that can be used with the CFDE Type II Cas proteins of the disclosure. Exemplary features of the gRNAs of the disclosure are described in Section 6.3 and specific embodiments 288 to 608, infra. [0019] In further aspects, the disclosure provides systems comprising a Type II Cas protein of the disclosure and one or more gRNAs, e.g., sgRNAs. For example, a system can comprise a ribonucleoprotein (RNP) comprising a Type II Cas protein complexed with a gRNA, e.g., an sgRNA or separate crRNA and tracrRNA. Exemplary features of systems are described in Section 6.4 and specific embodiments 609 to 680, infra. [0020] In another aspect, the disclosure provides nucleic acids and pluralities of nucleic acids encoding a Type II Cas protein of the disclosure and, optionally, a guide RNA, for example a sgRNA. In some embodiments, the nucleic acids comprise a Type II Cas protein of the disclosure operably linked to a heterologous promoter, e.g., a mammalian promoter, for example a human promoter. [0021] In another aspect, the disclosure provides nucleic acids encoding a gRNA, for example a sgRNA, of the disclosure and, optionally, a Type II Cas protein, for example a DGHJ Type II Cas protein, CUAZ Type II Cas protein, AHWY Type II Cas protein, CBGI Type II Cas protein, ASDR Type II Cas protein, BCZZ Type II Cas protein, DRCY Type II Cas protein, DRPY Type II Cas protein, EAJR Type II Cas protein, or CFDE Type II Cas protein. [0022] Exemplary features of nucleic and pluralities of nucleic acids of the disclosure are described in Section 6.5 and specific embodiments 681 to 735, infra. [0023] In further aspects, the disclosure provides particles comprising the Type II Cas proteins, gRNAs, nucleic acids, and systems of the disclosure. Exemplary features of particles of the disclosure are described in Section 6.6 and specific embodiments 741 to 756, infra. [0024] In another aspect, the disclosure provides cells and populations of cells containing or contacted with a Type II Cas protein, gRNA, nucleic acid, plurality of nucleic acids, system, or particle of the disclosure. Exemplary features of such cells and cell populations are described in Section 6.6 and specific embodiments 758 to 766 and 794, infra. [0025] In another aspect, the disclosure provides pharmaceutical compositions comprising a Type II Cas protein, gRNA, nucleic acid, plurality of nucleic acids, system, particle, cell, or population of cells together with one or more excipients. Exemplary features of pharmaceutical compositions are described in Section 6.7 and specific embodiment 757, infra. [0026] In another aspect, the disclosure provides methods of altering cells (e.g., editing the genome of a cell) using the Type II Cas proteins, gRNAs, nucleic acids, systems, particles, and pharmaceutical compositions of the disclosure. Cells altered according to the methods of the disclosure can be used, for example, to treat subjects having a disease or disorder, e.g., genetic disease or disorder. Features of exemplary methods of altering cells are described in Section 6.8 and specific embodiments 767 to 793, infra. 5. BRIEF DESCRIPTION OF THE FIGURES [0027] FIGS.1A-1B schematically show a hairpin structure generated for visualization after in silico folding using RNA folding form v2.3 (www.unafold.org) of exemplary sgRNA scaffolds (not including the spacer sequence) designed from crRNAs and tracrRNAs identified for DGHJ Type II Cas protein. FIG.1A shows a longer version (SEQ ID NO:103) (v1) while FIG.1B shows a shorter version (SEQ ID NO:104) (v2) where the last stem-ORRS^ORFDWHG^DW^WKH^^¶-end of the scaffold is truncated. [0028] FIG.2 schematically shows a hairpin structure generated for visualization after in silico folding using RNA folding form v2.3 (www.unafold.org) of exemplary sgRNA scaffold (SEQ ID NO:105) (not including the spacer sequence) designed from crRNA and tracrRNA identified for CUAZ Type II Cas protein. For this particular structure, the folding was obtained by setting the folding temperature to 30°C. [0029] FIGS.3A-3B schematically show a hairpin structure generated for visualization after in silico folding using RNA folding form v2.3 (www.unafold.org) of exemplary sgRNA scaffolds (not including the spacer sequence) designed from crRNAs and tracrRNAs identified for AHWY Type II Cas protein. FIG. 3A shows a longer version (SEQ ID NO:107) (v1) while FIG.3B shows a shorter version (SEQ ID NO:108) (v2) where the last stem-loop locateG^DW^WKH^^¶-end of the scaffold is truncated. [0030] FIG.4 schematically shows a hairpin structure generated for visualization after in silico folding using RNA folding form v2.3 (www.unafold.org) of an exemplary sgRNA scaffold (SEQ ID NO:109) (not including the spacer sequence) designed from crRNAs and tracrRNAs identified for CBGI Type II Cas protein. [0031] FIG.5 schematically shows a hairpin structure generated for visualization after in silico folding using RNA folding form v2.3 (www.unafold.org) of an exemplary sgRNA scaffold (SEQ ID NO:110) (not including the spacer sequence) designed from crRNAs and tracrRNAs identified for ASDR Type II Cas protein. [0032] FIG.6 schematically shows a hairpin structure generated for visualization after in silico folding using RNA folding form v2.3 (www.unafold.org) of an exemplary sgRNA scaffold (SEQ ID NO:111) (not including the spacer sequence) designed from crRNAs and tracrRNAs identified for BCZZ Type II Cas protein. [0033] FIG.7 schematically shows a hairpin structure generated for visualization after in silico folding using RNA folding form v2.3 (www.unafold.org) of an exemplary sgRNA scaffold (SEQ ID NO:112) (not including the spacer sequence) designed from crRNAs and tracrRNAs identified for DRCY Type II Cas protein. [0034] FIG.8 schematically shows a hairpin structure generated for visualization after in silico folding using RNA folding form v2.3 (www.unafold.org) of an exemplary sgRNA scaffold (SEQ ID NO:113) (not including the spacer sequence) designed from crRNAs and tracrRNAs identified for DRPY Type II Cas protein. [0035] FIG.9 schematically shows a hairpin structure generated for visualization after in silico folding using RNA folding form v2.3 (www.unafold.org) of an exemplary sgRNA scaffold (SEQ ID NO:114) (not including the spacer sequence) designed from crRNAs and tracrRNAs identified for EAJR Type II Cas protein. [0036] FIG.10 schematically show a hairpin structure generated for visualization after in silico folding using RNA folding form v2.3 (www.unafold.org) of an exemplary sgRNA scaffold (SEQ ID NO:115) (not including the spacer sequence) designed from crRNAs and tracrRNAs identified for CFDE Type II Cas protein. [0037] FIGS.11A-11D show PAM sequence logos for DGHJ Type II Cas obtained using a longer (v1, FIG.11A) or shorter version of the sgRNA scaffold (v2, FIG.11C) in an in vitro PAM discovery assay and PAM enrichment heatmaps calculated from the same in vitro PAM discovery assay showing the nucleotide preferences at different positions along the PAM for DGHJ Type II Cas (positions 1, 2 and 3, 4) in combination with the same long (FIG.11B) and short (FIG.11D) sgRNA scaffolds. [0038] FIGS.12A-12B show a PAM sequence logo for CUAZ Type II Cas from an in vitro PAM discovery assay (FIG.12A) and the PAM enrichment heatmap calculated from the same in vitro PAM discovery assay showing the nucleotide preferences at different positions along the PAM for CUAZ Type II Cas (positions 2, 3 and 5, 7; FIG.12B). [0039] FIGS.13A-13D show PAM sequence logos for AHWY Type II Cas obtained using a longer (v1, FIG.13A) or shorter version of the sgRNA scaffold (v2, FIG.13C) in an in vitro PAM discovery assay and PAM enrichment heatmaps calculated from the same in vitro PAM discovery assay showing the nucleotide preferences at different positions along the PAM for AHWY Type II Cas (positions 2, 3 and 4, 6) in combination with the same long (FIG.13B) and short (FIG.13D) sgRNA scaffolds. [0040] FIGS.14A-14B show a PAM sequence logo for CBGI Type II Cas from an in vitro PAM discovery assay (FIG.14A) and the PAM enrichment heatmap calculated from the same in vitro PAM discovery assay showing the nucleotide preferences at different positions along the PAM for CBGI Type II Cas (positions 3, 4 and 5, 6; FIG.14B). [0041] FIGS.15A-15B show a PAM sequence logo for ASDR Type II Cas from an in vitro PAM discovery assay (FIG.15A) and the PAM enrichment heatmap calculated from the same in vitro PAM discovery assay showing the nucleotide preferences at different positions along the PAM for ASDR Type II Cas (positions 3, 4 and 5, 6; FIG.15B). [0042] FIGS.16A-16B show a PAM sequence logo for BCZZ Type II Cas from an in vitro PAM discovery assay (FIG.16A) and the PAM enrichment heatmap calculated from the same in vitro PAM discovery assay showing the nucleotide preferences at different positions along the PAM for BCZZ Type II Cas (positions 1,2 and 3,4; FIG.16B). [0043] FIGS.17A-C shows a PAM sequence logo for DRCY Type II Cas as predicted in silico (FIG. 17A) and determined in vitro (FIG.17B); and PAM enrichment heatmap (FIG.17C) calculated from the same in vitro PAM discovery assay showing the nucleotide preferences at different positions along the PAM for DRCY Type II Cas [0044] FIGS.18A-18F show PAM sequence logos for DRPY Type II Cas obtained in combination with its an exemplary DRPY sgRNA scaffold (FIG.18A) or DRCY sgRNA scaffold (FIG.18D) using an in vitro PAM discovery assay. PAM enrichment heatmaps calculated from the same in vitro PAM discovery assay showing the nucleotide preferences at different positions along the PAM for DRPY Type II Cas in combination with its the exemplary DRPY sgRNA (FIG.18B-18C) and DRCY sgRNA (FIG.18E-18F) scaffold. [0045] FIGS.19A-19B show a PAM sequence logo for EAJR Type II Cas from an in vitro PAM discovery assay (FIG.19A) and the PAM enrichment heatmap calculated from the same in vitro PAM discovery assay showing the nucleotide preferences at different positions along the PAM for EAJR Type II Cas (positions 2, 4 and 5, 6; FIG.19B). [0046] FIGS.20A-20B show a PAM sequence logo for CFDE Type II Cas from an in vitro PAM discovery assay (FIG.20A) and the PAM enrichment heatmap calculated from the same in vitro PAM discovery assay showing the nucleotide preferences at different positions along the PAM for CFDE Type II Cas (positions 2, 3 and 4, 5; FIG.20B). [0047] FIG.21 shows the activity of Type II Cas proteins against an EGFP reporter in mammalian cells. ³Y^´^DQG^³Y^´^UHIHU^WR^WZR^GLIIHUHQW^VJ51$^VFDIIROGV^HYDOXDWHG^ZLth AHWY and DGHJ Type II Cas proteins. Data presented as mean ± SEM of n=2 biologically independent runs. [0048] FIGS.22A-22C show the editing activity of AHWY, ASDR, CBGI, CFDE and DRPY Type II Cas in combination with panels of sgRNAs targeting the B2M (FIG.22A), TRAC (FIG.22B), PD-1 (FIG.22C) after transient plasmid transfection in HEK293T cells. Data presented as mean ± SEM for n=2 independent runs. [0049] FIGS.23A-23C show the editing activity of ASDR, CFDE and DRPY Type II Cas in combination with selected sgRNAs targeting TRAC, B2M, and PD1 (only for DRPY Type II Cas) after transient mRNA transfection in primary T cells. Indels (FIGS. 23A and 23C) and target downregulation as measured by cytofluorimetry (FIG.23B) are shown. Comparative data have been generated for the three loci also for the benchmark nuclease SpCas9 using previously published sgRNAs. Data presented as mean ± SEM for n=2 independent runs, except for the B2M guide for SpCas9 in (FIGS.23A-23B) and for panel (FIG.23C) where n=1. [0050] FIG.24 shows the editing activity of DRPY Type II Cas while targeting multiple loci simultaneously (TRAC, B2M, and PD1) in primary T cells after transient mRNA electroporation in combination with selected sgRNAs. Comparative data have been generated also for the benchmark nuclease SpCas9 using SUHYLRXVO\^SXEOLVKHG^VJ51$V^^'DWD^SUHVHQWHG^DV^PHDQ^^^6(0^IRU^Q^^^LQGHSHQGHQW^UXQV^ [0051] FIGS. 25A-25D show the genome-wide specificity profile of DRPY Type II Cas for selected guides targeting the B2M (FIG.25A) and PD1 (FIG.25B) loci in HEK293T cells after transient plasmid transfection. Comparative data have been generated also for the benchmark nuclease SpCas9 using previously published sgRNAs targeting B2M (FIG.25C) and PD1 (FIG.25D). Figure discloses SEQ ID NOS 578-581, respectively, in order of appearance. 6. DETAILED DESCRIPTION [0052] In one aspect, the disclosure provides Type II Cas proteins (e.g., DGHJ Type II Cas proteins, CUAZ Type II Cas proteins, AHWY Type II Cas proteins, CBGI Type II Cas proteins, ASDR Type II Cas proteins, BCZZ Type II Cas proteins, DRCY Type II Cas proteins, DRPY Type II Cas proteins, EAJR Type II Cas proteins, and CFDE Type II Cas proteins). Type II Cas proteins of the disclosure can be in the form of fusion proteins. Unless required otherwise by context, disclosures relating to Type II Cas proteins encompass Type II Cas proteins which are not fusion proteins and Type II Cas proteins which are in the form of fusion proteins (e.g., Type II Cas protein comprising one or more nuclear localization signals and/or one or more tags). [0053] In some embodiments, a Type II Cas protein of the disclosure comprises an amino acid sequence having at least 50% (e.g., at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 85%, at least 90%, at least 95%, or more) sequence identity to a RuvC-I domain, RuvC-II domain, RuvC-III domain, BH domain, REC1_2 domain, REC2 domain, REC1_2 domain, HNH domain, Topo domain or CTD of a DGHJ Type II Cas protein, CUAZ Type II Cas protein, AHWY Type II Cas protein, CBGI Type II Cas protein, ASDR Type II Cas protein, BCZZ Type II Cas protein, DRCY Type II Cas protein, DRPY Type II Cas protein, EAJR Type II Cas protein, or CFDE Type II Cas protein. In some embodiments, a Type II Cas protein of the disclosure is a chimeric Type II Cas protein, for example, comprising one or more domains from a DGHJ Type II Cas protein and/or CUAZ Type II Cas protein and/or AHWY Type II Cas protein and/or CBGI Type II Cas protein and/or ASDR Type II Cas protein and/or BCZZ Type II Cas protein and/or DRCY Type II Cas protein and/or DRPY Type II Cas protein and/or EAJR Type II Cas protein and/or CFDE Type II Cas protein, and one or more domains from a different Type II Cas protein such as SpCas9. [0054] Exemplary features of Type II Cas proteins of the disclosure are described in Section 6.2. [0055] In another aspect, the disclosure provides guide (gRNA) molecules, for example single guide RNAs (sgRNAs). Exemplary features of the gRNAs of the disclosure are described in Section 6.3. [0056] In further aspects, the disclosure provides systems comprising a Type II Cas protein of the disclosure and one or more gRNAs, e.g., sgRNAs. Exemplary features of systems are described in Section 6.4. [0057] In further aspects, the disclosure provides nucleic acids and pluralities of nucleic acids encoding a Type II Cas protein of the disclosure and, optionally, a guide RNA, for example a sgRNA, and provides nucleic acids encoding a gRNA, for example a sgRNA, of the disclosure and, optionally, a Type II Cas protein. Exemplary features of nucleic and pluralities of nucleic acids of the disclosure are described in Section 6.5. [0058] In further aspects, the disclosure provides particles comprising the Type II Cas proteins, gRNAs, nucleic acids, and systems of the disclosure. Exemplary features of particles of the disclosure are described in Section 6.6. [0059] In another aspect, the disclosure provides cells and populations of cells containing or contacted with a Type II Cas protein, gRNA, nucleic acid, plurality of nucleic acids, system, or particle of the disclosure. Exemplary features of such cells and cell populations are described in Section 6.6. [0060] In another aspect, the disclosure provides pharmaceutical compositions comprising a Type II Cas protein, gRNA, nucleic acid, plurality of nucleic acids, system, particle, cell, or population of cells together with one or more excipients. Exemplary features of pharmaceutical compositions are described in Section 6.7. [0061] In another aspect, the disclosure provides methods of altering cells (e.g., editing the genome of a cell) using the Type II Cas proteins, gRNAs, nucleic acids, systems, particles, and pharmaceutical compositions of the disclosure. Features of exemplary methods of altering cells are described in Section 6.8. [0062] Those skilled in the relevant art will recognize and appreciate that many changes can be made to the various embodiments described herein, while still obtaining the beneficial results of the present disclosure. It will also be apparent that some of the desired benefits of the present disclosure can be obtained by selecting some of the features of the present disclosure without utilizing other features. Accordingly, those who work in the art will recognize that many modifications and adaptations to the present disclosure are possible and can even be desirable in certain circumstances and are a part of the present disclosure. Thus, the following description is provided as illustrative of the principles of the present disclosure and not in limitation thereof. 6.1. Definitions [0063] Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood to one of ordinary skill in the art to which this invention belongs. The following definitions are provided for the full understanding of terms used in this specification. [0064] $V^XVHG^LQ^WKH^VSHFLILFDWLRQ^DQG^FODLPV^^WKH^VLQJXODU^IRUP^³D^´^³DQ^´^DQG^³WKH´^LQFOXGH^SOXUDO^ references unless the context clearO\^GLFWDWHV^RWKHUZLVH^^)RU^H[DPSOH^^WKH^WHUP^³DQ^DJHQW´^LQFOXGHV^D^ plurality of agents, including mixtures thereof. [0065] 8QOHVV^LQGLFDWHG^RWKHUZLVH^^DQ^³RU´^FRQMXQFWLRQ^LV^LQWHQGHG^WR^EH^XVHG^LQ^LWV^FRUUHFW^VHQVH^DV^D^ Boolean logical operator, encompassing both the selection of features in the alternative (A or B, where the selection of A is mutually exclusive from B) and the selection of features in conjunction (A or B, where ERWK^$^DQG^%^DUH^VHOHFWHG^^^,Q^VRPH^SODFHV^LQ^WKH^WH[W^^WKH^WHUP^³DQG^RU´^LV^XVHG^IRU^the same purpose, ZKLFK^VKDOO^QRW^EH^FRQVWUXHG^WR^LPSO\^WKDW^³RU´^LV^XVHG^ZLWK^UHIHUHQFH^WR^PXWXDOO\^H[FOXVLYH^DOWHUQDWLYHV^ [0066] A Type II Cas protein refers to a wild-type or engineered Type II Cas protein. Engineered Type II Cas proteins can also be referred to as Type II Cas variants. For the avoidance of doubt, any GLVFORVXUH^SHUWDLQLQJ^WR^D^³7\SH^,,^&DV´^RU^³7\SH^,,^&DV^SURWHLQ´^SHUWDLQV^WR^ZLOG-type Type II Cas proteins and Type II Cas variants, unless the context dictates otherwise. A Type II Cas protein can have nuclease activity or be catalytically inactive (e.g., as in a dCas). [0067] As used herein, the percentage identity between two nucleotide sequences or between two amino acid sequences is calculated by multiplying the number of matches between a pair of aligned sequences by 100, and dividing by the length of the aligned region. Identity scoring only counts perfect matches and does not consider the degree of similarity of amino acids to one another, nor does it consider substitutions or deletions as matches. Alignment for purposes of determining percent sequence identity can be achieved in various ways that are within the skill in the art, for instance, by manual alignment or using publicly available computer software such as BLAST, BLAST-2, ALIGN, ALIGN-2 or Megalign (DNASTAR) software. Those skilled in the art can determine appropriate parameters for achieving maximum alignment. [0068] Guide RNA molecule (gRNA) refers to an RNA capable of forming a complex with a Type II Cas protein and which can direct the Type II Cas protein to a target DNA. gRNAs typically comprise a spacer of 15 to 30 nucleotides in length. gRNAs of the disclosure are in some embodiments single guide RNAs (sgRNAs), which typically FRPSULVH^D^VSDFHU^DW^WKH^^¶^HQG^RI^WKH^PROHFXOH^DQG^D^^¶^VJ51$^VFDIIROG^^ Various non-OLPLWLQJ^H[DPSOHV^RI^^¶^VJ51$^VFDIIROGV^DUH^GHVFULEHG^LQ^6HFWLRQ^6.3. [0069] $Q^VJ51$^FDQ^LQ^VRPH^HPERGLPHQWV^FRPSULVH^QR^XUDFLO^EDVH^DW^WKH^^¶^HQG^RI^WKH^VJ51$^ sequence. AlternaWLYHO\^^D^VJ51$^FDQ^FRPSULVH^RQH^RU^PRUH^XUDFLO^EDVHV^DW^WKH^^¶^HQG^RI^WKH^VJ51$^ VHTXHQFH^^)RU^H[DPSOH^^D^VJ51$^FDQ^FRPSULVH^^^XUDFLO^^8^^DW^WKH^^¶^HQG^RI^WKH^VJ51$^VHTXHQFH^^^^ XUDFLO^^88^^DW^WKH^^¶^HQG^RI^WKH^VJ51$^VHTXHQFH^^^^XUDFLO^^888^^DW^WKH^^¶^HQG^Rf the sgRNA sequence, 4 XUDFLO^^8888^^DW^WKH^^¶^HQG^RI^WKH^VJ51$^VHTXHQFH^^^^XUDFLO^^88888^^DW^WKH^^¶^HQG^RI^WKH^VJ51$^ VHTXHQFH^^^^XUDFLO^^888888^^DW^WKH^^¶^HQG^RI^WKH^VJ51$^VHTXHQFH^^^^XUDFLO^^8888888^^DW^WKH^^¶^HQG^ of the sgRNA sequence, or 8 uracil (UUU88888^^DW^WKH^^¶^HQG^RI^WKH^VJ51$^VHTXHQFH^^'LIIHUHQW^OHQJWK^ VWUHWFKHV^RI^XUDFLO^FDQ^EH^DSSHQGHG^DW^WKH^^¶^HQG^RI^D^VJ51$^DV^WHUPLQDWRUV^^7KXV^^IRU^H[DPSOH^^WKH^^¶^ sgRNA scaffolds set forth in Section 6.3 can be modified by adding or removing one or more uracils at the end of the sequence. [0070] Peptide, protein, and polypeptide are used interchangeably to refer to a natural or synthetic molecule comprising two or more amino acids linked by the carboxyl group of one amino acid to the alpha amino group of another. The amino acids may be natural or synthetic, and can contain chemical modifications such as disulfide bridges, substitution of radioisotopes, phosphorylation, substrate chelation (e.g., chelation of iron or copper atoms), glycosylation, acetylation, formylation, amidation, biotinylation, and a wide range of other modifications. A polypeptide may be attached to other molecules, for instance molecules required for function. Examples of molecules which may be attached to a polypeptide include, without limitation, cofactors, polynucleotides, lipids, metal ions, phosphate, etc. Non-limiting examples of polypeptides include peptide fragments, denatured/unstructured polypeptides, polypeptides having quaternary or aggregated structures, etc. There is expressly no requirement that a polypeptide must contain an intended function; a polypeptide can be functional, non-functional, function for unexpected/unintended purposes, or have unknown function. A polypeptide is comprised of approximately twenty, standard naturally occurring amino acids, although natural and synthetic amino acids which are not members of the standard twenty amino acids may also be used. The standard twenty amino acids include alanine (Ala, A), arginine (Arg, R), asparagine (Asn, N), aspartic acid (Asp, D), cysteine (Cys, C), glutamine (Gln, Q), glutamic acid (Glu, E), glycine (Gly, G), histidine, (His, H), isoleucine (Ile, I), leucine (Leu, L), lysine (Lys, K), methionine (Met, M), phenylalanine (Phe, F), proline (Pro, P), serine (Ser, S), threonine (Thr, T), tryptophan (Trp, W), tyrosine (Tyr, Y), and valine (Val, V). 7KH^WHUPV^³SRO\SHSWLGH^VHTXHQFH´^RU^³DPLQR^DFLG^VHTXHQFH´^DUH^DQ^DOSKDEHWLFDO^UHSUHVHQWDWLRQ^RI^D^ polypeptide molecule. [0071] Polynucleotide and oligonucleotide are used interchangeably and refer to a polymeric form of nucleotides of any length, either deoxyribonucleotides or ribonucleotides, or analogs thereof. Polynucleotides may have any three-dimensional structure, and may perform any function, known or unknown. The following are non-limiting examples of polynucleotides: a gene or gene fragment, exons, introns, messenger RNA (mRNA), transfer RNA, ribosomal RNA, ribozymes, cDNA, recombinant polynucleotides, branched polynucleotides, plasmids, vectors, isolated DNA of any sequence, isolated RNA of any sequence, nucleic acid probes, primers and gRNAs. A polynucleotide may comprise modified nucleotides, such as methylated nucleotides and nucleotide analogs. If present, modifications to the nucleotide structure may be imparted before or after assembly of the polymer. The sequence of nucleotides may be interrupted by non-nucleotide components. A polynucleotide may be further modified after polymerization, such as by conjugation with a labeling component. A polynucleotide is composed of a specific sequence of four nucleotide bases: adenine (A); cytosine (C); guanine (G); thymine (T); and XUDFLO^^8^^IRU^WK\PLQH^^7^^ZKHQ^WKH^SRO\QXFOHRWLGH^LV^51$^^7KXV^^WKH^WHUP^³QXFOHRWLGH^VHTXHQFH´^LV^WKH^ alphabetical representation of a polynucleotide molecule. The letters used in polynucleotide sequences GHVFULEHG^KHUHLQ^FRUUHVSRQG^WR^,83$&^QRWDWLRQ^^)RU^H[DPSOH^^WKH^OHWWHU^³1´^LQ^D^QXFOHRWLGH^VHTXHQFH^ represents a nucleotide which can be A, T, C, or G in a DNA sequence, or A, U, C, or G in a RNA VHTXHQFH^^WKH^OHWWHU^³5´^LQ^D^QXFOHRWLGH^VHTXHQFH^UHSUHVHQWV^D^QXFOHRWLGH^ZKLFK^FDQ^EH^$^RU^*^^DQG^WKH^ OHWWHU^³9´^LQ^D^QXFOHRWLGH^VHTXHQFH^UHSUHVHQWV^D^QXFOHRWLGH^ZKLFK^FDQ^EH^³$^^&^^RU^*^ [0072] Protospacer adjacent motif (PAM) refers to a DNA sequence downstream (e.g., immediately downstream) of a target sequence on the non-target strand recognized by a Type II Cas protein. A PAM VHTXHQFH^LV^ORFDWHG^^¶^RI^WKH^WDUJHW^VHTXHQFH^RQ^WKH^QRQ-target strand. [0073] Spacer refers to a region of a gRNA molecule which is partially or fully complementary to a target sequence found in the + or - strand of genomic DNA. When complexed with a Type II Cas protein, the gRNA directs the Type II Cas to the target sequence in the genomic DNA. A spacer of a Type II Cas gRNA is typically 15 to 30 nucleotides in length (e.g., 20-25 nucleotides). The nucleotide sequence of a spacer can be, but is not necessarily, fully complementary to the target sequence. For example, a spacer can contain one or more mismatches with a target sequence, e.g., the spacer can comprise one, two, or three mismatches with the target sequence. 6.2. Type II Cas Proteins 6.2.1.1. DGHJ Type II Cas Proteins [0074] In one aspect, the disclosure provides DGHJ Type II Cas proteins. DGHJ Type II Cas proteins can be further classified as Type IIA Cas proteins. The DGHJ Type II Cas proteins typically comprise an amino acid sequence that is at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 85%, at least 90%, or at least 95% identical to SEQ ID NO:1. In some embodiments, the DGHJ Type II Cas proteins comprise an amino acid sequence that is at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical to SEQ ID NO:1. In some embodiments, a DGHJ Type II Cas protein comprises an amino acid sequence that is identical to SEQ ID NO:1. [0075] Exemplary DGHJ Type II Cas protein sequences and nucleotide sequences encoding exemplary DGHJ Type II Cas proteins are set forth in Table 1A. TABLE 1A DGHJ Type II Cas Sequences Name Sequence SEQ ID NO DGHJ Type II RKDADYNIGLDIGVASVGWCVTNDENNILKRNGKNMWGSRLFSEASTAK 1 Cas amino NTRMLRSSRRRLERRKNRILYLQQIFQDDMEKEYPNFFPMLKETANIPED acid KRITKLLDGKKYNLFSEQNFTDVQYYNQKTIYHLRKYLIESNKKADLRLVY sequence LALHHIIKYRGNFLYEGDFNNQVDEIENDILIVNNFVKENYFINSKIEEKEIIN (without N- ILKDKTKTKAEKKEILIKVFEFDSASKPVITNIVKAIIGYNFDVSKIFEIDYEK terminal GNISFAKEIENEEEIEELLGNQVEIYEAMKNIYSWFTLQDILKGNKYISDAFI methionine) EKYDKYKGDLILLKNIYKKYYSNEYKSFFKEENINNYVHYNGKQIRKLDKS CDNNEFFKKVSQKIKSLPEDYEKLYDEVNKKVKIENEKNGSNIELDDRYR DKENILQDIESGNFICKLNVTDNGAIPHQLHEIELKKIIENQKQYYEFLNEN AEKIIKIFETRIPYYVGPLIEGKKGSRWSWLIKNNEQEKAHIRPWNMEETIN KNETAEKFINELTNYCTYLQDEKVMPKQSILYSKYCVLNELNNIRINGKHIA KDFKQKIINEYFERKTKVTKKDIISFYDKEGINVSSFEGLSDVNNFNSNMK SYKDMIEILGKVDDSNIEMCEKIIYYVTIFEDKKILREKIKNEYSGINDEQLR RIMKLKYSGWSRLSRKFLTGIKSYNNETIMDRLMNTHENLMQIINNKELG FDKIIENNIPKKSGRITYEDVDEIQTSPANKRAIWQTICVVNEIKKIMKKEPK TABLE 1A DGHJ Type II Cas Sequences Name Sequence SEQ ID NO NIYIEFARSEDRNKRLKDSRLKNLLKIYEDIKEQIDEIKKFDPDVYKELKKH QDDKDVTEKMYLYFIQNGKCLYSGEKLDIDELDKYEVDHIIPRSYIKDDSI DNKALVKRRENQRKKDSLLLTEDVIRKNISWWKNLYDNNLISAKKYHNLT RRKMFETESEKNKFVQRQLVETRQITKYVTNLLKDGEENVEIYALRAELT HWFREKYEIYKNRNVNNCHHAQDAYIISTIGNIISKEWPKKEDFQYGQYV DNYLREAKAKNEKYGMVIGFISKRVEISKIKQAMNYKDYFISRMLEEQTG EFYNQTIQKAGQGKIKLKPNKDTNRYGGYTNENEAYSVIYKYIDKSGKQQ YKMIGIPIRIASDIKRKRTNLQEYITDVDFNNNDNIELQIIRNKIMKNQIYLDQ NDEPLMLCSSKEIRPAKELILNEEMSKLVYMMNSSENKLTDEEKEEVKRS YSKMYDYLMSKLEKEYKIHKNSYMRLINKKEEFETLEDEDKKKAINGIIDL MFKGQGNLKVLGMSDREGRMSGKTFKTDKLIHMTFVDKSVTGMYERRQ KINGMENNSSK DGHJ Type II MRKDADYNIGLDIGVASVGWCVTNDENNILKRNGKNMWGSRLFSEAST 2 Cas amino AKNTRMLRSSRRRLERRKNRILYLQQIFQDDMEKEYPNFFPMLKETANIP acid EDKRITKLLDGKKYNLFSEQNFTDVQYYNQKTIYHLRKYLIESNKKADLRL sequence VYLALHHIIKYRGNFLYEGDFNNQVDEIENDILIVNNFVKENYFINSKIEEKE IINILKDKTKTKAEKKEILIKVFEFDSASKPVITNIVKAIIGYNFDVSKIFEIDYE KGNISFAKEIENEEEIEELLGNQVEIYEAMKNIYSWFTLQDILKGNKYISDA FIEKYDKYKGDLILLKNIYKKYYSNEYKSFFKEENINNYVHYNGKQIRKLDK SCDNNEFFKKVSQKIKSLPEDYEKLYDEVNKKVKIENEKNGSNIELDDRY RDKENILQDIESGNFICKLNVTDNGAIPHQLHEIELKKIIENQKQYYEFLNE NAEKIIKIFETRIPYYVGPLIEGKKGSRWSWLIKNNEQEKAHIRPWNMEETI NKNETAEKFINELTNYCTYLQDEKVMPKQSILYSKYCVLNELNNIRINGKH IAKDFKQKIINEYFERKTKVTKKDIISFYDKEGINVSSFEGLSDVNNFNSNM KSYKDMIEILGKVDDSNIEMCEKIIYYVTIFEDKKILREKIKNEYSGINDEQL RRIMKLKYSGWSRLSRKFLTGIKSYNNETIMDRLMNTHENLMQIINNKEL GFDKIIENNIPKKSGRITYEDVDEIQTSPANKRAIWQTICVVNEIKKIMKKEP KNIYIEFARSEDRNKRLKDSRLKNLLKIYEDIKEQIDEIKKFDPDVYKELKK HQDDKDVTEKMYLYFIQNGKCLYSGEKLDIDELDKYEVDHIIPRSYIKDDS IDNKALVKRRENQRKKDSLLLTEDVIRKNISWWKNLYDNNLISAKKYHNLT RRKMFETESEKNKFVQRQLVETRQITKYVTNLLKDGEENVEIYALRAELT HWFREKYEIYKNRNVNNCHHAQDAYIISTIGNIISKEWPKKEDFQYGQYV DNYLREAKAKNEKYGMVIGFISKRVEISKIKQAMNYKDYFISRMLEEQTG EFYNQTIQKAGQGKIKLKPNKDTNRYGGYTNENEAYSVIYKYIDKSGKQQ YKMIGIPIRIASDIKRKRTNLQEYITDVDFNNNDNIELQIIRNKIMKNQIYLDQ NDEPLMLCSSKEIRPAKELILNEEMSKLVYMMNSSENKLTDEEKEEVKRS YSKMYDYLMSKLEKEYKIHKNSYMRLINKKEEFETLEDEDKKKAINGIIDL MFKGQGNLKVLGMSDREGRMSGKTFKTDKLIHMTFVDKSVTGMYERRQ KINGMENNSSK DGHJ Type II MGKPIPNPLLGLDSTKRTADGSEFESPKKKRKVGSRKDADYNIGLDIGVA 3 Cas SVGWCVTNDENNILKRNGKNMWGSRLFSEASTAKNTRMLRSSRRRLER mammalian RKNRILYLQQIFQDDMEKEYPNFFPMLKETANIPEDKRITKLLDGKKYNLF expression SEQNFTDVQYYNQFKTIYHLRKYLIESNKKADLRLVYLALHHIIKYRGNFL construct YEGDFNNQVDEIENDILIVNNFVKENYFINSKIEEKEIINILKDKTKTKAEKK (with N- EILIKVFEFDSASKPVITNIVKAIIGYNFDVSKIFEIDYEKGNISFAKEIENEEE terminal IEELLGNQVEIYEAMKNIYSWFTLQDILKGNKYISDAFIEKYDKYKGDLILLK methionine, NIYKKYYSNEYKSFFKEENINNYVHYNGKQIRKLDKSCDNNEFFKKVSQK stop codon, IKSLPEDYEKLYDEVNKKVKIENEKNGSNIELDDRYRDKENILQDIESGNFI V5 tag, N- CKLNVTDNGAIPHQLHEIELKKIIENQKQYYEFLNENAEKIIKIFETRIPYYV terminal NLS GPLIEGKKGSRWSWLIKNNEQEKAHIRPWNMEETINKNETAEKFINELTN and C- YCTYLQDEKVMPKQSILYSKYCVLNELNNIRINGKHIAKDFKQKIINEYFER terminal NLS) KTKVTKKDIISFYDKEGINVSSFEGLSDVNNFNSNMKSYKDMIEILGKVDD (aa) SNIEMCEKIIYYVTIFEDKKILREKIKNEYSGINDEQLRRIMKLKYSGWSRL SRKFLTGIKSYNNETIMDRLMNTHENLMQIINNKELGFDKIIENNIPKKSGR ITYEDVDEIQTSPANKRAIWQTICVVNEIKKIMKKEPKNIYIEFARSEDRNK RLKDSRLKNLLKIYEDIKEQIDEIKKFDPDVYKELKKHQDDKDVTEKMYLY FIQNGKCLYSGEKLDIDELDKYEVDHIIPRSYIKDDSIDNKALVKRRENQR TABLE 1A DGHJ Type II Cas Sequences Name Sequence SEQ ID NO KKDSLLLTEDVIRKNISWWKNLYDNNLISAKKYHNLTRRKMFETESEKNK FVQRQLVETRQITKYVTNLLKDGEENVEIYALRAELTHWFREKYEIYKNR NVNNCHHAQDAYIISTIGNIISKEWPKKEDFQYGQYVDNYLREAKAKNEK YGMVIGFISKRVEISKIKQAMNYKDYFISRMLEEQTGEFYNQTIQKAGQG KIKLKPNKDTNRYGGYTNENEAYSVIYKYIDKSGKQQYKMIGIPIRIASDIK RKRTNLQEYITDVDFNNNDNIELQIIRNKIMKNQIYLDQNDEPLMLCSSKEI RPAKELILNEEMSKLVYMMNSSENKLTDEEKEEVKRSYSKMYDYLMSKL EKEYKIHKNSYMRLINKKEEFETLEDEDKKKAINGIIDLMFKGQGNLKVLG MSDREGRMSGKTFKTDKLIHMTFVDKSVTGMYERRQKINGMENNSSKS RKRTADGSEFESPKKKRKV DGHJ Type II ATGAGAAAAGACGCAGATTATAATATAGGATTAGATATAGGAGTAGCT 4 Cas coding TCTGTTGGTTGGTGTGTAACAAATGATGAAAATAATATATTGAAAAGAA sequence (nt) ATGGAAAAAACATGTGGGGGTCACGATTGTTTAGTGAAGCATCAACA (not codon GCTAAAAATACGCGAATGTTAAGATCATCTAGAAGGAGACTAGAAAGA optimized) AGAAAAAATAGAATTTTATATTTGCAGCAAATATTTCAAGATGACATGG AAAAAGAATATCCTAATTTCTTTCCTATGCTTAAAGAAACTGCGAATAT CCCAGAAGACAAAAGAATAACAAAATTATTGGATGGAAAAAAATATAA CTTATTTTCAGAACAGAATTTTACAGATGTACAATATTATAATCAATTCA AAACTATTTATCATCTTAGAAAATATTTAATTGAATCAAACAAAAAAGC CGATTTAAGGCTTGTTTATCTTGCTCTTCACCATATCATAAAATACAGA GGAAATTTCTTATATGAGGGAGATTTTAACAATCAAGTAGACGAAATT GAAAATGATATTTTAATAGTTAATAACTTTGTAAAAGAGAACTATTTTAT AAATTCAAAAATAGAAGAAAAAGAAATAATAAATATTTTAAAAGATAAA ACTAAAACTAAGGCTGAAAAGAAAGAGATATTGATAAAAGTATTCGAA TTTGATAGTGCATCAAAACCTGTGATAACAAATATTGTAAAAGCTATTA TTGGCTATAATTTTGATGTAAGTAAAATATTTGAAATTGATTATGAGAA AGGAAATATTTCATTTGCAAAAGAAATAGAAAACGAAGAAGAAATTGA AGAACTATTAGGAAATCAAGTAGAAATCTATGAAGCAATGAAAAACAT ATATAGTTGGTTTACATTACAAGATATCTTAAAGGGAAATAAATATATA TCAGATGCATTTATAGAAAAATATGATAAATATAAAGGAGATCTAATAC TACTAAAAAATATATACAAAAAATATTATTCTAATGAATATAAATCTTTT TTCAAAGAAGAAAACATAAATAATTATGTACATTATAATGGAAAACAAA TAAGAAAATTAGACAAGTCTTGTGATAACAATGAATTTTTCAAAAAAGT ATCTCAAAAAATAAAAAGCCTTCCAGAGGACTATGAAAAATTATATGAT GAAGTAAATAAAAAAGTAAAAATAGAAAATGAAAAAAACGGAAGTAAT ATTGAGCTGGATGATAGATACAGGGATAAAGAAAATATTTTACAAGAC ATAGAAAGCGGAAACTTCATATGTAAATTAAATGTGACAGATAATGGT GCAATACCACATCAATTACATGAAATCGAATTAAAGAAAATTATTGAGA ATCAAAAACAATATTATGAATTTTTAAATGAAAATGCAGAAAAAATTATA AAAATATTTGAGACAAGAATCCCATATTATGTTGGACCATTAATTGAAG GAAAAAAAGGAAGCAGATGGTCATGGCTTATAAAGAATAATGAACAAG AAAAGGCACACATTAGACCTTGGAACATGGAAGAAACAATTAATAAAA ATGAAACAGCAGAGAAATTCATAAATGAATTAACAAACTATTGCACTTA TTTGCAAGATGAAAAAGTAATGCCAAAGCAATCAATCTTATATTCCAAA TATTGTGTATTGAATGAATTGAACAATATAAGAATAAATGGAAAGCATA TAGCGAAAGACTTTAAACAGAAGATAATTAACGAATATTTTGAGAGAA AAACCAAGGTTACTAAGAAAGACATAATAAGTTTTTATGATAAAGAGG GAATAAATGTTTCAAGCTTTGAAGGATTGTCAGATGTAAACAACTTTAA CTCAAATATGAAATCATATAAAGACATGATAGAGATATTGGGCAAAGT AGATGATAGTAACATAGAAATGTGCGAAAAAATAATATATTATGTAACA ATATTTGAAGATAAAAAAATACTAAGAGAAAAGATAAAAAATGAATATT CAGGAATAAATGATGAACAATTAAGAAGAATAATGAAGTTAAAATATTC TGGCTGGTCAAGACTTTCTAGAAAATTCTTGACAGGAATTAAATCATA TAACAATGAAACTATAATGGATAGACTAATGAATACTCATGAAAATCTT ATGCAAATAATAAACAATAAAGAACTTGGTTTTGATAAAATAATTGAAA ATAATATACCTAAAAAAAGCGGAAGGATTACTTATGAGGATGTTGATG AAATTCAGACGTCACCAGCAAATAAAAGAGCAATTTGGCAAACAATAT TABLE 1A DGHJ Type II Cas Sequences Name Sequence SEQ ID NO GTGTTGTTAATGAAATAAAAAAGATAATGAAAAAAGAACCTAAAAATAT TTATATAGAGTTTGCGAGAAGTGAGGATAGAAATAAAAGGCTAAAAGA TTCTAGATTAAAAAATCTTTTAAAAATTTATGAGGACATAAAAGAACAA ATAGACGAAATAAAGAAATTTGATCCAGATGTATATAAAGAGCTAAAG AAACATCAAGACGATAAAGATGTTACTGAAAAAATGTATTTGTATTTTA TTCAAAATGGAAAATGTTTATATAGTGGAGAAAAGTTGGATATAGACG AACTAGACAAGTATGAAGTTGACCATATCATACCTCGTTCTTATATAAA AGATGATAGCATCGATAATAAGGCTTTGGTTAAGAGAAGAGAAAATCA AAGGAAGAAAGATAGCTTACTTTTAACAGAAGATGTTATCAGAAAAAA TATTTCGTGGTGGAAAAACTTATATGACAATAATTTAATTAGTGCAAAA AAATATCATAATTTAACAAGAAGAAAAATGTTTGAAACTGAAAGTGAAA AAAACAAATTTGTTCAAAGACAGCTTGTCGAAACAAGGCAGATAACTA AGTATGTTACAAATTTACTAAAAGATGGGGAAGAAAATGTTGAAATTTA TGCATTAAGAGCAGAGTTAACACATTGGTTCAGAGAAAAATACGAAAT ATATAAAAATAGAAATGTAAATAATTGTCACCATGCTCAGGATGCATAT ATAATTTCGACTATTGGAAATATAATAAGTAAAGAATGGCCAAAAAAAG AAGATTTCCAATATGGACAATATGTGGATAATTATTTAAGAGAAGCAAA AGCTAAGAATGAAAAATATGGCATGGTTATAGGGTTCATTAGCAAAAG AGTTGAAATAAGCAAAATAAAGCAAGCTATGAACTATAAAGATTACTTC ATAAGTAGAATGTTAGAAGAGCAAACTGGAGAATTTTATAATCAAACA ATTCAAAAAGCTGGGCAAGGAAAAATCAAACTCAAACCTAATAAAGAT ACAAACAGATATGGGGGATATACCAACGAAAATGAGGCATATAGTGTT ATATACAAATATATTGACAAATCTGGAAAACAACAATATAAGATGATAG GAATTCCAATTAGAATTGCAAGTGATATTAAGAGAAAAAGGACTAATTT ACAAGAGTATATTACAGATGTGGATTTCAACAATAATGATAATATTGAA CTGCAAATTATAAGAAACAAAATAATGAAGAATCAAATATATCTTGACC AGAATGATGAGCCATTAATGCTATGTAGTTCAAAAGAAATAAGACCTG CAAAAGAACTGATATTAAATGAAGAAATGTCAAAACTAGTATATATGAT GAACAGTTCGGAAAATAAATTAACAGATGAAGAAAAAGAAGAAGTAAA AAGATCGTATTCAAAAATGTATGACTATTTAATGAGCAAACTAGAAAAA GAATATAAAATCCATAAAAACAGTTATATGAGACTAATAAACAAAAAAG AAGAATTTGAAACACTAGAAGACGAGGATAAGAAAAAAGCAATCAATG GGATTATTGATTTGATGTTCAAAGGACAAGGAAATTTAAAAGTTTTAG GGATGTCAGACAGAGAAGGAAGAATGAGTGGCAAAACATTCAAAACT GATAAATTAATACATATGACGTTTGTTGATAAATCGGTAACAGGAATGT ACGAAAGAAGGCAAAAGATAAATGGGATGGAGAACAATAGTAGTAAG TAA DGHJ Type II AGAAAAGATGCTGACTACAACATCGGTCTGGACATCGGCGTGGCCTC 5 Cas coding CGTGGGCTGGTGCGTGACCAACGACGAGAACAATATCCTGAAACGG sequence (nt) AACGGCAAGAACATGTGGGGCTCTCGGCTGTTCAGCGAAGCGTCTA (human CCGCCAAGAATACCAGAATGCTCAGATCCAGCAGACGTAGACTGGAA codon- AGACGGAAGAACAGAATCCTGTACCTGCAGCAGATCTTTCAGGACGA optimized; CATGGAAAAGGAGTACCCCAACTTCTTTCCAATGCTGAAGGAAACCG lacking N- CCAACATCCCAGAGGACAAGAGAATCACAAAGCTCCTGGACGGCAAA terminal AAGTACAACCTGTTCAGCGAGCAGAACTTCACCGATGTCCAGTATTAC methionine; AACCAGTTCAAGACCATCTACCACCTGCGGAAGTACCTGATCGAGAG no stop CAACAAGAAGGCCGATCTGAGACTGGTTTACCTGGCCCTCCATCACA codon) TCATCAAGTACAGAGGCAACTTCCTGTACGAGGGCGACTTCAACAAC CAAGTGGACGAAATCGAGAATGACATCCTGATCGTGAACAACTTCGT GAAAGAGAACTACTTCATCAACTCTAAGATCGAGGAAAAGGAAATCAT CAATATCCTGAAGGACAAGACCAAAACCAAAGCTGAGAAAAAGGAAA TCCTGATCAAGGTGTTCGAGTTTGACAGCGCCTCTAAACCTGTGATCA CCAACATCGTGAAGGCTATCATTGGCTACAACTTCGACGTGTCCAAG ATCTTCGAGATCGACTACGAAAAAGGCAATATCAGCTTCGCCAAAGAA ATCGAAAATGAAGAAGAGATCGAAGAGCTGCTTGGCAACCAGGTGGA AATCTATGAAGCCATGAAGAACATCTACTCCTGGTTCACCCTGCAGGA CATCCTGAAGGGCAACAAGTACATCAGCGACGCCTTCATAGAGAAGT TABLE 1A DGHJ Type II Cas Sequences Name Sequence SEQ ID NO ACGACAAGTACAAGGGCGATCTGATCCTGCTGAAGAACATCTATAAA AAGTACTACTCTAACGAGTACAAGAGCTTTTTTAAGGAAGAAAACATT AACAATTACGTGCACTACAATGGAAAGCAGATCAGAAAGCTGGACAA GTCATGCGACAACAATGAGTTCTTCAAGAAGGTTTCTCAGAAAATCAA GAGTCTGCCTGAGGACTACGAGAAGCTGTACGACGAGGTGAACAAG AAGGTGAAGATCGAGAATGAGAAGAACGGCAGCAACATCGAGCTGG ACGACAGATACAGAGACAAGGAAAACATCCTGCAGGACATTGAGAGC GGCAACTTTATTTGTAAACTGAACGTGACAGACAACGGCGCCATCCC CCACCAGCTGCACGAGATCGAGCTGAAGAAGATCATCGAGAACCAAA AGCAGTACTACGAGTTCCTGAACGAGAATGCCGAGAAGATTATTAAG ATATTCGAAACCAGAATCCCTTACTATGTGGGACCTCTGATCGAGGG AAAAAAGGGCAGCAGATGGAGCTGGCTGATCAAAAACAACGAGCAAG AGAAGGCCCACATCAGACCTTGGAACATGGAAGAAACCATCAATAAA AACGAAACAGCTGAAAAGTTCATCAACGAATTAACTAACTACTGCACC TACCTGCAAGACGAAAAGGTGATGCCCAAACAGAGCATCCTGTACAG CAAGTATTGTGTGCTGAACGAGCTGAACAACATTAGAATCAACGGAAA GCACATCGCCAAGGATTTCAAGCAGAAGATAATCAACGAATACTTCGA AAGAAAGACCAAGGTCACCAAGAAGGACATTATCAGCTTTTACGACAA GGAAGGCATCAACGTGTCCTCTTTCGAAGGCCTGAGCGACGTGAACA ACTTCAACTCAAATATGAAGTCCTACAAGGATATGATAGAGATTCTGG GAAAGGTGGACGATAGCAACATCGAGATGTGCGAGAAAATCATCTAC TACGTGACAATCTTCGAGGATAAAAAGATTCTGAGGGAAAAGATCAAG AATGAGTATAGCGGCATCAACGACGAACAGTTGAGAAGAATCATGAA ACTTAAGTACAGCGGGTGGTCCAGACTGAGCAGAAAATTCCTGACCG GCATCAAGTCTTACAACAATGAAACGATTATGGATCGACTGATGAACA CCCATGAGAACCTGATGCAGATCATCAACAACAAGGAGTTAGGATTT GATAAGATCATCGAGAACAACATCCCAAAGAAGAGCGGCAGAATCAC CTACGAGGACGTGGACGAGATTCAGACCAGCCCTGCTAACAAAAGG GCCATCTGGCAGACCATCTGCGTGGTGAACGAGATCAAGAAAATCAT GAAAAAGGAGCCTAAGAACATCTACATCGAGTTCGCCAGAAGCGAGG ATAGAAACAAGAGACTTAAAGATTCCAGACTGAAGAACCTGCTGAAGA TCTACGAGGACATCAAGGAGCAGATCGACGAGATCAAGAAATTCGAT CCTGACGTGTACAAAGAACTGAAGAAGCACCAGGATGATAAGGACGT GACCGAGAAGATGTACCTGTATTTCATCCAGAACGGCAAGTGTCTGT ACAGCGGAGAGAAGCTGGACATCGACGAGCTGGACAAGTACGAGGT GGACCACATAATACCCAGATCATACATCAAGGACGACAGCATCGATA ACAAGGCCCTGGTGAAGCGGAGAGAGAATCAGAGAAAGAAGGATTC TCTGCTGCTGACCGAGGACGTGATCAGGAAGAACATCAGCTGGTGG AAGAATCTGTATGACAACAACCTGATAAGCGCCAAGAAGTACCACAA CCTGACCAGACGGAAAATGTTCGAGACAGAGAGCGAGAAAAACAAAT TCGTGCAGCGGCAGCTGGTCGAGACAAGACAGATCACAAAGTACGT GACAAACCTGCTGAAGGATGGCGAAGAGAACGTGGAAATCTACGCC CTGCGGGCCGAGCTGACCCACTGGTTTAGAGAGAAATACGAGATATA TAAGAATCGGAACGTGAACAACTGCCACCACGCCCAGGACGCCTACA TCATCAGCACAATCGGCAATATCATCAGCAAGGAGTGGCCTAAGAAG GAGGATTTCCAGTACGGGCAGTACGTGGACAACTACCTGAGAGAGG CCAAGGCCAAGAACGAGAAGTACGGCATGGTGATCGGCTTCATCTCT AAAAGAGTGGAAATCAGCAAGATCAAACAGGCCATGAACTACAAGGA TTACTTTATCAGCAGAATGCTGGAAGAGCAAACCGGCGAGTTCTACA ACCAGACAATCCAAAAGGCCGGCCAGGGAAAGATCAAGCTGAAACCT AACAAGGACACCAATAGATACGGCGGATACACCAACGAAAACGAAGC ATATTCTGTGATCTACAAGTACATCGACAAGAGCGGCAAACAGCAGTA TAAGATGATCGGCATCCCTATCAGAATCGCCTCTGACATCAAGAGAAA GCGGACAAATCTGCAGGAGTACATCACAGATGTGGATTTCAATAACA ACGACAACATAGAGCTGCAGATCATCCGCAACAAAATCATGAAAAAC CAGATCTACCTGGATCAGAACGACGAACCCCTGATGCTGTGCAGCAG CAAGGAAATCAGACCCGCTAAAGAACTGATCCTGAATGAAGAGATGA TABLE 1A DGHJ Type II Cas Sequences Name Sequence SEQ ID NO GCAAGCTCGTATACATGATGAACAGCAGCGAGAACAAGCTGACAGAT GAAGAAAAGGAAGAGGTGAAGCGGAGCTATAGCAAGATGTACGACTA CCTGATGTCCAAGCTGGAGAAGGAGTACAAAATCCACAAGAACTCCT ACATGAGACTGATCAACAAAAAGGAGGAATTTGAGACACTGGAGGAC GAGGATAAGAAAAAGGCCATCAACGGCATCATCGACCTGATGTTCAA GGGCCAGGGCAACCTGAAGGTCCTGGGCATGAGCGATCGGGAAGG CAGGATGAGTGGAAAGACCTTCAAGACCGACAAGCTGATCCACATGA CATTCGTGGACAAAAGCGTCACCGGCATGTACGAGCGACGGCAGAA GATCAACGGCATGGAAAATAACAGCTCCAAG DGHJ Type II ATGGGCAAACCTATCCCCAATCCCCTGCTGGGCCTGGACAGCACCAA 6 Cas GCGGACTGCCGACGGCAGTGAGTTCGAGTCTCCCAAGAAAAAGAGG mammalian AAAGTGggatccAGAAAAGATGCTGACTACAACATCGGTCTGGACATCG expression GCGTGGCCTCCGTGGGCTGGTGCGTGACCAACGACGAGAACAATAT construct CCTGAAACGGAACGGCAAGAACATGTGGGGCTCTCGGCTGTTCAGC (with N- GAAGCGTCTACCGCCAAGAATACCAGAATGCTCAGATCCAGCAGACG terminal TAGACTGGAAAGACGGAAGAACAGAATCCTGTACCTGCAGCAGATCT methionine, TTCAGGACGACATGGAAAAGGAGTACCCCAACTTCTTTCCAATGCTG stop codon, AAGGAAACCGCCAACATCCCAGAGGACAAGAGAATCACAAAGCTCCT V5 tag, N- GGACGGCAAAAAGTACAACCTGTTCAGCGAGCAGAACTTCACCGATG terminal NLS TCCAGTATTACAACCAGTTCAAGACCATCTACCACCTGCGGAAGTACC and C- TGATCGAGAGCAACAAGAAGGCCGATCTGAGACTGGTTTACCTGGCC terminal NLS) CTCCATCACATCATCAAGTACAGAGGCAACTTCCTGTACGAGGGCGA (nt) CTTCAACAACCAAGTGGACGAAATCGAGAATGACATCCTGATCGTGA ACAACTTCGTGAAAGAGAACTACTTCATCAACTCTAAGATCGAGGAAA AGGAAATCATCAATATCCTGAAGGACAAGACCAAAACCAAAGCTGAG AAAAAGGAAATCCTGATCAAGGTGTTCGAGTTTGACAGCGCCTCTAAA CCTGTGATCACCAACATCGTGAAGGCTATCATTGGCTACAACTTCGAC GTGTCCAAGATCTTCGAGATCGACTACGAAAAAGGCAATATCAGCTTC GCCAAAGAAATCGAAAATGAAGAAGAGATCGAAGAGCTGCTTGGCAA CCAGGTGGAAATCTATGAAGCCATGAAGAACATCTACTCCTGGTTCA CCCTGCAGGACATCCTGAAGGGCAACAAGTACATCAGCGACGCCTTC ATAGAGAAGTACGACAAGTACAAGGGCGATCTGATCCTGCTGAAGAA CATCTATAAAAAGTACTACTCTAACGAGTACAAGAGCTTTTTTAAGGAA GAAAACATTAACAATTACGTGCACTACAATGGAAAGCAGATCAGAAAG CTGGACAAGTCATGCGACAACAATGAGTTCTTCAAGAAGGTTTCTCAG AAAATCAAGAGTCTGCCTGAGGACTACGAGAAGCTGTACGACGAGGT GAACAAGAAGGTGAAGATCGAGAATGAGAAGAACGGCAGCAACATC GAGCTGGACGACAGATACAGAGACAAGGAAAACATCCTGCAGGACAT TGAGAGCGGCAACTTTATTTGTAAACTGAACGTGACAGACAACGGCG CCATCCCCCACCAGCTGCACGAGATCGAGCTGAAGAAGATCATCGAG AACCAAAAGCAGTACTACGAGTTCCTGAACGAGAATGCCGAGAAGAT TATTAAGATATTCGAAACCAGAATCCCTTACTATGTGGGACCTCTGAT CGAGGGAAAAAAGGGCAGCAGATGGAGCTGGCTGATCAAAAACAAC GAGCAAGAGAAGGCCCACATCAGACCTTGGAACATGGAAGAAACCAT CAATAAAAACGAAACAGCTGAAAAGTTCATCAACGAATTAACTAACTA CTGCACCTACCTGCAAGACGAAAAGGTGATGCCCAAACAGAGCATCC TGTACAGCAAGTATTGTGTGCTGAACGAGCTGAACAACATTAGAATCA ACGGAAAGCACATCGCCAAGGATTTCAAGCAGAAGATAATCAACGAA TACTTCGAAAGAAAGACCAAGGTCACCAAGAAGGACATTATCAGCTTT TACGACAAGGAAGGCATCAACGTGTCCTCTTTCGAAGGCCTGAGCGA CGTGAACAACTTCAACTCAAATATGAAGTCCTACAAGGATATGATAGA GATTCTGGGAAAGGTGGACGATAGCAACATCGAGATGTGCGAGAAAA TCATCTACTACGTGACAATCTTCGAGGATAAAAAGATTCTGAGGGAAA AGATCAAGAATGAGTATAGCGGCATCAACGACGAACAGTTGAGAAGA ATCATGAAACTTAAGTACAGCGGGTGGTCCAGACTGAGCAGAAAATT CCTGACCGGCATCAAGTCTTACAACAATGAAACGATTATGGATCGACT GATGAACACCCATGAGAACCTGATGCAGATCATCAACAACAAGGAGT TABLE 1A DGHJ Type II Cas Sequences Name Sequence SEQ ID NO TAGGATTTGATAAGATCATCGAGAACAACATCCCAAAGAAGAGCGGC AGAATCACCTACGAGGACGTGGACGAGATTCAGACCAGCCCTGCTAA CAAAAGGGCCATCTGGCAGACCATCTGCGTGGTGAACGAGATCAAGA AAATCATGAAAAAGGAGCCTAAGAACATCTACATCGAGTTCGCCAGAA GCGAGGATAGAAACAAGAGACTTAAAGATTCCAGACTGAAGAACCTG CTGAAGATCTACGAGGACATCAAGGAGCAGATCGACGAGATCAAGAA ATTCGATCCTGACGTGTACAAAGAACTGAAGAAGCACCAGGATGATA AGGACGTGACCGAGAAGATGTACCTGTATTTCATCCAGAACGGCAAG TGTCTGTACAGCGGAGAGAAGCTGGACATCGACGAGCTGGACAAGT ACGAGGTGGACCACATAATACCCAGATCATACATCAAGGACGACAGC ATCGATAACAAGGCCCTGGTGAAGCGGAGAGAGAATCAGAGAAAGAA GGATTCTCTGCTGCTGACCGAGGACGTGATCAGGAAGAACATCAGCT GGTGGAAGAATCTGTATGACAACAACCTGATAAGCGCCAAGAAGTAC CACAACCTGACCAGACGGAAAATGTTCGAGACAGAGAGCGAGAAAAA CAAATTCGTGCAGCGGCAGCTGGTCGAGACAAGACAGATCACAAAGT ACGTGACAAACCTGCTGAAGGATGGCGAAGAGAACGTGGAAATCTAC GCCCTGCGGGCCGAGCTGACCCACTGGTTTAGAGAGAAATACGAGA TATATAAGAATCGGAACGTGAACAACTGCCACCACGCCCAGGACGCC TACATCATCAGCACAATCGGCAATATCATCAGCAAGGAGTGGCCTAA GAAGGAGGATTTCCAGTACGGGCAGTACGTGGACAACTACCTGAGA GAGGCCAAGGCCAAGAACGAGAAGTACGGCATGGTGATCGGCTTCA TCTCTAAAAGAGTGGAAATCAGCAAGATCAAACAGGCCATGAACTACA AGGATTACTTTATCAGCAGAATGCTGGAAGAGCAAACCGGCGAGTTC TACAACCAGACAATCCAAAAGGCCGGCCAGGGAAAGATCAAGCTGAA ACCTAACAAGGACACCAATAGATACGGCGGATACACCAACGAAAACG AAGCATATTCTGTGATCTACAAGTACATCGACAAGAGCGGCAAACAG CAGTATAAGATGATCGGCATCCCTATCAGAATCGCCTCTGACATCAAG AGAAAGCGGACAAATCTGCAGGAGTACATCACAGATGTGGATTTCAA TAACAACGACAACATAGAGCTGCAGATCATCCGCAACAAAATCATGAA AAACCAGATCTACCTGGATCAGAACGACGAACCCCTGATGCTGTGCA GCAGCAAGGAAATCAGACCCGCTAAAGAACTGATCCTGAATGAAGAG ATGAGCAAGCTCGTATACATGATGAACAGCAGCGAGAACAAGCTGAC AGATGAAGAAAAGGAAGAGGTGAAGCGGAGCTATAGCAAGATGTACG ACTACCTGATGTCCAAGCTGGAGAAGGAGTACAAAATCCACAAGAAC TCCTACATGAGACTGATCAACAAAAAGGAGGAATTTGAGACACTGGA GGACGAGGATAAGAAAAAGGCCATCAACGGCATCATCGACCTGATGT TCAAGGGCCAGGGCAACCTGAAGGTCCTGGGCATGAGCGATCGGGA AGGCAGGATGAGTGGAAAGACCTTCAAGACCGACAAGCTGATCCACA TGACATTCGTGGACAAAAGCGTCACCGGCATGTACGAGCGACGGCA GAAGATCAACGGCATGGAAAATAACAGCTCCAAGtctagaAAGCGGACA GCAGACGGCTCCGAATTTGAAAGCCCTAAGAAAAAGAGAAAGGTGTG A [0076] In some embodiments a DGHJ Type II Cas protein comprises an amino acid sequence of SEQ ID NO:1, SEQ ID NO:2, or SEQ ID NO:3. In some embodiments, a DGHJ Type II Cas protein has nickase activity, for example resulting from one or more amino acid substitutions relative to the sequence of SEQ ID NO:1, SEQ ID NO:2, or SEQ ID NO:3. In some embodiments, the one or more amino acid substitutions providing nickase activity comprise a D12A substitution, wherein the position of the D12A substitution is defined with respect to the amino acid numbering of SEQ ID NO:2. In some embodiments, the one or more amino acid substitutions providing nickase activity comprise an H868A substitution, wherein the position of the H868A substitution is defined with respect to the amino acid numbering of SEQ ID NO:2. In some embodiments, a DGHJ Type II Cas protein is catalytically inactive, for example due to a D12A substitution in combination with an H868A substitution. 6.2.1.2. CUAZ Type II Cas Proteins [0077] In one aspect, the disclosure provides CUAZ Type II Cas proteins. CUAZ Type II Cas proteins can be further classified as Type IIA Cas proteins. The CUAZ Type II Cas proteins typically comprise an amino acid sequence that is at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 85%, at least 90%, or at least 95% identical to SEQ ID NO:7. In some embodiments, the CUAZ Type II Cas proteins comprise an amino acid sequence that is at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical to SEQ ID NO:7. In some embodiments, a CUAZ Type II Cas protein comprises an amino acid sequence that is identical to SEQ ID NO:7. [0078] Exemplary CUAZ Type II Cas protein sequences and nucleotide sequences encoding exemplary CUAZ Type II Cas proteins are set forth in Table 1B. TABLE 1B CUAZ Type II Cas Sequences Name Sequence SEQ ID NO CUAZ Type II KTNKKIYKDNQYTLSIDPGLNNVGWALFDAKNKKMVEMGVRKFEPAKEA 7 Cas amino KISRLKRSARRNNGRKKWRKQQLKQAFIDFNLIGEDEFKNSQFYEFHPE acid QYKAAGVKNLMDLRCYALQNKVTYQELFQCIYNILKYRGNFLRDDIDFEN sequence GESLTEELLIQEITNFMIKSLQLDIDNLDKLKESISTMCNQNIDLKEIPYTEIS (without N- DDKVSQKSIENVFKALKDQKFNPFMIEDDPILIDEEEGKSIIPEAKTKTIYM terminal KSVQKCGDLSNRFEQLLNITDSYRLYKVLNGEKYICTAYKKKLDRVADYY methionine) SFKDSPEGCSKYSDIESAKIEEEITYITNNMNATKGKPEKYGYTRKNPRG PIHVIHNVLNRYPNGLLVKEAVDILRTQQNYYGKEVISDEFIEVIRYILKGRI PYYIGPLSETAKNQWCTKTGKFKYSYNYTIEHMDGVDEERSIKDWKDRM RGHCTYLPEAFALPKGSFVAETLLILNELNVLTAIDKNNNTYYFTTNDKIKV FDYLFLDENHYGDVKYKEICDLLDLAYFGPKNQKSQVKTFKQNYTLYKSI TSICTALKLHSIIDLFSTKFQENQLKTIEEIVLNLNLYDEKKHKYQALTKMGI SDENSKKLSALNSKKFYGLSEKFVLKTSLNKDGRCLLEILFEDNQPGYTN EMMTAITNACDMDGNPVSFIANKYEAAFKELEKNGELELNFNSLLEIMNK TGNGFQIPASRPVLRSINQMMKVIQAVYTYYGVPKRIVLEIPRDMSGKNT NHEMPAKHYKTMEALHQDIMKQCKDKDHPRKTCIDQKWNDLKDFELNQ KNRIKLELYLRQNGMDMITGEKINLNELENYEIDHILPRGFGDDSMDDKILI RFVSNALKKDRTPLMWITDPTDPVINKSGKEITADQYINRVNELFDMKLIS ENKKKRLLLKDTEDLDRFINQNLVDTRYITREMIGILKAYNRVKGYPTKVIA LKPAFTKNYSNAFDIYKNRDFGLQHHALDAAVLGITDIMLNRLFPNYESG RISTSFAKHHQMLSEVGNDKKLKDSTKYVIRKMYEKVFGNSPLDYGSLV YVCKNTMPLYSQKSNRNWKGEKFPASRRLPYQKYDIKNGNIVGVYDDK SEENIGKQNSEVLRLFEINNKNHAFDSVNCVAYDIYRIPEKKNNKVIWKNI GIHIPKCIVDEDGTIQKDKYIKLIKKYYGFDDNLIDEKGDLITAYFKLRLYKN DLVGNTEKHEIRDLVLGSIADKKLELNKIDCFVYQKIAQKKASLFKMIIDNQ GSSKYDIHNYKEYQECECKEEILRDMIFDDMNIKNQEFASLLYAKIINDKK LDLENINKICWILAYYQQFLNEITGKKEFNTGRGIGALKKDESINYFKLKTS PLGVRFNTEKNQWTGPVGYENAFKIIKKEAFTWKITV CUAZ Type II MKTNKKIYKDNQYTLSIDPGLNNVGWALFDAKNKKMVEMGVRKFEPAKE 8 Cas amino AKISRLKRSARRNNGRKKWRKQQLKQAFIDFNLIGEDEFKNSQFYEFHP acid EQYKAAGVKNLMDLRCYALQNKVTYQELFQCIYNILKYRGNFLRDDIDFE sequence NGESLTEELLIQEITNFMIKSLQLDIDNLDKLKESISTMCNQNIDLKEIPYTEI SDDKVSQKSIENVFKALKDQKFNPFMIEDDPILIDEEEGKSIIPEAKTKTIY MKSVQKCGDLSNRFEQLLNITDSYRLYKVLNGEKYICTAYKKKLDRVADY YSFKDSPEGCSKYSDIESAKIEEEITYITNNMNATKGKPEKYGYTRKNPR GPIHVIHNVLNRYPNGLLVKEAVDILRTQQNYYGKEVISDEFIEVIRYILKG RIPYYIGPLSETAKNQWCTKTGKFKYSYNYTIEHMDGVDEERSIKDWKD TABLE 1B CUAZ Type II Cas Sequences Name Sequence SEQ ID NO RMRGHCTYLPEAFALPKGSFVAETLLILNELNVLTAIDKNNNTYYFTTNDK IKVFDYLFLDENHYGDVKYKEICDLLDLAYFGPKNQKSQVKTFKQNYTLY KSITSICTALKLHSIIDLFSTKFQENQLKTIEEIVLNLNLYDEKKHKYQALTK MGISDENSKKLSALNSKKFYGLSEKFVLKTSLNKDGRCLLEILFEDNQPG YTNEMMTAITNACDMDGNPVSFIANKYEAAFKELEKNGELELNFNSLLEI MNKTGNGFQIPASRPVLRSINQMMKVIQAVYTYYGVPKRIVLEIPRDMSG KNTNHEMPAKHYKTMEALHQDIMKQCKDKDHPRKTCIDQKWNDLKDFE LNQKNRIKLELYLRQNGMDMITGEKINLNELENYEIDHILPRGFGDDSMD DKILIRFVSNALKKDRTPLMWITDPTDPVINKSGKEITADQYINRVNELFDM KLISENKKKRLLLKDTEDLDRFINQNLVDTRYITREMIGILKAYNRVKGYPT KVIALKPAFTKNYSNAFDIYKNRDFGLQHHALDAAVLGITDIMLNRLFPNY ESGRISTSFAKHHQMLSEVGNDKKLKDSTKYVIRKMYEKVFGNSPLDYG SLVYVCKNTMPLYSQKSNRNWKGEKFPASRRLPYQKYDIKNGNIVGVYD DKSEENIGKQNSEVLRLFEINNKNHAFDSVNCVAYDIYRIPEKKNNKVIWK NIGIHIPKCIVDEDGTIQKDKYIKLIKKYYGFDDNLIDEKGDLITAYFKLRLYK NDLVGNTEKHEIRDLVLGSIADKKLELNKIDCFVYQKIAQKKASLFKMIIDN QGSSKYDIHNYKEYQECECKEEILRDMIFDDMNIKNQEFASLLYAKIINDK KLDLENINKICWILAYYQQFLNEITGKKEFNTGRGIGALKKDESINYFKLKT SPLGVRFNTEKNQWTGPVGYENAFKIIKKEAFTWKITV CUAZ Type II MGKPIPNPLLGLDSTKRTADGSEFESPKKKRKVGSKTNKKIYKDNQYTLS 9 Cas IDPGLNNVGWALFDAKNKKMVEMGVRKFEPAKEAKISRLKRSARRNNG mammalian RKKWRKQQLKQAFIDFNLIGEDEFKNSQFYEFHPEQYKAAGVKNLMDLR expression CYALQNKVTYQELFQCIYNILKYRGNFLRDDIDFENGESLTEELLIQEITNF construct MIKSLQLDIDNLDKLKESISTMCNQNIDLKEIPYTEISDDKVSQKSIENVFK (with N- ALKDQKFNPFMIEDDPILIDEEEGKSIIPEAKTKTIYMKSVQKCGDLSNRFE terminal QLLNITDSYRLYKVLNGEKYICTAYKKKLDRVADYYSFKDSPEGCSKYSDI methionine, ESAKIEEEITYITNNMNATKGKPEKYGYTRKNPRGPIHVIHNVLNRYPNGL stop codon, LVKEAVDILRTQQNYYGKEVISDEFIEVIRYILKGRIPYYIGPLSETAKNQW V5 tag, N- CTKTGKFKYSYNYTIEHMDGVDEERSIKDWKDRMRGHCTYLPEAFALPK terminal NLS GSFVAETLLILNELNVLTAIDKNNNTYYFTTNDKIKVFDYLFLDENHYGDV and C- KYKEICDLLDLAYFGPKNQKSQVKTFKQNYTLYKSITSICTALKLHSIIDLF terminal NLS) STKFQENQLKTIEEIVLNLNLYDEKKHKYQALTKMGISDENSKKLSALNSK (aa) KFYGLSEKFVLKTSLNKDGRCLLEILFEDNQPGYTNEMMTAITNACDMD GNPVSFIANKYEAAFKELEKNGELELNFNSLLEIMNKTGNGFQIPASRPVL RSINQMMKVIQAVYTYYGVPKRIVLEIPRDMSGKNTNHEMPAKHYKTME ALHQDIMKQCKDKDHPRKTCIDQKWNDLKDFELNQKNRIKLELYLRQNG MDMITGEKINLNELENYEIDHILPRGFGDDSMDDKILIRFVSNALKKDRTPL MWITDPTDPVINKSGKEITADQYINRVNELFDMKLISENKKKRLLLKDTED LDRFINQNLVDTRYITREMIGILKAYNRVKGYPTKVIALKPAFTKNYSNAFD IYKNRDFGLQHHALDAAVLGITDIMLNRLFPNYESGRISTSFAKHHQMLSE VGNDKKLKDSTKYVIRKMYEKVFGNSPLDYGSLVYVCKNTMPLYSQKSN RNWKGEKFPASRRLPYQKYDIKNGNIVGVYDDKSEENIGKQNSEVLRLF EINNKNHAFDSVNCVAYDIYRIPEKKNNKVIWKNIGIHIPKCIVDEDGTIQK DKYIKLIKKYYGFDDNLIDEKGDLITAYFKLRLYKNDLVGNTEKHEIRDLVL GSIADKKLELNKIDCFVYQKIAQKKASLFKMIIDNQGSSKYDIHNYKEYQE CECKEEILRDMIFDDMNIKNQEFASLLYAKIINDKKLDLENINKICWILAYYQ QFLNEITGKKEFNTGRGIGALKKDESINYFKLKTSPLGVRFNTEKNQWTG PVGYENAFKIIKKEAFTWKITVSRKRTADGSEFESPKKKRKV CUAZ Type II ATGAAAACAAATAAAAAAATCTATAAAGATAATCAATATACATTATCAAT 10 Cas coding TGATCCAGGTTTAAATAATGTAGGATGGGCATTATTTGATGCAAAAAA sequence (nt) TAAAAAAATGGTCGAGATGGGTGTAAGAAAGTTTGAACCTGCTAAAGA (not codon AGCTAAGATATCTCGATTAAAGCGAAGTGCTAGAAGAAATAATGGAAG optimized) AAAAAAATGGAGAAAGCAGCAGCTTAAGCAGGCATTTATAGATTTTAA TTTGATAGGGGAAGATGAATTTAAGAATAGTCAGTTTTATGAATTTCAT CCTGAGCAATATAAAGCAGCCGGTGTTAAAAATTTAATGGATTTAAGA TGTTATGCACTTCAGAATAAAGTTACTTATCAAGAATTATTTCAATGCA TATATAACATATTAAAGTATCGTGGTAATTTTTTAAGAGATGATATTGA TABLE 1B CUAZ Type II Cas Sequences Name Sequence SEQ ID NO TTTTGAAAATGGAGAATCTTTAACAGAAGAGCTTTTGATCCAGGAAAT TACAAACTTTATGATTAAGTCATTGCAATTGGATATTGATAATCTAGAT AAATTAAAAGAATCTATTAGTACAATGTGTAATCAAAATATTGATTTAAA AGAAATTCCATATACAGAAATATCAGATGATAAAGTATCTCAAAAGAGT ATTGAAAATGTGTTTAAGGCACTTAAAGATCAAAAATTCAATCCTTTTA TGATTGAAGATGATCCGATTCTGATTGATGAGGAAGAGGGTAAATCTA TTATTCCAGAGGCAAAAACAAAGACAATTTATATGAAATCGGTACAAA AATGTGGTGATTTATCGAATCGATTTGAACAGTTATTAAATATTACAGA TTCATATCGACTTTATAAAGTGTTGAACGGGGAAAAATATATTTGCACT GCGTATAAAAAGAAGCTTGATCGAGTTGCGGATTATTATTCTTTTAAA GATAGCCCAGAAGGGTGCTCTAAATATTCAGATATTGAATCCGCAAAG ATTGAAGAAGAAATAACATATATTACGAATAATATGAATGCTACTAAGG GAAAACCAGAAAAGTATGGTTATACTCGTAAGAATCCACGAGGTCCTA TTCATGTTATACATAATGTTTTGAACAGATATCCTAATGGTCTGCTTGT TAAAGAGGCTGTTGATATTTTAAGAACACAACAGAACTATTATGGTAA AGAAGTGATTTCTGATGAATTTATTGAGGTAATTCGATATATCTTAAAA GGTCGAATTCCATATTATATTGGGCCATTGTCAGAAACGGCAAAAAAT CAGTGGTGTACAAAGACTGGAAAATTCAAATATAGCTACAACTATACA ATTGAACACATGGATGGTGTGGACGAAGAACGTTCAATAAAAGATTG GAAAGATCGTATGCGTGGACATTGTACATATCTTCCTGAGGCATTTGC ACTTCCAAAAGGATCGTTTGTAGCTGAAACATTGCTTATCTTAAATGAA TTAAATGTGCTTACGGCGATAGACAAAAATAATAACACGTATTATTTCA CAACGAATGACAAAATTAAAGTATTTGATTATCTATTTTTAGATGAGAA TCATTACGGGGATGTTAAATATAAAGAAATTTGTGATCTACTGGATTTG GCATATTTTGGACCGAAAAATCAAAAATCACAGGTCAAAACATTTAAG CAGAATTATACGTTATATAAAAGTATAACAAGTATTTGTACTGCTTTAA AATTGCATTCAATTATTGATTTGTTTAGTACAAAATTTCAAGAAAATCAA CTAAAGACAATAGAAGAAATTGTTCTGAATCTTAACCTTTACGATGAG AAGAAGCACAAGTATCAGGCATTAACAAAAATGGGAATATCTGATGAA AATTCAAAAAAACTTTCTGCATTGAACTCAAAGAAATTTTATGGATTGT CAGAAAAATTTGTTTTAAAAACATCATTAAATAAAGATGGACGATGTCT TCTAGAAATATTATTTGAAGATAACCAGCCCGGTTATACAAATGAAAT GATGACAGCTATTACGAATGCTTGTGATATGGATGGAAATCCTGTTTC GTTTATTGCCAATAAATATGAGGCAGCTTTTAAAGAGTTAGAAAAAAAT GGAGAGTTAGAATTAAATTTTAATAGTCTTTTAGAAATAATGAATAAAA CTGGAAATGGTTTCCAAATTCCTGCTTCTCGTCCAGTACTTCGTTCTA TTAATCAAATGATGAAAGTGATTCAGGCTGTATATACATATTACGGGG TTCCTAAACGCATTGTATTAGAAATTCCTAGAGATATGTCTGGAAAAAA TACTAATCATGAAATGCCTGCAAAACACTATAAAACTATGGAAGCTTTA CATCAAGATATTATGAAGCAGTGTAAGGATAAAGATCATCCTAGAAAA ACTTGCATCGATCAAAAATGGAATGATTTAAAAGATTTTGAGCTAAATC AAAAAAATAGAATAAAACTTGAATTGTATCTTCGCCAAAATGGAATGG ATATGATAACAGGAGAAAAAATCAATTTGAATGAGCTTGAAAACTATG AAATTGACCATATTCTTCCAAGAGGATTTGGGGATGATTCAATGGATG ATAAGATACTAATACGTTTTGTATCTAATGCATTGAAAAAGGATAGAAC ACCTCTTATGTGGATTACAGATCCAACGGATCCAGTTATAAATAAATC AGGTAAAGAAATTACAGCCGATCAGTATATAAATCGAGTGAACGAGCT CTTTGATATGAAGTTAATATCTGAAAATAAAAAGAAGAGATTGTTATTA AAAGATACAGAAGATTTGGATCGATTTATTAATCAGAATCTTGTTGATA CTCGATATATCACAAGAGAAATGATTGGAATTTTAAAAGCATATAATCG TGTAAAGGGTTATCCTACAAAAGTGATCGCTTTAAAACCTGCTTTTAC AAAGAATTATTCAAATGCATTCGACATTTATAAGAACAGAGATTTTGGT TTACAACACCACGCGTTGGACGCAGCGGTATTAGGTATTACGGATAT TATGTTAAATAGATTATTTCCAAATTATGAAAGTGGACGAATATCTACA TCATTCGCTAAGCATCATCAGATGTTGTCAGAAGTTGGAAATGATAAA AAGTTAAAAGATTCGACTAAATATGTAATACGTAAAATGTATGAAAAAG TATTTGGTAATTCGCCATTAGATTATGGTTCACTTGTATATGTTTGTAA TABLE 1B CUAZ Type II Cas Sequences Name Sequence SEQ ID NO AAATACCATGCCTTTATATTCTCAAAAATCAAATAGAAACTGGAAGGG TGAAAAGTTTCCGGCTTCTAGAAGATTGCCTTATCAAAAATATGACAT AAAAAATGGAAATATAGTTGGAGTATATGATGATAAAAGTGAAGAAAA TATAGGTAAACAAAACTCTGAAGTATTACGATTATTTGAAATCAACAAT AAAAATCATGCGTTTGATAGTGTGAATTGTGTTGCCTATGATATATATA GAATTCCTGAAAAGAAAAATAATAAAGTGATTTGGAAAAATATAGGTAT ACATATTCCAAAATGTATTGTAGATGAAGATGGCACTATACAAAAGGA TAAGTATATTAAGCTAATTAAGAAGTATTATGGTTTTGATGATAATCTT ATAGATGAAAAAGGTGACCTTATTACAGCTTATTTCAAACTACGTCTTT ATAAAAATGATTTAGTAGGAAATACTGAAAAACATGAAATTAGAGATTT AGTATTGGGAAGTATTGCTGATAAAAAATTAGAATTAAATAAAATTGAT TGTTTTGTATATCAGAAAATTGCGCAGAAAAAAGCAAGTTTATTTAAAA TGATTATCGATAATCAAGGATCTTCAAAATACGATATTCATAATTATAA GGAATATCAGGAGTGTGAATGTAAAGAAGAGATTCTACGAGATATGAT TTTTGATGATATGAATATCAAAAATCAGGAGTTTGCTTCTCTTCTATAC GCTAAGATTATTAATGATAAAAAATTAGATTTGGAAAATATAAACAAGA TTTGTTGGATACTTGCCTATTATCAGCAATTTTTAAATGAAATTACTGG TAAAAAAGAATTTAATACAGGACGAGGAATAGGTGCTTTAAAGAAAGA TGAGAGTATTAATTACTTCAAGCTAAAAACATCACCGTTGGGGGTGCG ATTTAATACAGAAAAAAACCAATGGACCGGACCTGTTGGATATGAAAA TGCCTTTAAAATTATAAAAAAAGAAGCATTTACTTGGAAAATTACAGTT TAA CUAZ Type II AAAACCAATAAAAAGATCTACAAGGACAACCAGTATACCCTGTCTATC 11 Cas coding GATCCTGGACTGAACAACGTGGGCTGGGCCCTGTTCGATGCCAAGA sequence (nt) ACAAGAAAATGGTGGAGATGGGCGTGCGGAAGTTCGAGCCTGCCAA (human GGAGGCTAAGATCTCCAGACTGAAGAGAAGCGCACGGAGGAACAAC codon- GGCCGCAAGAAGTGGCGGAAGCAGCAGCTGAAGCAGGCCTTTATCG optimized; ACTTCAACCTTATCGGGGAGGATGAGTTCAAGAACAGCCAGTTCTAC lacking N- GAGTTCCACCCTGAGCAGTATAAGGCCGCTGGCGTGAAGAATCTGAT terminal GGACCTGCGCTGCTACGCTCTCCAGAACAAGGTGACCTACCAAGAG methionine; CTGTTCCAGTGTATCTACAACATCCTGAAGTACCGGGGAAACTTCCTG no stop AGGGATGACATCGACTTTGAAAATGGAGAGTCCCTGACAGAAGAACT codon) GTTGATCCAGGAAATTACCAATTTCATGATCAAGTCCCTGCAGTTGGA TATTGACAACTTGGACAAGCTGAAGGAGTCCATCAGCACCATGTGCA ACCAGAACATAGACCTGAAGGAAATCCCCTACACCGAAATCAGCGAC GACAAGGTGAGCCAGAAGAGCATTGAGAACGTGTTCAAGGCCCTGAA GGACCAGAAGTTCAACCCCTTCATGATCGAGGATGATCCTATCCTGA TCGACGAGGAAGAAGGCAAGAGCATCATCCCTGAGGCCAAGACCAA GACTATCTACATGAAATCTGTGCAGAAATGCGGCGACCTGAGCAACA GATTCGAGCAACTGCTGAACATCACCGACTCCTACAGACTGTACAAG GTGCTGAACGGCGAGAAGTACATCTGCACCGCTTACAAGAAGAAACT GGACCGGGTGGCCGATTACTACTCCTTCAAGGACAGCCCTGAGGGT TGTAGCAAATACAGCGACATCGAGTCCGCCAAGATCGAGGAAGAAAT CACCTATATCACCAACAACATGAACGCCACAAAAGGAAAACCCGAGA AGTACGGATACACAAGAAAGAATCCTAGAGGCCCCATCCACGTGATC CACAACGTGCTGAACAGATACCCTAACGGCCTTCTCGTGAAGGAAGC GGTGGACATCCTGAGAACCCAGCAAAACTACTACGGCAAGGAGGTTA TAAGCGACGAGTTCATCGAGGTGATCCGCTACATCTTGAAGGGCAGA ATCCCATACTACATCGGCCCTCTGTCTGAAACAGCCAAGAACCAGTG GTGCACCAAGACCGGCAAATTCAAGTACAGCTACAACTACACCATCG AGCACATGGATGGCGTGGACGAAGAGAGATCTATCAAGGACTGGAA GGATAGAATGAGAGGCCACTGTACCTACCTGCCTGAGGCCTTCGCCC TGCCAAAGGGCAGCTTCGTGGCCGAGACACTGCTGATCCTGAACGA GCTGAACGTGCTGACCGCCATCGACAAGAACAACAACACCTATTACT TCACAACAAACGACAAGATCAAAGTGTTCGATTATCTGTTTCTGGATG AGAATCATTACGGCGATGTGAAGTACAAGGAAATCTGTGATCTCCTG GACCTGGCCTACTTCGGCCCTAAGAACCAGAAATCTCAGGTGAAAAC TABLE 1B CUAZ Type II Cas Sequences Name Sequence SEQ ID NO CTTCAAGCAAAACTACACCCTGTACAAAAGCATCACAAGTATCTGCAC AGCCCTGAAGCTGCACAGCATTATCGATCTGTTCAGCACAAAGTTCC AGGAAAACCAGCTGAAAACCATCGAGGAAATCGTGCTGAATCTGAAT CTGTACGACGAAAAGAAGCACAAGTATCAAGCCTTGACCAAGATGGG AATCAGCGACGAGAATAGCAAGAAGCTGAGTGCCCTGAATAGCAAGA AATTTTACGGCCTGAGCGAGAAGTTTGTGCTGAAAACAAGCCTGAAC AAGGACGGCAGATGCCTGCTGGAAATCCTGTTTGAAGATAATCAACC TGGCTATACCAATGAAATGATGACCGCCATCACCAATGCTTGTGACAT GGATGGCAACCCTGTATCCTTCATCGCTAATAAATACGAGGCCGCTTT TAAAGAACTGGAAAAAAACGGAGAGCTGGAACTGAACTTCAATAGCC TTCTGGAAATCATGAACAAGACCGGCAACGGATTCCAGATCCCGGCT TCCAGACCCGTCCTGAGAAGCATCAACCAGATGATGAAGGTGATCCA GGCCGTTTACACCTACTACGGCGTGCCCAAGCGGATCGTGCTGGAA ATCCCTAGAGATATGAGCGGCAAGAATACCAACCATGAGATGCCCGC CAAGCACTACAAGACGATGGAAGCCCTGCACCAGGACATTATGAAGC AGTGTAAAGACAAGGATCACCCCAGAAAGACATGCATCGATCAGAAG TGGAACGACCTGAAGGACTTCGAGCTGAACCAGAAGAACCGGATCAA GCTCGAACTGTACCTGAGGCAGAACGGCATGGACATGATCACCGGC GAGAAGATCAATCTGAACGAGCTGGAAAACTACGAGATCGACCACAT CCTGCCTAGAGGCTTCGGAGATGACAGCATGGATGACAAGATTCTGA TCAGATTCGTGAGCAATGCCCTGAAGAAGGATAGAACACCTCTGATG TGGATCACCGACCCTACCGATCCTGTGATCAATAAGTCCGGCAAAGA GATCACTGCCGACCAGTACATCAACCGGGTCAACGAACTGTTCGACA TGAAACTGATAAGCGAGAATAAGAAGAAGCGGCTGCTGCTGAAGGAC ACAGAGGATCTGGACAGATTTATCAACCAGAACCTGGTCGACACCAG ATACATCACTAGAGAAATGATCGGCATCCTGAAGGCCTACAACAGAG TTAAGGGTTATCCTACAAAGGTGATCGCTCTGAAGCCCGCCTTTACCA AGAACTACTCTAACGCCTTCGACATCTACAAGAACCGGGATTTCGGC CTGCAGCACCACGCCCTGGATGCTGCTGTGCTGGGCATCACAGACA TCATGCTGAACAGGCTGTTCCCTAATTACGAGAGCGGCAGAATCAGC ACCAGCTTCGCCAAGCACCATCAAATGCTGAGCGAGGTGGGCAACG ACAAGAAGTTAAAAGACTCCACCAAGTACGTGATCAGAAAGATGTAC GAGAAGGTGTTCGGCAACAGCCCTCTGGACTACGGCTCTCTGGTGTA CGTGTGCAAGAACACCATGCCACTGTATTCACAAAAGTCTAACAGAAA TTGGAAGGGCGAAAAGTTCCCCGCCAGCAGACGGCTGCCCTACCAG AAATACGATATTAAGAATGGCAACATCGTCGGTGTCTACGACGACAA GTCCGAGGAGAACATCGGCAAGCAGAACAGCGAAGTGTTACGGCTG TTCGAGATCAACAATAAGAATCACGCCTTTGACAGCGTGAACTGCGT GGCCTACGACATCTACAGAATCCCCGAGAAGAAAAACAACAAGGTGA TCTGGAAGAACATCGGCATTCACATCCCTAAGTGCATCGTGGACGAG GACGGCACAATCCAGAAGGACAAATACATAAAGCTGATCAAGAAGTA CTACGGCTTCGACGACAACCTGATCGATGAGAAAGGCGACCTTATTA CCGCCTACTTTAAACTGCGGCTGTACAAAAATGATCTGGTGGGCAAC ACCGAAAAGCACGAGATCAGAGATCTTGTGCTGGGAAGCATCGCCGA TAAAAAACTGGAGTTAAACAAGATTGACTGCTTTGTGTACCAGAAGAT CGCCCAGAAGAAGGCCAGCCTGTTTAAAATGATCATCGACAACCAGG GCAGCTCTAAGTACGACATCCACAACTACAAAGAGTACCAGGAATGC GAGTGCAAGGAAGAGATCCTGAGAGACATGATCTTCGACGACATGAA TATCAAAAATCAGGAGTTCGCCAGCCTGCTGTACGCCAAGATTATCAA CGACAAGAAGCTGGACCTGGAAAACATCAACAAGATTTGCTGGATTC TGGCCTACTACCAGCAGTTTCTGAATGAGATCACAGGCAAGAAGGAA TTTAACACCGGCCGGGGCATCGGCGCCCTGAAAAAGGACGAGTCTA TCAACTACTTCAAGCTGAAAACCTCTCCACTGGGAGTGCGGTTCAAC ACCGAGAAGAACCAGTGGACCGGCCCTGTGGGCTACGAGAACGCCT TCAAGATCATCAAGAAAGAGGCCTTCACCTGGAAAATTACAGTG CUAZ Type II ATGGGCAAACCTATCCCCAATCCCCTGCTGGGCCTGGACAGCACCAA 12 Cas GCGGACTGCCGACGGCAGTGAGTTCGAGTCTCCCAAGAAAAAGAGG TABLE 1B CUAZ Type II Cas Sequences Name Sequence SEQ ID NO mammalian AAAGTGGGATCCAAAACCAATAAAAAGATCTACAAGGACAACCAGTAT expression ACCCTGTCTATCGATCCTGGACTGAACAACGTGGGCTGGGCCCTGTT construct CGATGCCAAGAACAAGAAAATGGTGGAGATGGGCGTGCGGAAGTTC (with N- GAGCCTGCCAAGGAGGCTAAGATCTCCAGACTGAAGAGAAGCGCAC terminal GGAGGAACAACGGCCGCAAGAAGTGGCGGAAGCAGCAGCTGAAGCA methionine, GGCCTTTATCGACTTCAACCTTATCGGGGAGGATGAGTTCAAGAACA stop codon, GCCAGTTCTACGAGTTCCACCCTGAGCAGTATAAGGCCGCTGGCGTG V5 tag, N- AAGAATCTGATGGACCTGCGCTGCTACGCTCTCCAGAACAAGGTGAC terminal NLS CTACCAAGAGCTGTTCCAGTGTATCTACAACATCCTGAAGTACCGGG and C- GAAACTTCCTGAGGGATGACATCGACTTTGAAAATGGAGAGTCCCTG terminal NLS) ACAGAAGAACTGTTGATCCAGGAAATTACCAATTTCATGATCAAGTCC (nt) CTGCAGTTGGATATTGACAACTTGGACAAGCTGAAGGAGTCCATCAG CACCATGTGCAACCAGAACATAGACCTGAAGGAAATCCCCTACACCG AAATCAGCGACGACAAGGTGAGCCAGAAGAGCATTGAGAACGTGTTC AAGGCCCTGAAGGACCAGAAGTTCAACCCCTTCATGATCGAGGATGA TCCTATCCTGATCGACGAGGAAGAAGGCAAGAGCATCATCCCTGAGG CCAAGACCAAGACTATCTACATGAAATCTGTGCAGAAATGCGGCGAC CTGAGCAACAGATTCGAGCAACTGCTGAACATCACCGACTCCTACAG ACTGTACAAGGTGCTGAACGGCGAGAAGTACATCTGCACCGCTTACA AGAAGAAACTGGACCGGGTGGCCGATTACTACTCCTTCAAGGACAGC CCTGAGGGTTGTAGCAAATACAGCGACATCGAGTCCGCCAAGATCGA GGAAGAAATCACCTATATCACCAACAACATGAACGCCACAAAAGGAA AACCCGAGAAGTACGGATACACAAGAAAGAATCCTAGAGGCCCCATC CACGTGATCCACAACGTGCTGAACAGATACCCTAACGGCCTTCTCGT GAAGGAAGCGGTGGACATCCTGAGAACCCAGCAAAACTACTACGGC AAGGAGGTTATAAGCGACGAGTTCATCGAGGTGATCCGCTACATCTT GAAGGGCAGAATCCCATACTACATCGGCCCTCTGTCTGAAACAGCCA AGAACCAGTGGTGCACCAAGACCGGCAAATTCAAGTACAGCTACAAC TACACCATCGAGCACATGGATGGCGTGGACGAAGAGAGATCTATCAA GGACTGGAAGGATAGAATGAGAGGCCACTGTACCTACCTGCCTGAG GCCTTCGCCCTGCCAAAGGGCAGCTTCGTGGCCGAGACACTGCTGA TCCTGAACGAGCTGAACGTGCTGACCGCCATCGACAAGAACAACAAC ACCTATTACTTCACAACAAACGACAAGATCAAAGTGTTCGATTATCTG TTTCTGGATGAGAATCATTACGGCGATGTGAAGTACAAGGAAATCTGT GATCTCCTGGACCTGGCCTACTTCGGCCCTAAGAACCAGAAATCTCA GGTGAAAACCTTCAAGCAAAACTACACCCTGTACAAAAGCATCACAAG TATCTGCACAGCCCTGAAGCTGCACAGCATTATCGATCTGTTCAGCA CAAAGTTCCAGGAAAACCAGCTGAAAACCATCGAGGAAATCGTGCTG AATCTGAATCTGTACGACGAAAAGAAGCACAAGTATCAAGCCTTGACC AAGATGGGAATCAGCGACGAGAATAGCAAGAAGCTGAGTGCCCTGAA TAGCAAGAAATTTTACGGCCTGAGCGAGAAGTTTGTGCTGAAAACAA GCCTGAACAAGGACGGCAGATGCCTGCTGGAAATCCTGTTTGAAGAT AATCAACCTGGCTATACCAATGAAATGATGACCGCCATCACCAATGCT TGTGACATGGATGGCAACCCTGTATCCTTCATCGCTAATAAATACGAG GCCGCTTTTAAAGAACTGGAAAAAAACGGAGAGCTGGAACTGAACTT CAATAGCCTTCTGGAAATCATGAACAAGACCGGCAACGGATTCCAGA TCCCGGCTTCCAGACCCGTCCTGAGAAGCATCAACCAGATGATGAAG GTGATCCAGGCCGTTTACACCTACTACGGCGTGCCCAAGCGGATCGT GCTGGAAATCCCTAGAGATATGAGCGGCAAGAATACCAACCATGAGA TGCCCGCCAAGCACTACAAGACGATGGAAGCCCTGCACCAGGACATT ATGAAGCAGTGTAAAGACAAGGATCACCCCAGAAAGACATGCATCGA TCAGAAGTGGAACGACCTGAAGGACTTCGAGCTGAACCAGAAGAACC GGATCAAGCTCGAACTGTACCTGAGGCAGAACGGCATGGACATGATC ACCGGCGAGAAGATCAATCTGAACGAGCTGGAAAACTACGAGATCGA CCACATCCTGCCTAGAGGCTTCGGAGATGACAGCATGGATGACAAGA TTCTGATCAGATTCGTGAGCAATGCCCTGAAGAAGGATAGAACACCT CTGATGTGGATCACCGACCCTACCGATCCTGTGATCAATAAGTCCGG TABLE 1B CUAZ Type II Cas Sequences Name Sequence SEQ ID NO CAAAGAGATCACTGCCGACCAGTACATCAACCGGGTCAACGAACTGT TCGACATGAAACTGATAAGCGAGAATAAGAAGAAGCGGCTGCTGCTG AAGGACACAGAGGATCTGGACAGATTTATCAACCAGAACCTGGTCGA CACCAGATACATCACTAGAGAAATGATCGGCATCCTGAAGGCCTACA ACAGAGTTAAGGGTTATCCTACAAAGGTGATCGCTCTGAAGCCCGCC TTTACCAAGAACTACTCTAACGCCTTCGACATCTACAAGAACCGGGAT TTCGGCCTGCAGCACCACGCCCTGGATGCTGCTGTGCTGGGCATCA CAGACATCATGCTGAACAGGCTGTTCCCTAATTACGAGAGCGGCAGA ATCAGCACCAGCTTCGCCAAGCACCATCAAATGCTGAGCGAGGTGG GCAACGACAAGAAGTTAAAAGACTCCACCAAGTACGTGATCAGAAAG ATGTACGAGAAGGTGTTCGGCAACAGCCCTCTGGACTACGGCTCTCT GGTGTACGTGTGCAAGAACACCATGCCACTGTATTCACAAAAGTCTAA CAGAAATTGGAAGGGCGAAAAGTTCCCCGCCAGCAGACGGCTGCCC TACCAGAAATACGATATTAAGAATGGCAACATCGTCGGTGTCTACGAC GACAAGTCCGAGGAGAACATCGGCAAGCAGAACAGCGAAGTGTTAC GGCTGTTCGAGATCAACAATAAGAATCACGCCTTTGACAGCGTGAAC TGCGTGGCCTACGACATCTACAGAATCCCCGAGAAGAAAAACAACAA GGTGATCTGGAAGAACATCGGCATTCACATCCCTAAGTGCATCGTGG ACGAGGACGGCACAATCCAGAAGGACAAATACATAAAGCTGATCAAG AAGTACTACGGCTTCGACGACAACCTGATCGATGAGAAAGGCGACCT TATTACCGCCTACTTTAAACTGCGGCTGTACAAAAATGATCTGGTGGG CAACACCGAAAAGCACGAGATCAGAGATCTTGTGCTGGGAAGCATCG CCGATAAAAAACTGGAGTTAAACAAGATTGACTGCTTTGTGTACCAGA AGATCGCCCAGAAGAAGGCCAGCCTGTTTAAAATGATCATCGACAAC CAGGGCAGCTCTAAGTACGACATCCACAACTACAAAGAGTACCAGGA ATGCGAGTGCAAGGAAGAGATCCTGAGAGACATGATCTTCGACGACA TGAATATCAAAAATCAGGAGTTCGCCAGCCTGCTGTACGCCAAGATTA TCAACGACAAGAAGCTGGACCTGGAAAACATCAACAAGATTTGCTGG ATTCTGGCCTACTACCAGCAGTTTCTGAATGAGATCACAGGCAAGAA GGAATTTAACACCGGCCGGGGCATCGGCGCCCTGAAAAAGGACGAG TCTATCAACTACTTCAAGCTGAAAACCTCTCCACTGGGAGTGCGGTTC AACACCGAGAAGAACCAGTGGACCGGCCCTGTGGGCTACGAGAACG CCTTCAAGATCATCAAGAAAGAGGCCTTCACCTGGAAAATTACAGTGT CTAGAAAGCGGACAGCAGACGGCTCCGAATTTGAAAGCCCTAAGAAA AAGAGAAAGGTGTGA [0079] In some embodiments a CUAZ Type II Cas protein comprises an amino acid sequence of SEQ ID NO:7, SEQ ID NO:8, or SEQ ID NO:9. In some embodiments, a CUAZ Type II Cas protein has nickase activity, for example resulting from one or more amino acid substitutions relative to the sequence of SEQ ID NO:7, SEQ ID NO:8, or SEQ ID NO:9. In some embodiments, the one or more amino acid substitutions providing nickase activity comprise a D18A substitution, wherein the position of the D18A substitution is defined with respect to the amino acid numbering of SEQ ID NO:8. In some embodiments, the one or more amino acid substitutions providing nickase activity comprise an H845A substitution, wherein the position of the H845A substitution is defined with respect to the amino acid numbering of SEQ ID NO:8. In some embodiments, a CUAZ Type II Cas protein is catalytically inactive, for example due to a D18A substitution in combination with a H845A substitution. 6.2.1.3. AHWY Type II Cas Proteins [0080] In one aspect, the disclosure provides AHWY Type II Cas proteins. AHWY Type II Cas proteins can be further classified as Type IIA Cas proteins. The AHWY Type II Cas proteins typically comprise an amino acid sequence that is at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 85%, at least 90%, or at least 95% identical to SEQ ID NO:13. In some embodiments, the AHWY Type II Cas proteins comprise an amino acid sequence that is at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical to SEQ ID NO:13. In some embodiments, an AHWY Type II Cas protein comprises an amino acid sequence that is identical to SEQ ID NO:13. [0081] Exemplary AHWY Type II Cas protein sequences and nucleotide sequences encoding exemplary AHWY Type II Cas proteins are set forth in Table 1C. TABLE 1C AHWY Type II Cas Sequences Name Sequence SEQ ID NO AHWY Type II NLRNVGEYNIGLDIGTGSVGWAVIDSDGEICHFKGKPTWGSRLFPSALPA 13 Cas amino SEARVHRGQRRRYTRRKWRLFLLQELFAGAIEANDPEFFIRLNQSHLLPE acid DRQSDHKDYLFTLFNDSSFDEKAYFKKFPTIYHLRKWLMETDEKADPRLI sequence YLAFHNIVKHRGNFLQQDNTGLNSQNANVRESVEELCAQLQEYCDSLDI (without N- PCTVEDNVDKIVQVLSDTHSVRSHLQEEVQKLLGIGVCDLLDKTSAKKMA terminal KALSGALVGLSAELADVFFLVIEKPEDAKTKIYLSKDEDVESFEEICPDEAL methionine) PLFEAMNKVYSSFVLQEILSSCPGKTLSVNKVKDYEQYGEDLKLLKALVK EYAPEKYDDFFRGEFYRPSSLHPQKYVYNKSVAKGYTRYNEVRGVAYE DFKKDVEKLFRDTPAVEDARYLDMMERFKEEKFLRRLKTSDNGAIPFQL HLEEMDKIIQNQARFYPFLEEKKKELDSLVTFRIPYYVGPLTQKNARKDR KGSNRFSWSERIEGKEHEVIKPWNWEEIIDKDRSATEFILRMTGTCTYLQ GEPVLPRCSLLYEEYCVLNELNGAHFTQDGDKEHRFDYQDRSDMLKEL FRKGKVSYKKAADWMVQRGNSNVRVSGGQGEKGFESKLGSYIFFAKDV FGVDEIPESDYPMIEEIILWNTIFEDRDILKEKLKRNYGDCLSDEQIKKIVRK RFTGWGRLSKKLLCGIKVETDNGRMSIMDVLREGNPNNNGLSKAMVFM EIIRDTGLGFDKRIDELNKEAMIGSTGLAIEDLPGSPALRRGVNQALGIVEE IVHIAGHAPNNIFIEVTREEDGRKKGQRTKKRYESLKEAMSKLKKESPEF WDAATSSQLAEKAKESAALDEKLTLYFLQGGKSLYSGKPLEIGRLSEYQ VDHIIPQSYIKDDSFENKALVLASENQNKSDQMLLPVEMRRKMRPYWDA LLSAGLIGKKKHTNLLRDHIGEKQMKGFIARQLVETSQILKIVQSFLEEQY PETNTLPVKAGLSHELRERIGLVKCREVNDFHHAHDALLAAEIGRFIQKR HPGMYTNPIGYTQVMKQFVKNESDAVRRGHEPGTSPFVIASFMTSGFD EETGELFKDDWSASYEIAKLKSYFNYRQCFISRMPEETSGAFWDQTIYSP RGGKKTPSIPLKKGLSTEKHGGYSGQNPAYFVLIAAYDSKGKPKRLFEG VPIAVAELIKSKKQSIEEWAKQLVESRGFVFGQVLRARLLKYARIDHNGN EYYVPALDCVYSSRELILNENQTTIAAAVCDLARVSDEALVPSEVLELFDA LTEKCLILCTRFKPVYEKLVEGRERFVALSMQDKRKQIVGILKFFNCVTAR VDLSLIGGSPHAGGITNGFLQKPNDSFDIVDQSITGMFERRTHVGL AHWY Type II MNLRNVGEYNIGLDIGTGSVGWAVIDSDGEICHFKGKPTWGSRLFPSAL 14 Cas amino PASEARVHRGQRRRYTRRKWRLFLLQELFAGAIEANDPEFFIRLNQSHLL acid PEDRQSDHKDYLFTLFNDSSFDEKAYFKKFPTIYHLRKWLMETDEKADP sequence RLIYLAFHNIVKHRGNFLQQDNTGLNSQNANVRESVEELCAQLQEYCDS LDIPCTVEDNVDKIVQVLSDTHSVRSHLQEEVQKLLGIGVCDLLDKTSAK KMAKALSGALVGLSAELADVFFLVIEKPEDAKTKIYLSKDEDVESFEEICP DEALPLFEAMNKVYSSFVLQEILSSCPGKTLSVNKVKDYEQYGEDLKLLK ALVKEYAPEKYDDFFRGEFYRPSSLHPQKYVYNKSVAKGYTRYNEVRG VAYEDFKKDVEKLFRDTPAVEDARYLDMMERFKEEKFLRRLKTSDNGAI PFQLHLEEMDKIIQNQARFYPFLEEKKKELDSLVTFRIPYYVGPLTQKNAR KDRKGSNRFSWSERIEGKEHEVIKPWNWEEIIDKDRSATEFILRMTGTCT YLQGEPVLPRCSLLYEEYCVLNELNGAHFTQDGDKEHRFDYQDRSDML KELFRKGKVSYKKAADWMVQRGNSNVRVSGGQGEKGFESKLGSYIFFA KDVFGVDEIPESDYPMIEEIILWNTIFEDRDILKEKLKRNYGDCLSDEQIKKI VRKRFTGWGRLSKKLLCGIKVETDNGRMSIMDVLREGNPNNNGLSKAM VFMEIIRDTGLGFDKRIDELNKEAMIGSTGLAIEDLPGSPALRRGVNQALG IVEEIVHIAGHAPNNIFIEVTREEDGRKKGQRTKKRYESLKEAMSKLKKES TABLE 1C AHWY Type II Cas Sequences Name Sequence SEQ ID NO PEFWDAATSSQLAEKAKESAALDEKLTLYFLQGGKSLYSGKPLEIGRLSE YQVDHIIPQSYIKDDSFENKALVLASENQNKSDQMLLPVEMRRKMRPYW DALLSAGLIGKKKHTNLLRDHIGEKQMKGFIARQLVETSQILKIVQSFLEE QYPETNTLPVKAGLSHELRERIGLVKCREVNDFHHAHDALLAAEIGRFIQ KRHPGMYTNPIGYTQVMKQFVKNESDAVRRGHEPGTSPFVIASFMTSG FDEETGELFKDDWSASYEIAKLKSYFNYRQCFISRMPEETSGAFWDQTIY SPRGGKKTPSIPLKKGLSTEKHGGYSGQNPAYFVLIAAYDSKGKPKRLFE GVPIAVAELIKSKKQSIEEWAKQLVESRGFVFGQVLRARLLKYARIDHNG NEYYVPALDCVYSSRELILNENQTTIAAAVCDLARVSDEALVPSEVLELFD ALTEKCLILCTRFKPVYEKLVEGRERFVALSMQDKRKQIVGILKFFNCVTA RVDLSLIGGSPHAGGITNGFLQKPNDSFDIVDQSITGMFERRTHVGL AHWY Type II MGKPIPNPLLGLDSTKRTADGSEFESPKKKRKVGSNLRNVGEYNIGLDIG 15 Cas TGSVGWAVIDSDGEICHFKGKPTWGSRLFPSALPASEARVHRGQRRRY mammalian TRRKWRLFLLQELFAGAIEANDPEFFIRLNQSHLLPEDRQSDHKDYLFTL expression FNDSSFDEKAYFKKFPTIYHLRKWLMETDEKADPRLIYLAFHNIVKHRGN construct FLQQDNTGLNSQNANVRESVEELCAQLQEYCDSLDIPCTVEDNVDKIVQ (with N- VLSDTHSVRSHLQEEVQKLLGIGVCDLLDKTSAKKMAKALSGALVGLSAE terminal LADVFFLVIEKPEDAKTKIYLSKDEDVESFEEICPDEALPLFEAMNKVYSS methionine, FVLQEILSSCPGKTLSVNKVKDYEQYGEDLKLLKALVKEYAPEKYDDFFR stop codon, GEFYRPSSLHPQKYVYNKSVAKGYTRYNEVRGVAYEDFKKDVEKLFRD V5 tag, N- TPAVEDARYLDMMERFKEEKFLRRLKTSDNGAIPFQLHLEEMDKIIQNQA terminal NLS RFYPFLEEKKKELDSLVTFRIPYYVGPLTQKNARKDRKGSNRFSWSERIE and C- GKEHEVIKPWNWEEIIDKDRSATEFILRMTGTCTYLQGEPVLPRCSLLYE terminal NLS) EYCVLNELNGAHFTQDGDKEHRFDYQDRSDMLKELFRKGKVSYKKAAD (aa) WMVQRGNSNVRVSGGQGEKGFESKLGSYIFFAKDVFGVDEIPESDYPM IEEIILWNTIFEDRDILKEKLKRNYGDCLSDEQIKKIVRKRFTGWGRLSKKL LCGIKVETDNGRMSIMDVLREGNPNNNGLSKAMVFMEIIRDTGLGFDKRI DELNKEAMIGSTGLAIEDLPGSPALRRGVNQALGIVEEIVHIAGHAPNNIFI EVTREEDGRKKGQRTKKRYESLKEAMSKLKKESPEFWDAATSSQLAEK AKESAALDEKLTLYFLQGGKSLYSGKPLEIGRLSEYQVDHIIPQSYIKDDS FENKALVLASENQNKSDQMLLPVEMRRKMRPYWDALLSAGLIGKKKHT NLLRDHIGEKQMKGFIARQLVETSQILKIVQSFLEEQYPETNTLPVKAGLS HELRERIGLVKCREVNDFHHAHDALLAAEIGRFIQKRHPGMYTNPIGYTQ VMKQFVKNESDAVRRGHEPGTSPFVIASFMTSGFDEETGELFKDDWSA SYEIAKLKSYFNYRQCFISRMPEETSGAFWDQTIYSPRGGKKTPSIPLKK GLSTEKHGGYSGQNPAYFVLIAAYDSKGKPKRLFEGVPIAVAELIKSKKQ SIEEWAKQLVESRGFVFGQVLRARLLKYARIDHNGNEYYVPALDCVYSS RELILNENQTTIAAAVCDLARVSDEALVPSEVLELFDALTEKCLILCTRFKP VYEKLVEGRERFVALSMQDKRKQIVGILKFFNCVTARVDLSLIGGSPHAG GITNGFLQKPNDSFDIVDQSITGMFERRTHVGLSRKRTADGSEFESPKKK RKV AHWY Type II ATGAATTTAAGAAATGTCGGAGAGTATAATATCGGTCTCGATATTGGT 16 Cas coding ACGGGATCGGTTGGATGGGCTGTCATTGATTCTGATGGCGAGATCTG sequence (nt) CCATTTTAAAGGAAAACCAACATGGGGTAGCCGCTTGTTCCCATCTG (not codon CACTTCCTGCGTCAGAAGCTCGAGTGCACCGTGGGCAACGACGTCG optimized) CTACACCCGTCGTAAATGGCGTTTATTTCTTTTGCAAGAGTTGTTTGC AGGCGCGATCGAAGCAAACGATCCTGAATTCTTCATTCGCTTAAATCA GTCTCATCTGCTTCCTGAAGACCGTCAGTCAGATCATAAAGACTATCT CTTTACGCTCTTCAACGATTCATCGTTTGACGAAAAGGCATATTTCAA GAAATTTCCAACTATCTATCATCTTCGCAAATGGCTTATGGAGACAGA CGAGAAGGCAGATCCGCGTCTTATCTATCTTGCTTTCCACAACATTGT CAAGCACAGAGGGAATTTCCTTCAGCAAGATAATACAGGATTGAATTC GCAAAACGCCAATGTGCGAGAGTCTGTTGAAGAACTCTGCGCGCAGC TTCAAGAGTATTGCGATAGTCTCGATATTCCTTGCACGGTGGAAGATA ACGTCGATAAGATCGTTCAGGTACTGAGTGATACCCATTCAGTTCGCT CTCATCTTCAAGAAGAGGTTCAAAAGCTTCTTGGAATCGGTGTTTGTG ATCTTCTTGATAAAACATCGGCAAAGAAGATGGCGAAAGCCCTTTCAG TABLE 1C AHWY Type II Cas Sequences Name Sequence SEQ ID NO GTGCATTGGTGGGACTCTCAGCAGAGCTGGCAGATGTGTTCTTCTTG GTGATAGAGAAGCCAGAGGATGCCAAAACGAAGATATATCTTTCAAA GGATGAGGATGTTGAATCGTTCGAAGAGATCTGTCCAGACGAAGCGC TCCCGCTTTTTGAAGCAATGAACAAGGTTTATTCATCATTCGTTTTGCA GGAGATCTTGAGTAGTTGTCCAGGAAAGACGCTGTCTGTAAACAAGG TCAAAGATTACGAGCAGTATGGTGAAGACCTCAAGCTCCTGAAGGCT CTTGTGAAAGAGTATGCTCCAGAAAAGTACGATGATTTCTTCAGAGGG GAGTTCTACAGGCCAAGCAGCCTGCATCCTCAAAAGTACGTGTACAA CAAGTCAGTAGCAAAAGGGTATACCCGCTACAACGAAGTACGTGGGG TTGCTTATGAAGATTTCAAAAAGGATGTTGAAAAGCTCTTTCGCGATA CACCGGCCGTTGAGGATGCTCGTTACCTTGATATGATGGAGCGATTC AAAGAGGAGAAGTTCCTTCGAAGGCTTAAGACAAGCGACAATGGGGC GATACCCTTTCAACTGCATCTTGAGGAAATGGACAAGATCATTCAGAA TCAAGCACGTTTTTATCCATTCTTGGAAGAGAAGAAAAAAGAGCTTGA TTCGCTCGTGACGTTCCGTATCCCTTATTATGTGGGTCCTCTCACGCA GAAGAACGCACGCAAAGATCGCAAAGGTAGCAATCGGTTTTCGTGGT CGGAGCGCATCGAAGGAAAAGAGCACGAAGTTATCAAACCGTGGAAT TGGGAAGAGATCATCGATAAGGACAGGAGCGCAACCGAGTTCATATT GCGCATGACCGGCACCTGCACCTATCTTCAAGGTGAGCCAGTGCTTC CGCGTTGTTCTCTGCTCTATGAGGAATATTGCGTACTGAACGAACTTA ATGGTGCCCACTTCACACAAGATGGTGACAAAGAGCATCGTTTCGAT TATCAAGATCGTTCAGATATGCTGAAAGAACTCTTCCGAAAAGGAAAG GTGTCGTACAAGAAGGCTGCTGACTGGATGGTGCAGCGCGGCAATT CGAATGTACGTGTTTCGGGCGGCCAAGGCGAAAAGGGTTTCGAGTC GAAGTTAGGATCTTACATCTTCTTTGCAAAAGACGTCTTTGGAGTCGA CGAGATACCTGAGTCTGACTATCCCATGATAGAGGAGATCATTCTTTG GAATACGATCTTTGAAGATCGAGACATCCTTAAGGAAAAGCTAAAGCG CAACTATGGTGATTGTTTGTCGGATGAGCAGATCAAGAAGATCGTTC GAAAGCGCTTTACAGGATGGGGTCGCCTATCGAAGAAGCTTCTTTGT GGGATAAAAGTTGAGACCGATAATGGGCGAATGTCCATTATGGATGT TTTGCGTGAAGGAAATCCGAACAATAATGGATTATCGAAAGCGATGGT CTTTATGGAGATCATCAGAGACACTGGTCTCGGTTTCGATAAGCGAAT CGATGAACTCAACAAAGAAGCGATGATCGGTTCAACCGGTCTTGCTA TCGAGGATCTTCCTGGGTCCCCTGCGCTTCGTCGTGGAGTGAATCAG GCTCTTGGTATCGTGGAAGAGATCGTCCATATTGCTGGGCATGCACC GAACAATATCTTCATCGAGGTAACCCGTGAGGAAGATGGACGAAAAA AGGGACAGAGGACGAAGAAGCGCTATGAGAGCCTCAAAGAGGCTAT GAGCAAGCTGAAGAAAGAATCTCCGGAATTCTGGGATGCAGCAACTT CATCTCAGCTTGCAGAGAAAGCGAAAGAATCGGCTGCTCTTGATGAG AAGCTCACCCTGTATTTCTTGCAAGGCGGAAAATCTCTCTATTCAGGG AAACCGCTTGAGATTGGTCGCCTCTCCGAATATCAGGTCGATCACAT CATTCCGCAGTCGTACATCAAGGATGATAGTTTTGAGAATAAAGCACT TGTTCTTGCGTCGGAGAACCAAAACAAATCTGATCAGATGCTCCTTCC TGTGGAAATGCGTCGCAAGATGCGCCCATATTGGGATGCTCTTTTGA GCGCTGGTCTTATTGGGAAGAAGAAGCACACGAACCTTCTTCGTGAT CATATTGGCGAAAAGCAGATGAAAGGCTTCATAGCGCGTCAGCTTGT AGAAACGAGCCAGATATTGAAGATTGTTCAGAGCTTCTTGGAGGAGC AGTATCCCGAAACGAATACTCTTCCTGTAAAAGCGGGGCTTTCGCAT GAGTTGCGTGAGCGCATTGGTTTGGTGAAGTGCCGCGAAGTGAACG ATTTCCATCATGCTCACGATGCGCTTTTGGCTGCAGAGATCGGCCGT TTCATCCAAAAACGGCATCCTGGTATGTATACGAATCCGATCGGCTAT ACGCAGGTGATGAAGCAATTCGTCAAGAATGAAAGCGATGCTGTTAG AAGAGGCCACGAGCCTGGTACTTCGCCATTCGTTATAGCAAGCTTCA TGACTTCAGGGTTCGATGAAGAGACCGGAGAACTTTTCAAAGATGAC TGGTCGGCATCGTATGAGATTGCAAAGCTAAAGAGCTATTTCAACTAT CGCCAGTGTTTCATATCGAGAATGCCGGAAGAAACATCGGGCGCCTT TTGGGATCAAACCATCTATTCACCACGCGGCGGGAAGAAAACACCAA TABLE 1C AHWY Type II Cas Sequences Name Sequence SEQ ID NO GTATTCCACTGAAAAAAGGACTCTCTACCGAGAAGCATGGAGGATATT CAGGTCAAAATCCTGCTTACTTTGTCTTGATTGCTGCATATGATTCAAA AGGGAAACCAAAAAGACTATTTGAAGGAGTTCCTATTGCTGTTGCCGA GCTAATCAAGAGCAAAAAGCAGTCAATAGAAGAGTGGGCGAAACAGC TGGTCGAATCACGTGGGTTTGTTTTTGGCCAAGTTTTAAGAGCCCGAT TGCTTAAATATGCGCGCATTGATCATAACGGGAATGAATACTATGTTC CAGCGCTAGATTGCGTTTACTCGTCACGTGAGTTGATCTTGAACGAAA ACCAGACTACAATTGCTGCAGCAGTTTGCGATCTTGCTCGTGTTTCG GATGAAGCTTTAGTGCCATCCGAAGTACTAGAGCTTTTTGATGCCCTA ACAGAGAAATGCCTGATTCTTTGTACACGATTCAAGCCAGTCTATGAA AAGCTTGTAGAAGGAAGAGAGCGATTCGTAGCTCTATCGATGCAAGA TAAGCGAAAGCAAATCGTTGGAATACTGAAGTTCTTCAATTGCGTTAC AGCTCGTGTTGATTTGTCCTTAATAGGTGGATCGCCACATGCTGGAG GAATTACGAATGGCTTTTTACAAAAGCCCAATGACTCTTTTGATATCGT TGATCAGTCTATAACAGGCATGTTTGAAAGGCGGACTCACGTTGGCC TTTAG AHWY Type II AACCTGCGGAACGTGGGAGAGTACAACATCGGCCTTGACATCGGCA 17 Cas coding CCGGAAGCGTGGGATGGGCAGTGATTGACTCCGACGGCGAGATCTG sequence (nt) CCACTTCAAGGGCAAGCCTACCTGGGGGAGCAGACTGTTCCCTAGC (human GCCCTCCCTGCCTCTGAGGCCAGAGTGCATAGAGGCCAGAGAAGAA codon- GATACACACGTAGAAAGTGGCGGCTGTTCCTGCTGCAAGAGCTGTTC optimized; GCCGGCGCCATCGAGGCGAATGACCCTGAGTTCTTCATCAGACTGAA lacking N- CCAGAGCCACCTGCTCCCTGAGGATCGGCAGTCTGATCACAAGGACT terminal ACCTGTTTACCCTGTTTAACGACAGCTCTTTCGACGAGAAAGCCTATT methionine; TCAAAAAATTCCCAACAATCTATCACCTCAGAAAATGGCTGATGGAAA no stop CAGATGAAAAAGCCGATCCTAGACTGATCTATCTGGCCTTCCACAACA codon) TCGTGAAGCACAGAGGAAATTTCCTGCAGCAGGACAATACCGGCCTG AACAGCCAGAACGCCAACGTGAGGGAGTCTGTTGAAGAGCTGTGCG CTCAGCTGCAGGAGTATTGCGACTCGCTGGATATCCCCTGCACCGTG GAGGATAATGTGGATAAGATCGTGCAGGTGCTGTCTGATACCCACAG CGTGCGGAGCCACCTGCAGGAGGAGGTGCAGAAGCTGCTGGGAATC GGTGTCTGCGACCTGCTTGATAAGACATCCGCAAAGAAGATGGCCAA GGCCCTGAGCGGCGCCCTGGTGGGCCTGAGCGCTGAACTGGCCGA TGTGTTTTTCCTGGTGATCGAAAAACCCGAAGATGCCAAGACGAAGA TCTACCTGAGCAAGGATGAGGACGTGGAGAGCTTCGAAGAGATCTGC CCTGATGAGGCTCTTCCACTGTTCGAAGCCATGAACAAGGTGTACAG CAGCTTTGTGCTGCAGGAGATCCTGAGCTCATGCCCCGGCAAGACAC TGTCTGTGAACAAGGTGAAGGACTACGAGCAGTATGGCGAGGACCT GAAGCTGCTGAAGGCCCTGGTCAAGGAATACGCCCCTGAAAAGTAC GATGATTTCTTTCGGGGCGAGTTCTACAGACCTAGCTCTCTGCACCC GCAGAAATACGTGTACAACAAAAGCGTGGCTAAGGGGTACACCCGGT ACAACGAGGTGCGGGGCGTGGCCTACGAGGATTTCAAGAAGGACGT GGAAAAGCTGTTTAGAGACACACCTGCTGTGGAAGATGCTAGATACC TGGACATGATGGAAAGATTCAAGGAGGAAAAATTCCTGCGCAGACTG AAAACAAGCGACAATGGAGCTATTCCTTTCCAGCTGCACCTCGAAGA GATGGACAAAATCATCCAGAACCAGGCCAGATTCTACCCTTTTCTGGA GGAGAAGAAGAAGGAACTGGACTCTCTGGTGACCTTCAGAATCCCTT ACTACGTGGGCCCTCTGACCCAGAAGAACGCCAGAAAGGACAGAAA AGGCAGCAACAGATTCAGCTGGTCCGAGCGGATCGAGGGAAAGGAA CACGAGGTGATCAAGCCTTGGAATTGGGAGGAGATCATCGACAAGGA CCGGTCGGCCACAGAGTTCATCCTGAGAATGACAGGCACCTGTACCT ACCTGCAAGGCGAGCCTGTGCTGCCTAGATGCAGCCTGCTGTATGA GGAATACTGTGTGCTGAACGAGCTGAACGGCGCCCACTTCACCCAG GACGGCGATAAAGAGCACCGGTTTGACTACCAGGATAGAAGCGATAT GCTGAAAGAATTATTCAGAAAGGGAAAGGTGTCTTATAAGAAAGCTGC TGATTGGATGGTGCAGCGGGGCAATAGCAACGTGCGAGTGTCCGGC GGCCAGGGCGAGAAGGGATTTGAATCTAAACTGGGCAGCTACATCTT TABLE 1C AHWY Type II Cas Sequences Name Sequence SEQ ID NO CTTCGCCAAGGATGTATTCGGAGTGGACGAGATCCCTGAGAGCGATT ACCCTATGATCGAGGAAATTATCCTGTGGAACACCATCTTCGAGGATC GAGACATCCTGAAGGAGAAGTTAAAGCGCAACTACGGCGACTGTCTG TCTGACGAGCAGATCAAAAAAATCGTGCGCAAACGCTTCACAGGCTG GGGCAGACTCAGCAAGAAGCTCCTGTGCGGCATTAAGGTGGAAACC GACAACGGCAGAATGAGCATCATGGACGTGCTGCGCGAGGGCAACC CCAACAACAACGGCCTGTCTAAGGCTATGGTGTTTATGGAAATCATCA GGGATACCGGCCTGGGCTTCGACAAGAGAATCGACGAGCTGAACAA GGAAGCCATGATCGGATCTACCGGCCTCGCCATCGAGGACCTGCCC GGCAGTCCAGCCCTAAGACGGGGCGTGAACCAAGCCCTGGGCATCG TGGAAGAGATCGTGCACATAGCCGGACACGCCCCTAACAACATCTTC ATTGAAGTGACAAGAGAGGAAGATGGCCGGAAAAAGGGACAAAGAA CCAAGAAGCGGTACGAGTCCCTGAAGGAGGCTATGTCTAAACTGAAG AAAGAGAGCCCCGAGTTCTGGGATGCCGCCACCAGCAGCCAGCTGG CCGAAAAAGCCAAGGAGAGCGCCGCGCTCGACGAAAAGCTGACACT GTACTTCCTGCAGGGAGGCAAGAGCCTGTACTCTGGCAAGCCCCTG GAAATCGGCCGCCTGAGCGAATACCAGGTTGATCATATCATCCCCCA GAGCTATATCAAGGACGATAGCTTCGAGAACAAGGCCCTGGTACTCG CCAGCGAGAACCAAAATAAGAGCGACCAGATGCTGTTACCAGTGGAA ATGCGGAGAAAGATGCGGCCTTACTGGGACGCCCTGCTGAGCGCCG GCCTCATCGGGAAGAAGAAGCATACCAACCTGCTGCGGGACCACAT CGGAGAGAAGCAGATGAAGGGCTTCATCGCAAGACAGCTGGTCGAA ACCTCTCAGATTCTTAAGATCGTGCAGAGCTTCCTGGAAGAACAGTAC CCCGAAACCAACACCCTGCCCGTCAAGGCCGGCTTGTCTCACGAACT GAGAGAGAGAATCGGCCTGGTCAAGTGCAGAGAGGTGAACGACTTC CACCACGCCCACGACGCTCTCCTGGCAGCAGAAATCGGCCGGTTTAT CCAGAAGAGACACCCTGGCATGTACACCAACCCCATCGGCTACACAC AAGTGATGAAGCAGTTCGTGAAAAACGAGTCCGACGCCGTTAGAAGA GGCCACGAGCCTGGAACCAGCCCCTTCGTGATCGCCAGCTTCATGA CAAGCGGGTTCGACGAAGAAACCGGCGAGCTGTTCAAAGACGACTG GTCCGCCAGCTACGAGATCGCCAAACTCAAGTCTTATTTTAACTACCG ACAGTGCTTCATAAGCAGAATGCCTGAAGAAACCAGCGGCGCTTTCT GGGACCAAACCATCTACAGCCCTCGGGGCGGCAAGAAAACCCCTTC TATCCCCCTGAAGAAAGGCCTGAGCACCGAGAAGCACGGCGGCTAC AGCGGACAGAATCCTGCATACTTCGTGCTGATCGCCGCCTACGACAG CAAGGGCAAGCCTAAGCGGCTGTTCGAGGGCGTGCCTATCGCCGTG GCCGAGCTCATCAAGAGTAAGAAACAGAGCATCGAGGAGTGGGCCA AGCAGCTGGTGGAAAGCCGGGGCTTCGTGTTCGGCCAGGTGCTGAG AGCCAGACTGCTGAAGTACGCCAGAATCGATCACAACGGCAACGAGT ACTACGTGCCTGCCCTGGACTGTGTGTACAGCAGCCGGGAACTGATC CTGAACGAGAATCAGACCACAATCGCCGCTGCCGTGTGTGACCTGG CCAGAGTGAGCGACGAGGCCCTGGTGCCTAGCGAAGTGCTGGAACT GTTTGATGCTCTGACCGAGAAGTGCCTGATCCTGTGCACCCGGTTCA AGCCAGTGTACGAGAAGCTGGTAGAAGGCAGAGAAAGATTTGTTGCC CTGTCCATGCAGGATAAGAGAAAGCAGATCGTGGGCATCCTGAAATT TTTCAACTGCGTGACCGCCAGAGTGGACCTGTCCCTGATCGGCGGCT CTCCACACGCCGGCGGAATCACCAATGGCTTTCTGCAAAAGCCTAAC GACAGCTTCGACATCGTTGACCAGAGCATCACAGGCATGTTTGAGCG GCGAACACACGTGGGACTC AHWY Type II ATGGGCAAACCTATCCCCAATCCCCTGCTGGGCCTGGACAGCACCAA 18 Cas GCGGACTGCCGACGGCAGTGAGTTCGAGTCTCCCAAGAAAAAGAGG mammalian AAAGTGGGATCCAACCTGCGGAACGTGGGAGAGTACAACATCGGCC expression TTGACATCGGCACCGGAAGCGTGGGATGGGCAGTGATTGACTCCGA construct CGGCGAGATCTGCCACTTCAAGGGCAAGCCTACCTGGGGGAGCAGA (with N- CTGTTCCCTAGCGCCCTCCCTGCCTCTGAGGCCAGAGTGCATAGAG terminal GCCAGAGAAGAAGATACACACGTAGAAAGTGGCGGCTGTTCCTGCTG methionine, CAAGAGCTGTTCGCCGGCGCCATCGAGGCGAATGACCCTGAGTTCTT TABLE 1C AHWY Type II Cas Sequences Name Sequence SEQ ID NO stop codon, CATCAGACTGAACCAGAGCCACCTGCTCCCTGAGGATCGGCAGTCTG V5 tag, N- ATCACAAGGACTACCTGTTTACCCTGTTTAACGACAGCTCTTTCGACG terminal NLS AGAAAGCCTATTTCAAAAAATTCCCAACAATCTATCACCTCAGAAAAT and C- GGCTGATGGAAACAGATGAAAAAGCCGATCCTAGACTGATCTATCTG terminal NLS) GCCTTCCACAACATCGTGAAGCACAGAGGAAATTTCCTGCAGCAGGA (nt) CAATACCGGCCTGAACAGCCAGAACGCCAACGTGAGGGAGTCTGTT GAAGAGCTGTGCGCTCAGCTGCAGGAGTATTGCGACTCGCTGGATAT CCCCTGCACCGTGGAGGATAATGTGGATAAGATCGTGCAGGTGCTGT CTGATACCCACAGCGTGCGGAGCCACCTGCAGGAGGAGGTGCAGAA GCTGCTGGGAATCGGTGTCTGCGACCTGCTTGATAAGACATCCGCAA AGAAGATGGCCAAGGCCCTGAGCGGCGCCCTGGTGGGCCTGAGCG CTGAACTGGCCGATGTGTTTTTCCTGGTGATCGAAAAACCCGAAGAT GCCAAGACGAAGATCTACCTGAGCAAGGATGAGGACGTGGAGAGCT TCGAAGAGATCTGCCCTGATGAGGCTCTTCCACTGTTCGAAGCCATG AACAAGGTGTACAGCAGCTTTGTGCTGCAGGAGATCCTGAGCTCATG CCCCGGCAAGACACTGTCTGTGAACAAGGTGAAGGACTACGAGCAG TATGGCGAGGACCTGAAGCTGCTGAAGGCCCTGGTCAAGGAATACG CCCCTGAAAAGTACGATGATTTCTTTCGGGGCGAGTTCTACAGACCT AGCTCTCTGCACCCGCAGAAATACGTGTACAACAAAAGCGTGGCTAA GGGGTACACCCGGTACAACGAGGTGCGGGGCGTGGCCTACGAGGA TTTCAAGAAGGACGTGGAAAAGCTGTTTAGAGACACACCTGCTGTGG AAGATGCTAGATACCTGGACATGATGGAAAGATTCAAGGAGGAAAAA TTCCTGCGCAGACTGAAAACAAGCGACAATGGAGCTATTCCTTTCCA GCTGCACCTCGAAGAGATGGACAAAATCATCCAGAACCAGGCCAGAT TCTACCCTTTTCTGGAGGAGAAGAAGAAGGAACTGGACTCTCTGGTG ACCTTCAGAATCCCTTACTACGTGGGCCCTCTGACCCAGAAGAACGC CAGAAAGGACAGAAAAGGCAGCAACAGATTCAGCTGGTCCGAGCGG ATCGAGGGAAAGGAACACGAGGTGATCAAGCCTTGGAATTGGGAGG AGATCATCGACAAGGACCGGTCGGCCACAGAGTTCATCCTGAGAATG ACAGGCACCTGTACCTACCTGCAAGGCGAGCCTGTGCTGCCTAGATG CAGCCTGCTGTATGAGGAATACTGTGTGCTGAACGAGCTGAACGGCG CCCACTTCACCCAGGACGGCGATAAAGAGCACCGGTTTGACTACCAG GATAGAAGCGATATGCTGAAAGAATTATTCAGAAAGGGAAAGGTGTC TTATAAGAAAGCTGCTGATTGGATGGTGCAGCGGGGCAATAGCAACG TGCGAGTGTCCGGCGGCCAGGGCGAGAAGGGATTTGAATCTAAACT GGGCAGCTACATCTTCTTCGCCAAGGATGTATTCGGAGTGGACGAGA TCCCTGAGAGCGATTACCCTATGATCGAGGAAATTATCCTGTGGAAC ACCATCTTCGAGGATCGAGACATCCTGAAGGAGAAGTTAAAGCGCAA CTACGGCGACTGTCTGTCTGACGAGCAGATCAAAAAAATCGTGCGCA AACGCTTCACAGGCTGGGGCAGACTCAGCAAGAAGCTCCTGTGCGG CATTAAGGTGGAAACCGACAACGGCAGAATGAGCATCATGGACGTGC TGCGCGAGGGCAACCCCAACAACAACGGCCTGTCTAAGGCTATGGT GTTTATGGAAATCATCAGGGATACCGGCCTGGGCTTCGACAAGAGAA TCGACGAGCTGAACAAGGAAGCCATGATCGGATCTACCGGCCTCGC CATCGAGGACCTGCCCGGCAGTCCAGCCCTAAGACGGGGCGTGAAC CAAGCCCTGGGCATCGTGGAAGAGATCGTGCACATAGCCGGACACG CCCCTAACAACATCTTCATTGAAGTGACAAGAGAGGAAGATGGCCGG AAAAAGGGACAAAGAACCAAGAAGCGGTACGAGTCCCTGAAGGAGG CTATGTCTAAACTGAAGAAAGAGAGCCCCGAGTTCTGGGATGCCGCC ACCAGCAGCCAGCTGGCCGAAAAAGCCAAGGAGAGCGCCGCGCTCG ACGAAAAGCTGACACTGTACTTCCTGCAGGGAGGCAAGAGCCTGTAC TCTGGCAAGCCCCTGGAAATCGGCCGCCTGAGCGAATACCAGGTTG ATCATATCATCCCCCAGAGCTATATCAAGGACGATAGCTTCGAGAACA AGGCCCTGGTACTCGCCAGCGAGAACCAAAATAAGAGCGACCAGAT GCTGTTACCAGTGGAAATGCGGAGAAAGATGCGGCCTTACTGGGAC GCCCTGCTGAGCGCCGGCCTCATCGGGAAGAAGAAGCATACCAACC TGCTGCGGGACCACATCGGAGAGAAGCAGATGAAGGGCTTCATCGC TABLE 1C AHWY Type II Cas Sequences Name Sequence SEQ ID NO AAGACAGCTGGTCGAAACCTCTCAGATTCTTAAGATCGTGCAGAGCT TCCTGGAAGAACAGTACCCCGAAACCAACACCCTGCCCGTCAAGGCC GGCTTGTCTCACGAACTGAGAGAGAGAATCGGCCTGGTCAAGTGCA GAGAGGTGAACGACTTCCACCACGCCCACGACGCTCTCCTGGCAGC AGAAATCGGCCGGTTTATCCAGAAGAGACACCCTGGCATGTACACCA ACCCCATCGGCTACACACAAGTGATGAAGCAGTTCGTGAAAAACGAG TCCGACGCCGTTAGAAGAGGCCACGAGCCTGGAACCAGCCCCTTCG TGATCGCCAGCTTCATGACAAGCGGGTTCGACGAAGAAACCGGCGA GCTGTTCAAAGACGACTGGTCCGCCAGCTACGAGATCGCCAAACTCA AGTCTTATTTTAACTACCGACAGTGCTTCATAAGCAGAATGCCTGAAG AAACCAGCGGCGCTTTCTGGGACCAAACCATCTACAGCCCTCGGGG CGGCAAGAAAACCCCTTCTATCCCCCTGAAGAAAGGCCTGAGCACCG AGAAGCACGGCGGCTACAGCGGACAGAATCCTGCATACTTCGTGCT GATCGCCGCCTACGACAGCAAGGGCAAGCCTAAGCGGCTGTTCGAG GGCGTGCCTATCGCCGTGGCCGAGCTCATCAAGAGTAAGAAACAGA GCATCGAGGAGTGGGCCAAGCAGCTGGTGGAAAGCCGGGGCTTCGT GTTCGGCCAGGTGCTGAGAGCCAGACTGCTGAAGTACGCCAGAATC GATCACAACGGCAACGAGTACTACGTGCCTGCCCTGGACTGTGTGTA CAGCAGCCGGGAACTGATCCTGAACGAGAATCAGACCACAATCGCC GCTGCCGTGTGTGACCTGGCCAGAGTGAGCGACGAGGCCCTGGTGC CTAGCGAAGTGCTGGAACTGTTTGATGCTCTGACCGAGAAGTGCCTG ATCCTGTGCACCCGGTTCAAGCCAGTGTACGAGAAGCTGGTAGAAGG CAGAGAAAGATTTGTTGCCCTGTCCATGCAGGATAAGAGAAAGCAGA TCGTGGGCATCCTGAAATTTTTCAACTGCGTGACCGCCAGAGTGGAC CTGTCCCTGATCGGCGGCTCTCCACACGCCGGCGGAATCACCAATG GCTTTCTGCAAAAGCCTAACGACAGCTTCGACATCGTTGACCAGAGC ATCACAGGCATGTTTGAGCGGCGAACACACGTGGGACTCTCTAGAAA GCGGACAGCAGACGGCTCCGAATTTGAAAGCCCTAAGAAAAAGAGAA AGGTGTGA [0082] In some embodiments an AHWY Type II Cas protein comprises an amino acid sequence of SEQ ID NO:13, SEQ ID NO:14 or SEQ ID NO:15. In some embodiments, an AHWY Type II Cas protein has nickase activity, for example resulting from one or more amino acid substitutions relative to the sequence of SEQ ID NO:13, SEQ ID NO:14, or SEQ ID NO:15. In some embodiments, the one or more amino acid substitutions providing nickase activity comprise a D14A substitution, wherein the position of the D14A substitution is defined with respect to the amino acid numbering of SEQ ID NO:14. In some embodiments, the one or more amino acid substitutions providing nickase activity comprise an H900A substitution, wherein the position of the H900A substitution is defined with respect to the amino acid numbering of SEQ ID NO:14. In some embodiments, an AHWY Type II Cas protein is catalytically inactive, for example due a D14A substitution in combination with an H900A substitution. 6.2.1.4. CBGI Type II Cas Proteins [0083] In one aspect, the disclosure provides CBGI Type II Cas proteins. CBGI Type II Cas proteins can be further classified as Type IIA Cas proteins. The CBGI Type II Cas proteins typically comprise an amino acid sequence that is at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 85%, at least 90%, or at least 95% identical to SEQ ID NO:19. In some embodiments, the CBGI Type II Cas proteins comprise an amino acid sequence that is at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical to SEQ ID NO:19. In some embodiments, a CBGI Type II Cas protein comprises an amino acid sequence that is identical to SEQ ID NO:19. [0084] Exemplary CBGI Type II Cas protein sequences and nucleotide sequences encoding exemplary CBGI Type II Cas proteins are set forth in Table 1D. TABLE 1D CBGI Type II Cas Sequences Name Sequence SEQ ID NO CBGI Type II KLRNIQGDYNIGLDLGTGSVGWAVTGVDGELLTQNGRPTLGSRVFPSGE 19 Cas amino TAADARLKRGQRRRYERRRWRLDLLQRFFENDMADVDRTFFIRLKQAR acid LLREDRDESCRDYHSPLFLTVDAERDYYKRFPTIYHLRVWLMTTEKKADL sequence REIYLALHNIVKHRGNFLHQDNPSLSASAANMEDSVEQLCLALEDRCITL (without N- DIPCSCDAASIQRVLEDSSLSRAEKSESVGKLFGLDKDAQKTMGRGISRA terminal IVGYKVDFAAVLGCEFEDSSFTLSDDEKVDAALAAIPDVAMSLFDAIREVY methionine) SAYVLLGILSSGNGAPISSGSLSSVSGRTISFCKVREYEVYRADLALLKSL VRTYAPSQYDAFFRGELIPGTCNYDPTKAKGYTRYDLVHKVAYADFFKE VKSLLDKTDAVTDERYKDMLGRFEEERFLRRLKTSDNGSIPYQLHLEEM DAILKNQGKYYPFLLENLDKIESLVSFRIPYYLGPLTQKNAALDHHGNPRF AWATRKPGMEDEPVYPWNWEEVIDKGHAAHTFIQRMTSDCTYLIGEGV LPRNSLLYEEFCVLNELNGARYSVDGDDWRRFDYADRMGIMDDLFRQR RSVTYKMVEDWMRANRGWARVHVRGGQGECKFESSLSAYRFFCKDVF KTDELSPSIIPMVETIILWSTLFEDRSILKEQLVRNFSDRLSSEQIKIICKKRL TGWGNLSEKFLAEVEAETDCGPRSIMDILREGSPVGGEQGRTMVLMEV LHDERLGFEDKIEEINAERIADAGRLEVVDLPGSPALRRTVNQAVRVVEEI VHIAGKPPVNIFIENTRDEDASKKGKRTKRRYDTIKEAVNAFKRENADLAR ELKEFKPADFDDERLALYFIQGGKSLYSKAPLDVTRLSEYEVDHIIPQSYI KDDSFENKALVLKSENQTKTNRLLLPQGVRVKMASYWQELHRCGLIGDK KLRNLMCSDVSERRVKGFIARQLVETSQIVKLTKMMLENRLPESRLVPIK ASLSHELREAKHYYKCREINDFHHAHDALLAAEIGRFLLLRHAGMYDNPI GYAHVVKDFVRVQADEAKRTGRLPGSAGFIVSSFLHSGFDKDTGEIFKD TWDAEFECERIRKYLNYRQVYLSRMPEETSGAFWDATIYSPRGKMKLSL PLKEGLDPSKYGGYSSEKYAYFFCYYAKDKKGKRLIDFAPVPVSRAAGG QVDIEAFGREVAEERGYAFESIARAKIAIKQLIEVDGCRLFITGADQVRSA VPLAFGQEETHLVERLLSGAPVGPRVRDSLFNQMMVGIRRFDRRLYDNL KLDDRSAAFERLGEEDKDLILKGLLALSSASNNMADMRPIGGAKTAGQL KIVFRNVLSNQGITFIDQSVTGMFERKTYIGL CBGI Type II MKLRNIQGDYNIGLDLGTGSVGWAVTGVDGELLTQNGRPTLGSRVFPS 20 Cas amino GETAADARLKRGQRRRYERRRWRLDLLQRFFENDMADVDRTFFIRLKQ acid ARLLREDRDESCRDYHSPLFLTVDAERDYYKRFPTIYHLRVWLMTTEKK sequence ADLREIYLALHNIVKHRGNFLHQDNPSLSASAANMEDSVEQLCLALEDRC ITLDIPCSCDAASIQRVLEDSSLSRAEKSESVGKLFGLDKDAQKTMGRGI SRAIVGYKVDFAAVLGCEFEDSSFTLSDDEKVDAALAAIPDVAMSLFDAIR EVYSAYVLLGILSSGNGAPISSGSLSSVSGRTISFCKVREYEVYRADLALL KSLVRTYAPSQYDAFFRGELIPGTCNYDPTKAKGYTRYDLVHKVAYADF FKEVKSLLDKTDAVTDERYKDMLGRFEEERFLRRLKTSDNGSIPYQLHLE EMDAILKNQGKYYPFLLENLDKIESLVSFRIPYYLGPLTQKNAALDHHGNP RFAWATRKPGMEDEPVYPWNWEEVIDKGHAAHTFIQRMTSDCTYLIGE GVLPRNSLLYEEFCVLNELNGARYSVDGDDWRRFDYADRMGIMDDLFR QRRSVTYKMVEDWMRANRGWARVHVRGGQGECKFESSLSAYRFFCK DVFKTDELSPSIIPMVETIILWSTLFEDRSILKEQLVRNFSDRLSSEQIKIICK KRLTGWGNLSEKFLAEVEAETDCGPRSIMDILREGSPVGGEQGRTMVL MEVLHDERLGFEDKIEEINAERIADAGRLEVVDLPGSPALRRTVNQAVRV VEEIVHIAGKPPVNIFIENTRDEDASKKGKRTKRRYDTIKEAVNAFKRENA DLARELKEFKPADFDDERLALYFIQGGKSLYSKAPLDVTRLSEYEVDHIIP QSYIKDDSFENKALVLKSENQTKTNRLLLPQGVRVKMASYWQELHRCGL IGDKKLRNLMCSDVSERRVKGFIARQLVETSQIVKLTKMMLENRLPESRL VPIKASLSHELREAKHYYKCREINDFHHAHDALLAAEIGRFLLLRHAGMY DNPIGYAHVVKDFVRVQADEAKRTGRLPGSAGFIVSSFLHSGFDKDTGEI FKDTWDAEFECERIRKYLNYRQVYLSRMPEETSGAFWDATIYSPRGKMK LSLPLKEGLDPSKYGGYSSEKYAYFFCYYAKDKKGKRLIDFAPVPVSRAA GGQVDIEAFGREVAEERGYAFESIARAKIAIKQLIEVDGCRLFITGADQVR TABLE 1D CBGI Type II Cas Sequences Name Sequence SEQ ID NO SAVPLAFGQEETHLVERLLSGAPVGPRVRDSLFNQMMVGIRRFDRRLYD NLKLDDRSAAFERLGEEDKDLILKGLLALSSASNNMADMRPIGGAKTAG QLKIVFRNVLSNQGITFIDQSVTGMFERKTYIGL CBGI Type II MGKPIPNPLLGLDSTKRTADGSEFESPKKKRKVGSKLRNIQGDYNIGLDL 21 Cas GTGSVGWAVTGVDGELLTQNGRPTLGSRVFPSGETAADARLKRGQRR mammalian RYERRRWRLDLLQRFFENDMADVDRTFFIRLKQARLLREDRDESCRDY expression HSPLFLTVDAERDYYKRFPTIYHLRVWLMTTEKKADLREIYLALHNIVKHR construct GNFLHQDNPSLSASAANMEDSVEQLCLALEDRCITLDIPCSCDAASIQRV (with N- LEDSSLSRAEKSESVGKLFGLDKDAQKTMGRGISRAIVGYKVDFAAVLG terminal CEFEDSSFTLSDDEKVDAALAAIPDVAMSLFDAIREVYSAYVLLGILSSGN methionine, GAPISSGSLSSVSGRTISFCKVREYEVYRADLALLKSLVRTYAPSQYDAF stop codon, FRGELIPGTCNYDPTKAKGYTRYDLVHKVAYADFFKEVKSLLDKTDAVTD V5 tag, N- ERYKDMLGRFEEERFLRRLKTSDNGSIPYQLHLEEMDAILKNQGKYYPFL terminal NLS LENLDKIESLVSFRIPYYLGPLTQKNAALDHHGNPRFAWATRKPGMEDE and C- PVYPWNWEEVIDKGHAAHTFIQRMTSDCTYLIGEGVLPRNSLLYEEFCVL terminal NLS) NELNGARYSVDGDDWRRFDYADRMGIMDDLFRQRRSVTYKMVEDWM (aa) RANRGWARVHVRGGQGECKFESSLSAYRFFCKDVFKTDELSPSIIPMVE TIILWSTLFEDRSILKEQLVRNFSDRLSSEQIKIICKKRLTGWGNLSEKFLA EVEAETDCGPRSIMDILREGSPVGGEQGRTMVLMEVLHDERLGFEDKIE EINAERIADAGRLEVVDLPGSPALRRTVNQAVRVVEEIVHIAGKPPVNIFIE NTRDEDASKKGKRTKRRYDTIKEAVNAFKRENADLARELKEFKPADFDD ERLALYFIQGGKSLYSKAPLDVTRLSEYEVDHIIPQSYIKDDSFENKALVLK SENQTKTNRLLLPQGVRVKMASYWQELHRCGLIGDKKLRNLMCSDVSE RRVKGFIARQLVETSQIVKLTKMMLENRLPESRLVPIKASLSHELREAKHY YKCREINDFHHAHDALLAAEIGRFLLLRHAGMYDNPIGYAHVVKDFVRVQ ADEAKRTGRLPGSAGFIVSSFLHSGFDKDTGEIFKDTWDAEFECERIRKY LNYRQVYLSRMPEETSGAFWDATIYSPRGKMKLSLPLKEGLDPSKYGGY SSEKYAYFFCYYAKDKKGKRLIDFAPVPVSRAAGGQVDIEAFGREVAEE RGYAFESIARAKIAIKQLIEVDGCRLFITGADQVRSAVPLAFGQEETHLVE RLLSGAPVGPRVRDSLFNQMMVGIRRFDRRLYDNLKLDDRSAAFERLGE EDKDLILKGLLALSSASNNMADMRPIGGAKTAGQLKIVFRNVLSNQGITFI DQSVTGMFERKTYIGLSRKRTADGSEFESPKKKRKV CBGI Type II ATGAAACTGCGCAATATTCAAGGTGACTACAACATTGGTCTTGACCTT 22 Cas coding GGAACCGGCTCTGTGGGGTGGGCGGTTACCGGTGTAGATGGCGAGC sequence (nt) TTCTCACGCAGAACGGTAGACCGACGTTGGGAAGTCGCGTTTTCCCT (not codon TCGGGCGAAACTGCTGCGGACGCTCGGCTCAAGCGCGGTCAGCGCC optimized) GTAGGTATGAGCGTCGTCGTTGGCGTCTGGATTTGCTGCAGCGCTTT TTCGAGAATGATATGGCTGATGTAGATCGGACTTTCTTTATCCGCCTT AAACAGGCTCGCCTCTTGCGTGAAGATAGAGATGAGTCGTGTCGCGA TTACCACAGCCCTCTCTTTCTCACCGTTGATGCGGAGAGAGACTATTA CAAACGCTTCCCCACAATTTACCACCTTCGTGTCTGGCTGATGACCAC CGAGAAAAAGGCCGACCTGCGGGAGATTTATCTCGCGCTGCACAACA TCGTCAAGCATCGAGGCAATTTCCTCCACCAAGACAATCCCAGCTTG AGCGCATCTGCTGCCAACATGGAAGATTCTGTTGAGCAGCTGTGTCT TGCGCTGGAGGATCGATGCATCACCCTGGACATTCCTTGCTCTTGCG ATGCGGCTTCTATCCAGCGGGTGCTCGAGGATTCGTCTCTCTCCCGC GCGGAGAAGTCGGAGTCGGTTGGCAAGCTGTTCGGCCTTGATAAGG ACGCCCAGAAGACGATGGGGAGGGGCATCTCTCGCGCGATTGTGGG CTATAAGGTTGATTTTGCAGCCGTGCTCGGATGCGAGTTCGAGGACT CGTCTTTCACCCTTTCCGACGATGAGAAGGTAGATGCTGCCCTTGCG GCAATTCCCGACGTTGCCATGAGCTTGTTCGATGCGATTCGAGAGGT ATATTCTGCATATGTGCTTTTGGGCATCTTGAGTTCCGGAAATGGTGC CCCTATCTCCTCGGGTTCTCTTTCATCTGTTTCTGGTAGAACGATTTC CTTTTGCAAGGTTCGGGAGTATGAGGTGTATAGAGCCGACCTCGCCC TGCTGAAGTCTCTTGTGCGTACGTATGCGCCCAGCCAGTATGACGCC TTCTTCCGTGGTGAGCTGATTCCGGGCACTTGCAACTATGACCCGAC AAAGGCCAAGGGCTATACGCGTTACGACTTGGTTCATAAGGTTGCAT TABLE 1D CBGI Type II Cas Sequences Name Sequence SEQ ID NO ATGCTGACTTCTTTAAAGAGGTCAAGAGCCTTCTCGATAAGACCGATG CCGTAACGGACGAACGTTATAAGGACATGCTGGGCCGCTTTGAGGA GGAGCGCTTCCTGCGGAGGCTCAAAACGAGCGATAACGGCAGCATC CCGTATCAACTCCATCTGGAGGAGATGGACGCGATTCTCAAGAATCA AGGCAAGTATTATCCGTTCCTGCTCGAGAACCTAGACAAAATCGAGT CGCTTGTCAGCTTCAGGATTCCATATTACTTGGGGCCTTTGACACAGA AAAACGCTGCTCTCGATCACCATGGCAATCCACGTTTTGCCTGGGCA ACTCGTAAGCCCGGTATGGAGGACGAGCCTGTGTATCCCTGGAATTG GGAGGAGGTTATCGACAAGGGTCATGCGGCGCATACGTTTATCCAGC GCATGACGAGCGATTGCACCTACTTGATAGGGGAGGGCGTTCTTCCT CGCAATTCTCTTTTGTATGAGGAGTTCTGTGTCCTCAATGAGCTTAAC GGCGCTCGGTATTCCGTTGATGGCGATGATTGGCGGCGCTTCGACTA TGCCGACCGTATGGGTATCATGGATGACCTGTTCAGGCAGCGTCGAA GTGTTACGTACAAAATGGTCGAAGATTGGATGCGCGCCAATCGCGGA TGGGCCCGTGTTCATGTCCGCGGTGGTCAGGGAGAGTGTAAATTCG AATCCTCCCTTTCGGCGTACCGCTTCTTTTGCAAAGACGTTTTCAAGA CCGACGAACTGTCGCCTTCGATAATTCCTATGGTTGAGACCATAATCC TGTGGAGTACCTTATTCGAGGATCGTTCGATTTTGAAAGAGCAGCTC GTCCGCAATTTCTCAGATCGATTGAGCTCCGAACAGATCAAGATTATC TGCAAAAAGCGTCTGACGGGCTGGGGCAACCTATCGGAGAAGTTCCT GGCTGAGGTCGAGGCTGAGACGGATTGCGGACCCCGCTCCATCATG GATATCCTTCGCGAAGGCAGCCCTGTAGGCGGTGAACAGGGTAGGA CCATGGTCCTTATGGAGGTGTTGCATGATGAACGCCTGGGCTTTGAA GATAAAATCGAAGAAATAAACGCTGAAAGAATCGCTGATGCCGGTCG CCTTGAGGTGGTTGACCTGCCTGGATCTCCTGCGCTGCGCAGGACG GTCAATCAGGCCGTCCGTGTTGTCGAGGAGATTGTCCATATAGCGGG CAAGCCTCCTGTGAACATCTTTATCGAGAACACGCGTGATGAAGATG CGTCGAAAAAGGGCAAGAGGACGAAGAGGCGCTACGATACGATAAA AGAGGCTGTTAATGCGTTTAAGAGGGAAAACGCTGATTTGGCGAGAG AATTGAAGGAGTTCAAGCCTGCCGATTTCGACGATGAGAGGCTGGCC TTGTATTTCATTCAGGGAGGTAAGAGCCTGTACTCCAAGGCTCCTCTT GACGTGACGAGGCTGTCTGAATATGAAGTCGATCACATCATTCCACA GTCCTACATCAAAGATGATAGCTTTGAAAACAAAGCCCTCGTTCTAAA GAGTGAAAATCAGACCAAGACGAATCGGCTATTGTTGCCTCAAGGGG TTCGCGTAAAGATGGCGAGCTATTGGCAAGAGCTGCACCGTTGCGG CCTTATAGGTGATAAGAAACTGAGAAACCTCATGTGCTCAGATGTCAG CGAGCGTCGGGTTAAGGGCTTTATCGCACGTCAGCTTGTTGAAACCT CGCAGATCGTCAAGTTGACGAAGATGATGCTCGAGAATCGCCTGCCT GAGTCGAGGCTGGTTCCCATCAAGGCGAGCCTGTCGCATGAGCTGC GCGAGGCCAAGCATTATTACAAGTGCCGTGAGATCAACGATTTTCAC CATGCGCACGATGCACTTCTCGCTGCGGAGATCGGTAGGTTTTTGCT GCTGAGGCATGCTGGAATGTATGACAATCCCATCGGCTATGCGCATG TTGTTAAGGATTTTGTGCGCGTTCAAGCAGACGAGGCAAAGAGGACG GGCCGCCTGCCTGGATCGGCTGGCTTCATCGTGTCGAGCTTCCTGC ATTCGGGTTTCGACAAAGATACGGGCGAGATCTTCAAGGATACGTGG GACGCCGAGTTCGAATGTGAGCGCATTCGCAAGTATCTCAACTATCG GCAGGTGTACCTGAGCCGTATGCCGGAGGAGACATCTGGTGCGTTTT GGGATGCGACTATCTATTCGCCGCGTGGAAAGATGAAACTCTCCCTC CCGCTTAAAGAAGGGCTCGATCCGTCTAAATACGGCGGCTATTCATC GGAGAAATATGCATATTTCTTCTGCTATTACGCGAAGGATAAGAAGGG CAAGCGACTTATTGACTTTGCTCCAGTTCCGGTGTCTCGCGCTGCTG GGGGGCAAGTCGATATCGAGGCTTTCGGGCGTGAGGTTGCCGAAGA GAGGGGATATGCGTTCGAGTCTATCGCTCGTGCGAAGATAGCTATCA AGCAGCTCATTGAAGTTGACGGATGCAGGTTGTTTATCACCGGTGCC GACCAGGTCCGGAGTGCCGTTCCATTGGCCTTTGGCCAGGAAGAGA CGCATTTGGTTGAGAGGCTGCTTTCGGGCGCTCCGGTTGGTCCTCG CGTACGTGATTCCCTGTTCAATCAAATGATGGTCGGTATTAGGAGGTT TABLE 1D CBGI Type II Cas Sequences Name Sequence SEQ ID NO TGACAGGCGTCTGTATGACAACTTGAAGTTGGATGACCGATCGGCTG CGTTTGAGCGGTTGGGGGAAGAAGATAAGGACCTCATCCTGAAAGG GCTTCTTGCGTTGTCCTCTGCTTCAAACAACATGGCAGACATGCGGC CGATTGGAGGCGCGAAGACTGCAGGCCAGTTGAAAATCGTCTTTAGA AACGTACTGAGCAATCAGGGCATTACGTTTATCGATCAGTCTGTGACC GGCATGTTTGAGAGGAAAACTTACATTGGGCTTTAG CBGI Type II AAGCTGAGAAACATCCAGGGCGACTACAACATTGGCCTGGACCTGG 23 Cas coding GCACAGGCAGCGTGGGCTGGGCCGTGACCGGAGTGGATGGCGAAC sequence (nt) TGCTGACACAGAACGGCAGACCTACCCTGGGCTCAAGGGTCTTTCCT (human AGCGGCGAGACAGCCGCTGATGCCAGACTGAAGCGGGGACAGCGC codon- AGGCGGTATGAAAGAAGGCGCTGGCGGCTGGACCTGCTTCAGAGAT optimized; TCTTCGAGAACGACATGGCCGACGTGGACCGGACCTTCTTCATTAGA lacking N- CTGAAGCAGGCCAGACTGCTCCGGGAGGATAGAGATGAGAGCTGCA terminal GAGACTACCACAGTCCCCTGTTTCTCACCGTTGATGCAGAAAGAGAC methionine; TACTACAAGAGATTCCCTACAATCTACCACCTGAGAGTGTGGCTGATG no stop ACGACCGAGAAGAAGGCTGATCTGAGAGAGATCTACCTGGCCTTGCA codon) CAACATCGTGAAGCACAGAGGCAATTTCCTGCACCAGGACAATCCCA GCCTGAGCGCCTCAGCAGCTAATATGGAAGATTCTGTGGAGCAGCTC TGTCTGGCCTTGGAGGACAGATGCATCACCCTGGATATCCCTTGTTC TTGTGACGCCGCCAGCATCCAGAGAGTCCTGGAGGACAGCAGCCTG AGCCGGGCTGAGAAGAGCGAGTCTGTGGGCAAGCTGTTCGGCCTGG ACAAAGACGCCCAGAAAACAATGGGCAGAGGAATCTCTCGGGCGAT CGTGGGCTACAAGGTTGACTTCGCCGCTGTGCTGGGATGTGAGTTTG AGGACAGCTCTTTTACCCTGTCTGACGACGAAAAGGTGGACGCCGCC CTGGCCGCCATCCCGGACGTTGCAATGAGCTTGTTCGACGCCATCAG AGAGGTGTACAGCGCTTACGTGCTGCTGGGCATCCTGAGCAGCGGC AACGGCGCTCCAATCAGCTCCGGCTCTTTGTCCTCTGTGAGTGGCAG AACCATCAGCTTCTGCAAGGTGAGAGAATATGAGGTGTACCGGGCCG ACCTGGCCCTATTGAAATCACTGGTGCGCACCTACGCTCCTAGTCAG TACGACGCCTTCTTTAGAGGCGAGTTAATTCCTGGAACATGCAACTAC GACCCCACCAAAGCCAAGGGCTACACCAGATACGACCTGGTCCATAA GGTGGCCTACGCTGATTTCTTCAAGGAAGTGAAGTCCCTGCTCGATA AGACCGACGCCGTGACCGATGAGAGATACAAGGACATGCTGGGTAG ATTCGAAGAGGAGCGCTTCCTGCGGAGACTGAAAACCAGCGATAATG GCAGCATCCCTTATCAGCTGCACCTGGAAGAGATGGATGCCATCCTG AAGAATCAGGGCAAGTACTACCCTTTCCTGCTGGAAAACCTGGACAA GATCGAGTCCCTGGTGAGCTTTAGAATCCCTTACTACCTGGGCCCTC TGACCCAGAAGAACGCCGCTCTGGATCACCACGGAAACCCTAGATTC GCCTGGGCGACCCGGAAGCCCGGGATGGAGGACGAGCCTGTGTAC CCCTGGAACTGGGAGGAAGTGATCGACAAAGGCCACGCCGCCCATA CATTTATCCAGCGGATGACCAGCGACTGCACCTACTTGATCGGCGAG GGTGTGCTGCCCAGAAACAGCCTGCTGTACGAGGAATTTTGTGTGCT GAACGAGCTGAACGGCGCCCGGTACAGCGTGGATGGCGATGACTGG AGAAGATTCGATTACGCTGATAGGATGGGCATCATGGACGACCTGTT TCGGCAAAGAAGAAGTGTAACCTACAAGATGGTCGAGGATTGGATGC GGGCCAATAGAGGCTGGGCTAGAGTGCACGTGAGAGGCGGCCAGG GCGAGTGCAAGTTCGAGAGCAGCCTGTCCGCCTATAGATTCTTTTGT AAGGACGTGTTCAAGACAGACGAGCTGTCCCCTTCTATCATCCCTAT GGTGGAAACCATCATCCTGTGGTCGACCCTGTTCGAGGACAGATCTA TCCTCAAAGAGCAGCTGGTTAGAAATTTCAGCGACCGGCTGAGCTCC GAGCAAATCAAGATCATCTGCAAGAAGAGACTGACCGGCTGGGGCAA CCTTAGTGAAAAATTCCTGGCCGAAGTGGAAGCCGAAACCGACTGCG GCCCCCGGAGCATCATGGACATCCTGAGAGAGGGAAGCCCTGTGGG AGGAGAGCAGGGCAGAACAATGGTGCTGATGGAAGTGCTCCACGAT GAGAGACTGGGTTTCGAGGATAAAATCGAGGAGATCAACGCCGAGC GGATCGCTGACGCCGGCAGACTGGAAGTGGTGGATCTGCCCGGATC TCCCGCCCTGCGGAGAACCGTGAACCAGGCTGTGCGAGTCGTAGAG TABLE 1D CBGI Type II Cas Sequences Name Sequence SEQ ID NO GAAATCGTGCACATCGCCGGCAAACCACCTGTGAACATCTTTATTGA GAACACCCGGGACGAGGACGCCAGCAAGAAGGGCAAGCGGACCAA GCGGCGATACGATACCATCAAGGAAGCCGTGAACGCCTTCAAGCGG GAGAACGCCGACCTGGCCAGAGAACTGAAAGAGTTCAAGCCTGCGG ATTTCGATGATGAGCGGCTCGCTCTGTATTTCATTCAGGGCGGCAAG TCTCTGTACAGCAAGGCCCCTCTGGATGTGACAAGACTGAGCGAGTA CGAGGTGGACCACATCATCCCCCAAAGCTACATCAAAGACGACTCTT TCGAAAACAAGGCTCTGGTGCTCAAGTCCGAGAACCAGACCAAGACA AACAGACTGCTGCTGCCACAAGGAGTGAGAGTGAAGATGGCCTCCTA CTGGCAGGAACTGCATAGATGCGGCCTGATCGGCGACAAGAAGCTG AGAAACCTGATGTGCAGCGATGTGTCCGAACGGAGAGTGAAGGGCT TCATCGCCAGACAGCTGGTGGAAACAAGCCAGATCGTGAAGCTGACT AAGATGATGCTGGAGAATAGACTGCCTGAGTCTAGGCTGGTGCCCAT CAAGGCCAGCCTGAGCCACGAGCTGAGGGAGGCCAAGCACTATTAC AAGTGCCGGGAAATCAACGACTTCCACCACGCCCACGACGCCCTGC TGGCCGCCGAGATCGGACGGTTTCTGCTGCTGCGGCACGCCGGCAT GTACGATAACCCCATCGGCTACGCCCACGTGGTCAAAGACTTTGTGC GCGTGCAGGCCGACGAAGCCAAGAGAACCGGCCGACTGCCTGGCTC TGCCGGTTTCATCGTTTCCAGCTTCCTGCACAGCGGCTTCGACAAGG ATACGGGCGAGATCTTCAAGGATACATGGGACGCCGAGTTCGAATGC GAGAGAATCAGAAAGTACCTGAATTACAGACAGGTGTACCTGAGTAG AATGCCTGAGGAAACAAGCGGCGCCTTTTGGGACGCCACAATCTACA GCCCTCGGGGAAAAATGAAGCTGAGCCTACCTCTGAAGGAGGGCCT GGACCCTAGCAAGTACGGCGGATACAGCAGCGAGAAGTATGCTTACT TCTTTTGCTACTATGCTAAAGACAAGAAAGGCAAACGGCTCATCGACT TCGCCCCTGTGCCAGTGAGCCGGGCCGCCGGAGGACAGGTGGACA TCGAGGCCTTCGGCAGAGAAGTGGCCGAAGAGCGGGGCTATGCCTT CGAAAGCATCGCCCGAGCTAAGATCGCCATCAAGCAGCTGATCGAG GTCGACGGCTGCAGACTGTTCATCACCGGCGCCGACCAGGTTAGAA GCGCTGTGCCTCTGGCCTTCGGCCAAGAAGAAACCCACCTGGTGGA ACGGCTGCTGAGCGGCGCCCCCGTGGGACCTAGAGTGAGAGATAGC CTGTTCAACCAGATGATGGTCGGCATCAGACGGTTTGACAGGCGGCT GTACGACAACCTGAAGCTGGATGACAGAAGCGCCGCCTTTGAACGG CTGGGCGAGGAGGATAAGGATCTCATCCTCAAAGGCCTGCTGGCCC TGTCGAGCGCCTCCAACAACATGGCTGACATGCGGCCTATCGGCGG CGCCAAAACGGCCGGTCAACTGAAGATCGTGTTCCGGAATGTGCTGT CCAACCAGGGGATCACCTTCATCGATCAGAGCGTGACCGGCATGTTC GAACGTAAAACATACATTGGGTTA CBGI Type II ATGGGCAAACCTATCCCCAATCCCCTGCTGGGCCTGGACAGCACCAA 24 Cas GCGGACTGCCGACGGCAGTGAGTTCGAGTCTCCCAAGAAAAAGAGG mammalian AAAGTGGGATCCAAGCTGAGAAACATCCAGGGCGACTACAACATTGG expression CCTGGACCTGGGCACAGGCAGCGTGGGCTGGGCCGTGACCGGAGT construct GGATGGCGAACTGCTGACACAGAACGGCAGACCTACCCTGGGCTCA (with N- AGGGTCTTTCCTAGCGGCGAGACAGCCGCTGATGCCAGACTGAAGC terminal GGGGACAGCGCAGGCGGTATGAAAGAAGGCGCTGGCGGCTGGACC methionine, TGCTTCAGAGATTCTTCGAGAACGACATGGCCGACGTGGACCGGACC stop codon, TTCTTCATTAGACTGAAGCAGGCCAGACTGCTCCGGGAGGATAGAGA V5 tag, N- TGAGAGCTGCAGAGACTACCACAGTCCCCTGTTTCTCACCGTTGATG terminal NLS CAGAAAGAGACTACTACAAGAGATTCCCTACAATCTACCACCTGAGA and C- GTGTGGCTGATGACGACCGAGAAGAAGGCTGATCTGAGAGAGATCTA terminal NLS) CCTGGCCTTGCACAACATCGTGAAGCACAGAGGCAATTTCCTGCACC (nt) AGGACAATCCCAGCCTGAGCGCCTCAGCAGCTAATATGGAAGATTCT GTGGAGCAGCTCTGTCTGGCCTTGGAGGACAGATGCATCACCCTGG ATATCCCTTGTTCTTGTGACGCCGCCAGCATCCAGAGAGTCCTGGAG GACAGCAGCCTGAGCCGGGCTGAGAAGAGCGAGTCTGTGGGCAAGC TGTTCGGCCTGGACAAAGACGCCCAGAAAACAATGGGCAGAGGAAT CTCTCGGGCGATCGTGGGCTACAAGGTTGACTTCGCCGCTGTGCTG TABLE 1D CBGI Type II Cas Sequences Name Sequence SEQ ID NO GGATGTGAGTTTGAGGACAGCTCTTTTACCCTGTCTGACGACGAAAA GGTGGACGCCGCCCTGGCCGCCATCCCGGACGTTGCAATGAGCTTG TTCGACGCCATCAGAGAGGTGTACAGCGCTTACGTGCTGCTGGGCAT CCTGAGCAGCGGCAACGGCGCTCCAATCAGCTCCGGCTCTTTGTCCT CTGTGAGTGGCAGAACCATCAGCTTCTGCAAGGTGAGAGAATATGAG GTGTACCGGGCCGACCTGGCCCTATTGAAATCACTGGTGCGCACCTA CGCTCCTAGTCAGTACGACGCCTTCTTTAGAGGCGAGTTAATTCCTG GAACATGCAACTACGACCCCACCAAAGCCAAGGGCTACACCAGATAC GACCTGGTCCATAAGGTGGCCTACGCTGATTTCTTCAAGGAAGTGAA GTCCCTGCTCGATAAGACCGACGCCGTGACCGATGAGAGATACAAG GACATGCTGGGTAGATTCGAAGAGGAGCGCTTCCTGCGGAGACTGA AAACCAGCGATAATGGCAGCATCCCTTATCAGCTGCACCTGGAAGAG ATGGATGCCATCCTGAAGAATCAGGGCAAGTACTACCCTTTCCTGCT GGAAAACCTGGACAAGATCGAGTCCCTGGTGAGCTTTAGAATCCCTT ACTACCTGGGCCCTCTGACCCAGAAGAACGCCGCTCTGGATCACCAC GGAAACCCTAGATTCGCCTGGGCGACCCGGAAGCCCGGGATGGAGG ACGAGCCTGTGTACCCCTGGAACTGGGAGGAAGTGATCGACAAAGG CCACGCCGCCCATACATTTATCCAGCGGATGACCAGCGACTGCACCT ACTTGATCGGCGAGGGTGTGCTGCCCAGAAACAGCCTGCTGTACGA GGAATTTTGTGTGCTGAACGAGCTGAACGGCGCCCGGTACAGCGTG GATGGCGATGACTGGAGAAGATTCGATTACGCTGATAGGATGGGCAT CATGGACGACCTGTTTCGGCAAAGAAGAAGTGTAACCTACAAGATGG TCGAGGATTGGATGCGGGCCAATAGAGGCTGGGCTAGAGTGCACGT GAGAGGCGGCCAGGGCGAGTGCAAGTTCGAGAGCAGCCTGTCCGC CTATAGATTCTTTTGTAAGGACGTGTTCAAGACAGACGAGCTGTCCCC TTCTATCATCCCTATGGTGGAAACCATCATCCTGTGGTCGACCCTGTT CGAGGACAGATCTATCCTCAAAGAGCAGCTGGTTAGAAATTTCAGCG ACCGGCTGAGCTCCGAGCAAATCAAGATCATCTGCAAGAAGAGACTG ACCGGCTGGGGCAACCTTAGTGAAAAATTCCTGGCCGAAGTGGAAG CCGAAACCGACTGCGGCCCCCGGAGCATCATGGACATCCTGAGAGA GGGAAGCCCTGTGGGAGGAGAGCAGGGCAGAACAATGGTGCTGATG GAAGTGCTCCACGATGAGAGACTGGGTTTCGAGGATAAAATCGAGGA GATCAACGCCGAGCGGATCGCTGACGCCGGCAGACTGGAAGTGGTG GATCTGCCCGGATCTCCCGCCCTGCGGAGAACCGTGAACCAGGCTG TGCGAGTCGTAGAGGAAATCGTGCACATCGCCGGCAAACCACCTGT GAACATCTTTATTGAGAACACCCGGGACGAGGACGCCAGCAAGAAG GGCAAGCGGACCAAGCGGCGATACGATACCATCAAGGAAGCCGTGA ACGCCTTCAAGCGGGAGAACGCCGACCTGGCCAGAGAACTGAAAGA GTTCAAGCCTGCGGATTTCGATGATGAGCGGCTCGCTCTGTATTTCA TTCAGGGCGGCAAGTCTCTGTACAGCAAGGCCCCTCTGGATGTGACA AGACTGAGCGAGTACGAGGTGGACCACATCATCCCCCAAAGCTACAT CAAAGACGACTCTTTCGAAAACAAGGCTCTGGTGCTCAAGTCCGAGA ACCAGACCAAGACAAACAGACTGCTGCTGCCACAAGGAGTGAGAGT GAAGATGGCCTCCTACTGGCAGGAACTGCATAGATGCGGCCTGATC GGCGACAAGAAGCTGAGAAACCTGATGTGCAGCGATGTGTCCGAAC GGAGAGTGAAGGGCTTCATCGCCAGACAGCTGGTGGAAACAAGCCA GATCGTGAAGCTGACTAAGATGATGCTGGAGAATAGACTGCCTGAGT CTAGGCTGGTGCCCATCAAGGCCAGCCTGAGCCACGAGCTGAGGGA GGCCAAGCACTATTACAAGTGCCGGGAAATCAACGACTTCCACCACG CCCACGACGCCCTGCTGGCCGCCGAGATCGGACGGTTTCTGCTGCT GCGGCACGCCGGCATGTACGATAACCCCATCGGCTACGCCCACGTG GTCAAAGACTTTGTGCGCGTGCAGGCCGACGAAGCCAAGAGAACCG GCCGACTGCCTGGCTCTGCCGGTTTCATCGTTTCCAGCTTCCTGCAC AGCGGCTTCGACAAGGATACGGGCGAGATCTTCAAGGATACATGGG ACGCCGAGTTCGAATGCGAGAGAATCAGAAAGTACCTGAATTACAGA CAGGTGTACCTGAGTAGAATGCCTGAGGAAACAAGCGGCGCCTTTTG GGACGCCACAATCTACAGCCCTCGGGGAAAAATGAAGCTGAGCCTAC TABLE 1D CBGI Type II Cas Sequences Name Sequence SEQ ID NO CTCTGAAGGAGGGCCTGGACCCTAGCAAGTACGGCGGATACAGCAG CGAGAAGTATGCTTACTTCTTTTGCTACTATGCTAAAGACAAGAAAGG CAAACGGCTCATCGACTTCGCCCCTGTGCCAGTGAGCCGGGCCGCC GGAGGACAGGTGGACATCGAGGCCTTCGGCAGAGAAGTGGCCGAAG AGCGGGGCTATGCCTTCGAAAGCATCGCCCGAGCTAAGATCGCCAT CAAGCAGCTGATCGAGGTCGACGGCTGCAGACTGTTCATCACCGGC GCCGACCAGGTTAGAAGCGCTGTGCCTCTGGCCTTCGGCCAAGAAG AAACCCACCTGGTGGAACGGCTGCTGAGCGGCGCCCCCGTGGGACC TAGAGTGAGAGATAGCCTGTTCAACCAGATGATGGTCGGCATCAGAC GGTTTGACAGGCGGCTGTACGACAACCTGAAGCTGGATGACAGAAG CGCCGCCTTTGAACGGCTGGGCGAGGAGGATAAGGATCTCATCCTC AAAGGCCTGCTGGCCCTGTCGAGCGCCTCCAACAACATGGCTGACAT GCGGCCTATCGGCGGCGCCAAAACGGCCGGTCAACTGAAGATCGTG TTCCGGAATGTGCTGTCCAACCAGGGGATCACCTTCATCGATCAGAG CGTGACCGGCATGTTCGAACGTAAAACATACATTGGGTTATCTAGAAA GCGGACAGCAGACGGCTCCGAATTTGAAAGCCCTAAGAAAAAGAGAA AGGTGTGA [0085] In some embodiments a CBGI Type II Cas protein comprises an amino acid sequence of SEQ ID NO:19, SEQ ID NO:20, or SEQ ID NO:21. In some embodiments, a CBGI Type II Cas protein has nickase activity, for example resulting from one or more amino acid substitutions relative to the sequence of SEQ ID NO:19, SEQ ID NO:20, or SEQ ID NO:21. In some embodiments, the one or more amino acid substitutions providing nickase activity comprise a D15A substitution, wherein the position of the D15A substitution is defined with respect to the amino acid numbering of SEQ ID NO:20. In some embodiments, the one or more amino acid substitutions providing nickase activity comprise an H890A substitution, wherein the position of the H890A substitution is defined with respect to the amino acid numbering of SEQ ID NO:20. In some embodiments, a CBGI Type II Cas protein is catalytically inactive, for example due to a D15A substitution in combination with an H890A substitution. 6.2.1.5. ASDR Type II Cas Proteins [0086] In one aspect, the disclosure provides ASDR Type II Cas proteins. ASDR Type II Cas proteins can be further classified as Type IIA Cas proteins. The ASDR Type II Cas proteins typically comprise an amino acid sequence that is at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 85%, at least 90%, or at least 95% identical to SEQ ID NO:25. In some embodiments, the ASDR Type II Cas proteins comprise an amino acid sequence that is at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical to SEQ ID NO:25. In some embodiments, an ASDR Type II Cas protein comprises an amino acid sequence that is identical to SEQ ID NO:25. [0087] Exemplary ASDR Type II Cas protein sequences and nucleotide sequences encoding exemplary ASDR Type II Cas proteins are set forth in Table 1E. TABLE 1E ASDR Type II Cas Sequences Name Sequence SEQ ID NO ASDR Type II NLRNYPGSFNIGLDLGPASAGWSVVDSEGKLFHFKKKPTWGSRLFDSA 25 Cas amino QTASEARMHRGQRRRYVRRRWRLNLLQALFTEEMEKVDPGFFMRLNH acid SRTVEGDPIFTKDFTKKDYYKRFPTIYHLRAHLMETDDPADLRLVYLAIHN sequence IVKHRGNFLRQEEKLTAKIANTSEALGKLSVSLKEWCDSRGYSVGKVDE TABLE 1E ASDR Type II Cas Sequences Name Sequence SEQ ID NO (without N- EAIAAVLADKRISRSQKAKNVTLLISVDTGDSKANAKLAKALANAIVGLQA terminal EFKDLFGDFECEATKLNLSEEEKLEALQAACPDDSAELLEAVCGAYSAY methionine) VLQGLLSYAEGQTISHNMIKKYDCYAEDLECLKALVRDYLPKTKGDDGLK PFDRFFRGVTYHDLDKSDPSRDYDASKAKGYTAYNLHKLSYDEFRKEVE KLLKGTGAQDDERYITMMDAFEKQRFLRRLKTSDNGAIYYQLHLEELQAI LENQGRFYPFLKQEASKIESLVTFRIPYYVGPLTSKNAAKDAHNKNRFQW SERKPGMEDAIITPWNWESVIDKNRSAENFIMRMTGDCTYLAGEKVLPR HSLLYEEFCVLNELNGVRFTNDGDNWHRLDSEQRAGIIRDLFHKKRSVK YADIAAWLVRMGYLERTPQVRGGQGESGLESKMSSYLLFAKDVFGVDE LDEALFPAIEQIILWNTLFEDRSIFEEKLRDEFGPNGNGMLNNEQIKKCKK RLSGWGRLSEKFLTGIRVVTQGGRQMSIMNVLREGSPNPDRRQGEALV MMEVLHDNTLGFQKKIDEVNRAYYAQNAKAMGVNELPGSPGIRRSINQA IRIVDEIVKIAGHAPDNIFIEVTRDEDEKKKGKRTVKRWDRIEAALNAFKAE GGDLAVMSDFKRLKNTEADLDERLTLYLLQNGKCLYSGRAIDLDALMTG SGEYEVDHIIPRAYIKDDSLDNKALVYRSENQRKTDQLLIDENVRQKMGE TWKQLHDAKMISDKKYNNLMRLTVTEGAMKGFIARQLVETSQIVKMTKA LLEARYGDAGTRIVPVKASMSHNLREVAGLVKCREANDFHHAHDAYLAC RMGLFIQKRYPGIYENPIGYAHAMKKYVLEQSELFKKTHRMPGSAGLIVN SFMTAGFDPETGEMFEDAWAGNDEIKHCKKKHYDDAWNAAAEVEGIRR ALNYRQCYVSRMPYEDSGKFWDANPISKRSNPKLALKNTLDPACFGGY NTKTAAYFFIYEVTDKKGKTGLRIAQMPIFMKAKCESDNAAFEKYCQDLE SESRVSFTRIVIPRLLKWQLVEIDGNLLRINAEEEMKSASQLAFSQEELER ILLVCTDGYLDPDVNRSVYMAIAKALDRAKCPMLTNDLDLISRFDAFCGLA VADQKDLLKKILLLCKGNKQSSPINLSSVGGKKKAGYLRPARSKVPDGM VLIDQSVTGMFERRTRVGV ASDR Type II MNLRNYPGSFNIGLDLGPASAGWSVVDSEGKLFHFKKKPTWGSRLFDS 26 Cas amino AQTASEARMHRGQRRRYVRRRWRLNLLQALFTEEMEKVDPGFFMRLN acid HSRTVEGDPIFTKDFTKKDYYKRFPTIYHLRAHLMETDDPADLRLVYLAIH sequence NIVKHRGNFLRQEEKLTAKIANTSEALGKLSVSLKEWCDSRGYSVGKVD EEAIAAVLADKRISRSQKAKNVTLLISVDTGDSKANAKLAKALANAIVGLQ AEFKDLFGDFECEATKLNLSEEEKLEALQAACPDDSAELLEAVCGAYSA YVLQGLLSYAEGQTISHNMIKKYDCYAEDLECLKALVRDYLPKTKGDDGL KPFDRFFRGVTYHDLDKSDPSRDYDASKAKGYTAYNLHKLSYDEFRKEV EKLLKGTGAQDDERYITMMDAFEKQRFLRRLKTSDNGAIYYQLHLEELQ AILENQGRFYPFLKQEASKIESLVTFRIPYYVGPLTSKNAAKDAHNKNRFQ WSERKPGMEDAIITPWNWESVIDKNRSAENFIMRMTGDCTYLAGEKVLP RHSLLYEEFCVLNELNGVRFTNDGDNWHRLDSEQRAGIIRDLFHKKRSV KYADIAAWLVRMGYLERTPQVRGGQGESGLESKMSSYLLFAKDVFGVD ELDEALFPAIEQIILWNTLFEDRSIFEEKLRDEFGPNGNGMLNNEQIKKCK KRLSGWGRLSEKFLTGIRVVTQGGRQMSIMNVLREGSPNPDRRQGEAL VMMEVLHDNTLGFQKKIDEVNRAYYAQNAKAMGVNELPGSPGIRRSINQ AIRIVDEIVKIAGHAPDNIFIEVTRDEDEKKKGKRTVKRWDRIEAALNAFKA EGGDLAVMSDFKRLKNTEADLDERLTLYLLQNGKCLYSGRAIDLDALMT GSGEYEVDHIIPRAYIKDDSLDNKALVYRSENQRKTDQLLIDENVRQKMG ETWKQLHDAKMISDKKYNNLMRLTVTEGAMKGFIARQLVETSQIVKMTK ALLEARYGDAGTRIVPVKASMSHNLREVAGLVKCREANDFHHAHDAYLA CRMGLFIQKRYPGIYENPIGYAHAMKKYVLEQSELFKKTHRMPGSAGLIV NSFMTAGFDPETGEMFEDAWAGNDEIKHCKKKHYDDAWNAAAEVEGIR RALNYRQCYVSRMPYEDSGKFWDANPISKRSNPKLALKNTLDPACFGG YNTKTAAYFFIYEVTDKKGKTGLRIAQMPIFMKAKCESDNAAFEKYCQDL ESESRVSFTRIVIPRLLKWQLVEIDGNLLRINAEEEMKSASQLAFSQEELE RILLVCTDGYLDPDVNRSVYMAIAKALDRAKCPMLTNDLDLISRFDAFCG LAVADQKDLLKKILLLCKGNKQSSPINLSSVGGKKKAGYLRPARSKVPDG MVLIDQSVTGMFERRTRVGV ASDR Type II MGKPIPNPLLGLDSTKRTADGSEFESPKKKRKVGSNLRNYPGSFNIGLDL 27 Cas GPASAGWSVVDSEGKLFHFKKKPTWGSRLFDSAQTASEARMHRGQRR mammalian RYVRRRWRLNLLQALFTEEMEKVDPGFFMRLNHSRTVEGDPIFTKDFTK TABLE 1E ASDR Type II Cas Sequences Name Sequence SEQ ID NO expression KDYYKRFPTIYHLRAHLMETDDPADLRLVYLAIHNIVKHRGNFLRQEEKLT construct AKIANTSEALGKLSVSLKEWCDSRGYSVGKVDEEAIAAVLADKRISRSQK (with N- AKNVTLLISVDTGDSKANAKLAKALANAIVGLQAEFKDLFGDFECEATKLN terminal LSEEEKLEALQAACPDDSAELLEAVCGAYSAYVLQGLLSYAEGQTISHN methionine, MIKKYDCYAEDLECLKALVRDYLPKTKGDDGLKPFDRFFRGVTYHDLDK stop codon, SDPSRDYDASKAKGYTAYNLHKLSYDEFRKEVEKLLKGTGAQDDERYIT V5 tag, N- MMDAFEKQRFLRRLKTSDNGAIYYQLHLEELQAILENQGRFYPFLKQEAS terminal NLS KIESLVTFRIPYYVGPLTSKNAAKDAHNKNRFQWSERKPGMEDAIITPWN and C- WESVIDKNRSAENFIMRMTGDCTYLAGEKVLPRHSLLYEEFCVLNELNG terminal NLS) VRFTNDGDNWHRLDSEQRAGIIRDLFHKKRSVKYADIAAWLVRMGYLER (aa) TPQVRGGQGESGLESKMSSYLLFAKDVFGVDELDEALFPAIEQIILWNTL FEDRSIFEEKLRDEFGPNGNGMLNNEQIKKCKKRLSGWGRLSEKFLTGI RVVTQGGRQMSIMNVLREGSPNPDRRQGEALVMMEVLHDNTLGFQKKI DEVNRAYYAQNAKAMGVNELPGSPGIRRSINQAIRIVDEIVKIAGHAPDNI FIEVTRDEDEKKKGKRTVKRWDRIEAALNAFKAEGGDLAVMSDFKRLKN TEADLDERLTLYLLQNGKCLYSGRAIDLDALMTGSGEYEVDHIIPRAYIKD DSLDNKALVYRSENQRKTDQLLIDENVRQKMGETWKQLHDAKMISDKKY NNLMRLTVTEGAMKGFIARQLVETSQIVKMTKALLEARYGDAGTRIVPVK ASMSHNLREVAGLVKCREANDFHHAHDAYLACRMGLFIQKRYPGIYENP IGYAHAMKKYVLEQSELFKKTHRMPGSAGLIVNSFMTAGFDPETGEMFE DAWAGNDEIKHCKKKHYDDAWNAAAEVEGIRRALNYRQCYVSRMPYED SGKFWDANPISKRSNPKLALKNTLDPACFGGYNTKTAAYFFIYEVTDKKG KTGLRIAQMPIFMKAKCESDNAAFEKYCQDLESESRVSFTRIVIPRLLKW QLVEIDGNLLRINAEEEMKSASQLAFSQEELERILLVCTDGYLDPDVNRS VYMAIAKALDRAKCPMLTNDLDLISRFDAFCGLAVADQKDLLKKILLLCKG NKQSSPINLSSVGGKKKAGYLRPARSKVPDGMVLIDQSVTGMFERRTRV GVSRKRTADGSEFESPKKKRKV ASDR Type II ATGAACCTGAGAAATTACCCTGGGTCGTTCAATATTGGTCTTGACCTT 28 Cas coding GGTCCCGCGTCGGCTGGCTGGTCGGTAGTGGACAGTGAGGGAAAAC sequence (nt) TCTTTCATTTCAAGAAAAAACCAACATGGGGAAGTCGTTTGTTTGATA (not codon GTGCGCAGACTGCCTCGGAGGCGCGTATGCATCGCGGTCAGCGTCG optimized) ACGTTATGTGCGTCGCCGTTGGCGCCTCAACTTGTTGCAGGCACTGT TTACTGAGGAGATGGAAAAGGTTGACCCTGGGTTCTTTATGAGGCTC AACCATTCCCGAACCGTTGAGGGCGATCCTATCTTCACCAAGGACTT TACCAAGAAGGACTATTACAAGCGTTTTCCTACCATCTATCACTTGCG TGCGCACCTTATGGAAACCGACGATCCGGCCGATCTCCGTTTGGTTT ACTTGGCTATTCATAACATCGTTAAGCACCGTGGCAACTTCCTGCGTC AGGAGGAAAAGCTTACGGCAAAAATCGCAAACACCTCGGAAGCACTT GGGAAGCTTTCTGTCTCGCTAAAAGAATGGTGCGATTCGAGGGGATA TAGTGTCGGAAAGGTTGATGAGGAAGCAATCGCGGCAGTTTTAGCAG ACAAACGCATATCGCGTTCTCAAAAAGCAAAGAATGTGACATTGCTTA TCTCCGTTGATACAGGCGACAGTAAGGCAAACGCGAAGCTCGCTAAG GCGTTGGCGAATGCTATCGTGGGTCTCCAAGCTGAGTTCAAAGACCT ATTTGGCGATTTTGAGTGTGAAGCAACCAAGCTCAATCTTTCTGAAGA GGAGAAGCTCGAAGCGCTTCAGGCGGCGTGTCCCGATGATAGCGCG GAGCTGCTTGAAGCTGTATGTGGTGCGTATTCTGCGTATGTGCTCCA GGGTTTGCTTTCATATGCGGAGGGGCAGACCATTTCGCATAATATGA TAAAAAAGTACGATTGCTACGCGGAAGATCTGGAGTGCCTTAAAGCTT TGGTTCGCGACTATTTGCCGAAGACGAAGGGCGATGACGGACTCAA GCCATTTGACCGGTTTTTTCGTGGCGTCACTTATCATGATTTGGATAA GAGTGATCCCTCGCGCGACTACGATGCGTCAAAAGCCAAAGGCTATA CCGCCTATAACTTGCATAAACTTTCGTACGACGAATTCAGAAAAGAAG TTGAAAAGCTGCTAAAGGGCACAGGTGCTCAGGACGATGAGCGCTAT ATCACCATGATGGATGCGTTTGAGAAGCAGCGTTTCTTGCGTCGCTT GAAGACGAGTGATAACGGGGCAATCTACTATCAGCTGCACTTGGAAG AGTTGCAGGCGATTCTTGAAAATCAAGGTCGTTTCTATCCCTTCCTTA AACAAGAGGCAAGCAAGATAGAAAGCCTGGTTACCTTCCGGATCCCC TABLE 1E ASDR Type II Cas Sequences Name Sequence SEQ ID NO TACTACGTGGGTCCGCTTACGTCTAAAAACGCTGCAAAGGATGCTCA TAACAAGAACCGTTTTCAGTGGTCGGAGCGCAAGCCGGGTATGGAG GATGCGATTATCACGCCGTGGAACTGGGAAAGCGTTATCGATAAGAA TCGGAGCGCGGAGAACTTTATTATGCGCATGACGGGGGATTGCACAT ACTTGGCGGGCGAGAAAGTACTGCCCAGGCATTCCTTGCTCTACGAG GAGTTCTGCGTACTCAATGAGCTAAACGGCGTTCGTTTCACTAATGAC GGCGATAATTGGCACCGCCTGGATTCCGAGCAGCGTGCAGGCATTAT TCGCGATTTGTTCCATAAGAAGCGCAGTGTGAAGTATGCAGACATTG CCGCCTGGCTTGTGCGGATGGGCTATCTTGAGAGAACTCCTCAGGTG CGCGGCGGACAGGGTGAAAGCGGCTTGGAATCAAAGATGAGCTCGT ACCTCTTGTTTGCTAAGGATGTTTTCGGCGTTGACGAGCTTGATGAAG CGCTATTCCCTGCCATTGAGCAGATCATTTTGTGGAACACGCTGTTTG AGGATCGCTCGATTTTTGAAGAAAAGCTTCGCGATGAATTCGGGCCT AACGGCAACGGCATGCTCAATAACGAGCAGATAAAGAAATGCAAAAA GCGCCTTTCGGGGTGGGGTAGGCTTTCTGAAAAGTTCCTTACTGGAA TCAGGGTCGTCACGCAGGGTGGTCGCCAGATGAGCATCATGAATGT GCTTCGCGAGGGTAGTCCGAATCCCGATCGCCGCCAGGGTGAAGCT TTGGTGATGATGGAGGTTCTTCATGACAACACCCTTGGCTTCCAGAAA AAGATCGACGAGGTAAACCGTGCGTACTATGCGCAAAATGCCAAAGC GATGGGCGTAAACGAACTTCCCGGCTCGCCTGGTATTCGCCGTAGCA TTAACCAAGCTATTCGCATTGTTGACGAAATTGTAAAGATTGCTGGTC ATGCGCCCGACAACATCTTTATTGAGGTCACGCGTGACGAAGATGAA AAGAAGAAGGGGAAGCGTACCGTCAAGCGCTGGGACCGCATAGAAG CGGCGCTCAATGCATTCAAGGCTGAGGGCGGCGACCTTGCGGTTAT GAGCGATTTCAAGCGGCTTAAGAACACAGAAGCCGATTTGGACGAAC GACTCACTTTGTATTTGCTGCAAAATGGCAAATGCCTGTATTCAGGCC GCGCTATCGACCTCGATGCGCTTATGACGGGATCGGGCGAGTATGA AGTCGACCACATTATTCCGCGTGCATATATTAAGGACGATAGCCTCGA CAACAAGGCGTTGGTGTATCGTTCCGAAAATCAACGAAAAACCGATC AACTTTTGATTGACGAGAACGTTCGCCAAAAGATGGGCGAAACATGG AAGCAGCTTCATGACGCGAAAATGATTAGCGACAAGAAGTATAACAA CCTGATGCGACTCACCGTTACCGAAGGAGCAATGAAGGGCTTTATTG CTCGCCAGCTGGTTGAGACAAGTCAGATAGTGAAGATGACAAAGGCG TTGCTTGAAGCGCGTTATGGCGATGCGGGGACGAGGATCGTCCCTG TTAAGGCAAGCATGTCCCACAACCTTCGTGAAGTGGCGGGCTTGGTT AAATGCCGTGAAGCCAATGATTTCCATCATGCTCATGACGCGTATCTG GCGTGCCGCATGGGTTTGTTTATCCAGAAGCGTTACCCAGGGATTTA CGAGAACCCCATTGGTTATGCGCATGCCATGAAAAAGTATGTTCTTGA ACAGTCGGAGCTGTTCAAGAAGACTCACCGGATGCCTGGATCTGCTG GCTTGATTGTAAACAGCTTTATGACGGCGGGGTTTGATCCCGAAACT GGCGAGATGTTCGAAGATGCCTGGGCTGGCAACGATGAAATCAAGC ATTGTAAGAAAAAGCACTATGACGATGCATGGAATGCAGCTGCGGAG GTGGAGGGTATTCGTCGTGCACTCAACTACCGACAGTGTTATGTCTC GAGGATGCCTTACGAAGATTCGGGCAAGTTCTGGGACGCGAACCCG ATTTCAAAGAGAAGCAATCCCAAGCTTGCGCTTAAAAACACCCTCGAC CCTGCGTGCTTTGGCGGCTACAACACAAAGACAGCGGCGTATTTCTT TATATATGAGGTTACTGATAAAAAAGGTAAGACCGGGTTACGAATAGC TCAGATGCCCATTTTCATGAAAGCTAAATGCGAGAGCGATAATGCCG CTTTTGAGAAATATTGCCAAGATCTGGAAAGCGAAAGTAGGGTTTCCT TTACTCGAATCGTTATCCCAAGGCTGTTGAAATGGCAACTCGTCGAAA TCGATGGAAACCTTCTGCGCATTAATGCGGAAGAGGAGATGAAATCT GCGTCTCAGTTGGCATTTTCGCAAGAAGAGCTTGAAAGAATCCTACTT GTCTGTACAGACGGCTATCTTGACCCTGACGTTAATCGATCTGTCTAT ATGGCAATTGCGAAAGCGCTCGATAGGGCAAAATGCCCCATGTTGAC GAATGATCTGGATCTTATAAGCAGATTCGATGCATTTTGCGGTCTTGC CGTGGCTGATCAGAAGGATTTATTAAAAAAGATTCTTCTTCTCTGCAA GGGAAATAAACAGTCCTCTCCAATTAATCTTTCGTCTGTTGGTGGAAA TABLE 1E ASDR Type II Cas Sequences Name Sequence SEQ ID NO GAAGAAGGCAGGGTATCTCCGCCCAGCTCGTAGTAAAGTGCCCGAT GGGATGGTGCTCATCGACCAATCCGTAACGGGTATGTTCGAGAGGA GAACCCGTGTCGGGGTTTAG ASDR Type II AACCTGAGAAACTACCCCGGCTCCTTCAACATCGGACTGGACTTGGG 29 Cas coding CCCTGCCTCTGCCGGCTGGAGCGTGGTCGATAGCGAGGGAAAGCTG sequence (nt) TTTCATTTCAAAAAGAAGCCTACATGGGGCAGCAGACTCTTTGACAGC (human GCTCAGACCGCCTCCGAGGCCAGAATGCACAGAGGACAGCGGCGG codon- CGGTATGTGCGCAGGAGATGGCGGCTCAACCTGCTGCAGGCGCTGT optimized; TCACCGAAGAGATGGAAAAAGTGGACCCTGGTTTTTTTATGAGACTGA lacking N- ACCACAGCCGGACAGTAGAGGGCGATCCCATCTTCACCAAGGACTTC terminal ACCAAGAAGGATTACTACAAGCGGTTCCCCACCATCTACCACCTGAG methionine; GGCTCACCTGATGGAAACCGACGATCCCGCCGACCTGAGACTGGTC no stop TACCTGGCCATCCACAACATCGTGAAACACAGAGGCAACTTCCTGAG codon) GCAGGAGGAGAAGCTGACCGCTAAAATCGCTAACACCAGCGAGGCA CTGGGCAAGCTGTCTGTCTCCCTGAAAGAGTGGTGCGACTCCAGAG GATATTCTGTGGGCAAGGTGGATGAAGAAGCCATTGCTGCCGTGCTG GCTGACAAGCGAATAAGCAGATCTCAAAAGGCCAAGAACGTGACCCT GCTGATCAGCGTGGACACCGGAGATAGCAAGGCCAACGCCAAGCTG GCCAAGGCCCTGGCCAATGCCATCGTGGGACTGCAAGCTGAATTTAA GGATCTGTTCGGAGATTTCGAATGCGAGGCTACAAAGCTGAACCTGA GCGAGGAAGAGAAGCTGGAAGCCCTGCAAGCTGCCTGTCCTGATGA CAGCGCCGAGCTGCTGGAGGCCGTGTGTGGCGCCTATAGCGCCTAC GTGCTGCAGGGCCTGCTGAGCTACGCCGAGGGCCAGACAATCAGCC ACAATATGATCAAGAAGTACGACTGCTACGCCGAGGACCTGGAGTGT CTGAAGGCCCTGGTGAGAGACTACCTGCCAAAGACCAAGGGCGACG ACGGCCTCAAGCCTTTCGACAGATTCTTCCGTGGCGTGACATACCAC GACCTGGACAAGAGCGACCCTAGCAGAGATTACGATGCAAGCAAGG CAAAGGGCTACACCGCCTACAACCTGCACAAGCTGTCTTATGACGAG TTCAGAAAGGAAGTGGAAAAGCTGCTGAAGGGCACCGGCGCCCAGG ACGACGAAAGATACATCACAATGATGGACGCTTTCGAAAAGCAGAGA TTCCTACGGCGGCTTAAGACATCTGACAACGGCGCCATCTACTACCA GCTGCACCTGGAGGAGCTGCAGGCCATCCTGGAAAACCAGGGCAGA TTTTACCCCTTCCTGAAACAGGAGGCCAGTAAAATCGAGTCTCTGGT GACCTTCAGAATCCCTTACTACGTGGGCCCACTGACGTCCAAGAACG CCGCCAAGGACGCCCACAACAAGAATAGATTTCAATGGTCCGAGAGA AAGCCTGGCATGGAAGATGCCATTATCACCCCTTGGAACTGGGAGTC TGTGATCGATAAGAATAGAAGCGCCGAGAACTTCATCATGCGGATGA CCGGCGATTGCACCTACCTGGCCGGCGAGAAGGTGCTGCCTAGACA CAGTCTGCTGTACGAGGAGTTCTGCGTGCTGAACGAGCTGAACGGC GTGCGGTTCACTAACGACGGCGACAACTGGCACCGGCTGGACTCAG AGCAGCGGGCTGGAATTATCAGAGATCTGTTTCACAAAAAGAGATCT GTGAAGTACGCCGACATCGCCGCCTGGCTGGTACGGATGGGCTATC TGGAGCGAACACCTCAGGTGCGCGGTGGACAGGGCGAAAGCGGCC TGGAATCTAAGATGAGCAGCTACCTGCTGTTCGCCAAAGACGTGTTC GGCGTGGACGAGCTGGACGAGGCCTTATTTCCTGCCATCGAACAGAT CATTCTGTGGAACACCCTGTTTGAGGATAGATCCATCTTCGAAGAGAA GCTGCGGGACGAGTTCGGCCCTAATGGCAATGGCATGCTGAACAAT GAACAGATCAAGAAATGCAAAAAAAGACTCAGCGGATGGGGCAGACT GAGCGAAAAGTTCCTGACAGGCATAAGGGTTGTGACACAGGGCGGC AGACAGATGAGCATCATGAACGTGCTGCGGGAAGGCAGCCCTAACC CTGACAGACGGCAGGGCGAGGCCCTCGTAATGATGGAAGTGCTGCA TGACAACACCCTGGGCTTCCAGAAAAAGATCGACGAAGTGAACAGAG CCTACTACGCCCAGAACGCTAAGGCAATGGGCGTGAATGAGCTGCC CGGGAGCCCTGGCATCCGGCGAAGCATCAATCAAGCCATTAGAATC GTTGATGAGATCGTGAAGATCGCTGGCCACGCTCCTGACAACATCTT TATCGAGGTTACCCGGGATGAGGACGAAAAGAAGAAGGGCAAGCGG ACCGTGAAAAGATGGGACCGGATCGAAGCCGCTCTCAACGCCTTTAA TABLE 1E ASDR Type II Cas Sequences Name Sequence SEQ ID NO AGCCGAGGGTGGAGATCTGGCAGTGATGAGCGATTTTAAGCGCCTG AAGAATACAGAAGCTGATCTGGATGAGCGCCTCACACTGTACTTACT GCAGAACGGCAAGTGCCTGTACAGCGGCAGAGCGATCGACTTAGAT GCCCTGATGACCGGCAGCGGCGAGTACGAAGTGGATCACATCATCC CTAGAGCCTACATCAAGGACGATTCCCTGGACAACAAGGCCCTTGTC TACCGGAGCGAGAACCAGAGAAAGACCGACCAACTGCTGATCGACG AGAACGTGCGGCAGAAAATGGGCGAAACCTGGAAGCAGCTTCATGAT GCCAAGATGATCAGCGACAAGAAGTACAACAACCTGATGAGACTGAC CGTGACAGAGGGAGCCATGAAAGGCTTCATCGCCAGACAGTTGGTC GAGACATCTCAGATCGTGAAGATGACCAAGGCCTTGCTGGAGGCCC GGTACGGCGATGCTGGCACAAGAATCGTGCCTGTGAAGGCCAGCAT GAGCCACAACCTGAGAGAGGTTGCGGGCTTGGTCAAGTGTAGAGAG GCCAACGACTTCCACCACGCCCATGACGCTTACCTGGCCTGCAGAAT GGGCCTGTTTATCCAGAAGAGATACCCAGGAATCTACGAAAACCCCA TCGGCTACGCTCACGCCATGAAGAAATACGTGCTGGAACAGAGCGAA CTATTCAAGAAAACCCACCGGATGCCTGGCTCCGCTGGCCTGATCGT GAACAGCTTCATGACAGCTGGATTCGACCCCGAGACAGGCGAGATGT TCGAGGACGCCTGGGCCGGAAACGATGAGATCAAGCACTGCAAGAA GAAGCACTACGACGACGCCTGGAACGCCGCCGCCGAGGTGGAAGG AATCAGACGGGCCCTGAATTACAGACAATGTTACGTGAGCCGGATGC CATACGAGGACTCCGGCAAGTTCTGGGATGCTAACCCTATCAGCAAG AGAAGCAACCCCAAGTTGGCTCTAAAGAACACCCTGGACCCTGCTTG CTTCGGAGGCTACAATACCAAGACCGCCGCCTACTTCTTCATCTACG AGGTGACCGACAAAAAAGGCAAGACCGGCCTGCGAATCGCCCAGAT GCCAATCTTCATGAAGGCTAAGTGCGAGAGCGATAACGCCGCCTTCG AAAAGTACTGCCAGGACCTGGAAAGCGAATCCAGAGTGAGCTTCACC CGGATCGTGATCCCCAGACTCCTCAAGTGGCAGCTCGTGGAAATCGA CGGCAACCTGCTGCGGATCAACGCCGAAGAGGAGATGAAATCCGCC TCTCAGCTGGCCTTCAGCCAGGAGGAGCTGGAGAGAATCCTGTTGGT CTGTACAGATGGCTATCTGGACCCCGACGTGAACAGAAGCGTGTACA TGGCCATCGCCAAAGCCCTCGACAGAGCGAAGTGCCCTATGCTGAC CAACGACCTCGATCTGATCAGCAGATTCGATGCCTTCTGCGGCCTCG CCGTGGCCGATCAGAAGGACCTTCTCAAGAAGATCCTGCTGCTGTGC AAGGGCAACAAACAGAGCAGTCCCATCAACCTGAGCAGCGTGGGCG GCAAGAAGAAGGCAGGCTACCTGCGGCCTGCCCGGAGCAAAGTTCC TGATGGAATGGTGCTGATCGACCAGTCCGTGACAGGGATGTTCGAGC GGAGGACAAGAGTGGGCGTG ASDR Type II ATGGGCAAACCTATCCCCAATCCCCTGCTGGGCCTGGACAGCACCAA 30 Cas GCGGACTGCCGACGGCAGTGAGTTCGAGTCTCCCAAGAAAAAGAGG mammalian AAAGTGGGATCCAACCTGAGAAACTACCCCGGCTCCTTCAACATCGG expression ACTGGACTTGGGCCCTGCCTCTGCCGGCTGGAGCGTGGTCGATAGC construct GAGGGAAAGCTGTTTCATTTCAAAAAGAAGCCTACATGGGGCAGCAG (with N- ACTCTTTGACAGCGCTCAGACCGCCTCCGAGGCCAGAATGCACAGA terminal GGACAGCGGCGGCGGTATGTGCGCAGGAGATGGCGGCTCAACCTG methionine, CTGCAGGCGCTGTTCACCGAAGAGATGGAAAAAGTGGACCCTGGTTT stop codon, TTTTATGAGACTGAACCACAGCCGGACAGTAGAGGGCGATCCCATCT V5 tag, N- TCACCAAGGACTTCACCAAGAAGGATTACTACAAGCGGTTCCCCACC terminal NLS ATCTACCACCTGAGGGCTCACCTGATGGAAACCGACGATCCCGCCGA and C- CCTGAGACTGGTCTACCTGGCCATCCACAACATCGTGAAACACAGAG terminal NLS) GCAACTTCCTGAGGCAGGAGGAGAAGCTGACCGCTAAAATCGCTAAC (nt) ACCAGCGAGGCACTGGGCAAGCTGTCTGTCTCCCTGAAAGAGTGGT GCGACTCCAGAGGATATTCTGTGGGCAAGGTGGATGAAGAAGCCATT GCTGCCGTGCTGGCTGACAAGCGAATAAGCAGATCTCAAAAGGCCAA GAACGTGACCCTGCTGATCAGCGTGGACACCGGAGATAGCAAGGCC AACGCCAAGCTGGCCAAGGCCCTGGCCAATGCCATCGTGGGACTGC AAGCTGAATTTAAGGATCTGTTCGGAGATTTCGAATGCGAGGCTACAA AGCTGAACCTGAGCGAGGAAGAGAAGCTGGAAGCCCTGCAAGCTGC TABLE 1E ASDR Type II Cas Sequences Name Sequence SEQ ID NO CTGTCCTGATGACAGCGCCGAGCTGCTGGAGGCCGTGTGTGGCGCC TATAGCGCCTACGTGCTGCAGGGCCTGCTGAGCTACGCCGAGGGCC AGACAATCAGCCACAATATGATCAAGAAGTACGACTGCTACGCCGAG GACCTGGAGTGTCTGAAGGCCCTGGTGAGAGACTACCTGCCAAAGA CCAAGGGCGACGACGGCCTCAAGCCTTTCGACAGATTCTTCCGTGG CGTGACATACCACGACCTGGACAAGAGCGACCCTAGCAGAGATTACG ATGCAAGCAAGGCAAAGGGCTACACCGCCTACAACCTGCACAAGCTG TCTTATGACGAGTTCAGAAAGGAAGTGGAAAAGCTGCTGAAGGGCAC CGGCGCCCAGGACGACGAAAGATACATCACAATGATGGACGCTTTCG AAAAGCAGAGATTCCTACGGCGGCTTAAGACATCTGACAACGGCGCC ATCTACTACCAGCTGCACCTGGAGGAGCTGCAGGCCATCCTGGAAAA CCAGGGCAGATTTTACCCCTTCCTGAAACAGGAGGCCAGTAAAATCG AGTCTCTGGTGACCTTCAGAATCCCTTACTACGTGGGCCCACTGACG TCCAAGAACGCCGCCAAGGACGCCCACAACAAGAATAGATTTCAATG GTCCGAGAGAAAGCCTGGCATGGAAGATGCCATTATCACCCCTTGGA ACTGGGAGTCTGTGATCGATAAGAATAGAAGCGCCGAGAACTTCATC ATGCGGATGACCGGCGATTGCACCTACCTGGCCGGCGAGAAGGTGC TGCCTAGACACAGTCTGCTGTACGAGGAGTTCTGCGTGCTGAACGAG CTGAACGGCGTGCGGTTCACTAACGACGGCGACAACTGGCACCGGC TGGACTCAGAGCAGCGGGCTGGAATTATCAGAGATCTGTTTCACAAA AAGAGATCTGTGAAGTACGCCGACATCGCCGCCTGGCTGGTACGGA TGGGCTATCTGGAGCGAACACCTCAGGTGCGCGGTGGACAGGGCGA AAGCGGCCTGGAATCTAAGATGAGCAGCTACCTGCTGTTCGCCAAAG ACGTGTTCGGCGTGGACGAGCTGGACGAGGCCTTATTTCCTGCCATC GAACAGATCATTCTGTGGAACACCCTGTTTGAGGATAGATCCATCTTC GAAGAGAAGCTGCGGGACGAGTTCGGCCCTAATGGCAATGGCATGC TGAACAATGAACAGATCAAGAAATGCAAAAAAAGACTCAGCGGATGG GGCAGACTGAGCGAAAAGTTCCTGACAGGCATAAGGGTTGTGACACA GGGCGGCAGACAGATGAGCATCATGAACGTGCTGCGGGAAGGCAGC CCTAACCCTGACAGACGGCAGGGCGAGGCCCTCGTAATGATGGAAG TGCTGCATGACAACACCCTGGGCTTCCAGAAAAAGATCGACGAAGTG AACAGAGCCTACTACGCCCAGAACGCTAAGGCAATGGGCGTGAATGA GCTGCCCGGGAGCCCTGGCATCCGGCGAAGCATCAATCAAGCCATT AGAATCGTTGATGAGATCGTGAAGATCGCTGGCCACGCTCCTGACAA CATCTTTATCGAGGTTACCCGGGATGAGGACGAAAAGAAGAAGGGCA AGCGGACCGTGAAAAGATGGGACCGGATCGAAGCCGCTCTCAACGC CTTTAAAGCCGAGGGTGGAGATCTGGCAGTGATGAGCGATTTTAAGC GCCTGAAGAATACAGAAGCTGATCTGGATGAGCGCCTCACACTGTAC TTACTGCAGAACGGCAAGTGCCTGTACAGCGGCAGAGCGATCGACTT AGATGCCCTGATGACCGGCAGCGGCGAGTACGAAGTGGATCACATC ATCCCTAGAGCCTACATCAAGGACGATTCCCTGGACAACAAGGCCCT TGTCTACCGGAGCGAGAACCAGAGAAAGACCGACCAACTGCTGATC GACGAGAACGTGCGGCAGAAAATGGGCGAAACCTGGAAGCAGCTTC ATGATGCCAAGATGATCAGCGACAAGAAGTACAACAACCTGATGAGA CTGACCGTGACAGAGGGAGCCATGAAAGGCTTCATCGCCAGACAGTT GGTCGAGACATCTCAGATCGTGAAGATGACCAAGGCCTTGCTGGAG GCCCGGTACGGCGATGCTGGCACAAGAATCGTGCCTGTGAAGGCCA GCATGAGCCACAACCTGAGAGAGGTTGCGGGCTTGGTCAAGTGTAG AGAGGCCAACGACTTCCACCACGCCCATGACGCTTACCTGGCCTGCA GAATGGGCCTGTTTATCCAGAAGAGATACCCAGGAATCTACGAAAAC CCCATCGGCTACGCTCACGCCATGAAGAAATACGTGCTGGAACAGAG CGAACTATTCAAGAAAACCCACCGGATGCCTGGCTCCGCTGGCCTGA TCGTGAACAGCTTCATGACAGCTGGATTCGACCCCGAGACAGGCGA GATGTTCGAGGACGCCTGGGCCGGAAACGATGAGATCAAGCACTGC AAGAAGAAGCACTACGACGACGCCTGGAACGCCGCCGCCGAGGTGG AAGGAATCAGACGGGCCCTGAATTACAGACAATGTTACGTGAGCCGG ATGCCATACGAGGACTCCGGCAAGTTCTGGGATGCTAACCCTATCAG TABLE 1E ASDR Type II Cas Sequences Name Sequence SEQ ID NO CAAGAGAAGCAACCCCAAGTTGGCTCTAAAGAACACCCTGGACCCTG CTTGCTTCGGAGGCTACAATACCAAGACCGCCGCCTACTTCTTCATCT ACGAGGTGACCGACAAAAAAGGCAAGACCGGCCTGCGAATCGCCCA GATGCCAATCTTCATGAAGGCTAAGTGCGAGAGCGATAACGCCGCCT TCGAAAAGTACTGCCAGGACCTGGAAAGCGAATCCAGAGTGAGCTTC ACCCGGATCGTGATCCCCAGACTCCTCAAGTGGCAGCTCGTGGAAAT CGACGGCAACCTGCTGCGGATCAACGCCGAAGAGGAGATGAAATCC GCCTCTCAGCTGGCCTTCAGCCAGGAGGAGCTGGAGAGAATCCTGT TGGTCTGTACAGATGGCTATCTGGACCCCGACGTGAACAGAAGCGTG TACATGGCCATCGCCAAAGCCCTCGACAGAGCGAAGTGCCCTATGCT GACCAACGACCTCGATCTGATCAGCAGATTCGATGCCTTCTGCGGCC TCGCCGTGGCCGATCAGAAGGACCTTCTCAAGAAGATCCTGCTGCTG TGCAAGGGCAACAAACAGAGCAGTCCCATCAACCTGAGCAGCGTGG GCGGCAAGAAGAAGGCAGGCTACCTGCGGCCTGCCCGGAGCAAAGT TCCTGATGGAATGGTGCTGATCGACCAGTCCGTGACAGGGATGTTCG AGCGGAGGACAAGAGTGGGCGTGTCTAGAAAGCGGACAGCAGACGG CTCCGAATTTGAAAGCCCTAAGAAAAAGAGAAAGGTGTGA [0088] In some embodiments an ASDR Type II Cas protein comprises an amino acid sequence of SEQ ID NO:25, SEQ ID NO:26, or SEQ ID NO:27. In some embodiments, an ASDR Type II Cas protein has nickase activity, for example resulting from one or more amino acid substitutions relative to the sequence of SEQ ID NO:25, SEQ ID NO:26, or SEQ ID NO:27. In some embodiments, the one or more amino acid substitutions providing nickase activity comrpise a D15A substitution, wherein the position of the D15A substitution is defined with respect to the amino acid numbering of SEQ ID NO:26. In some embodiments, the one or more amino acid substitutions providing nickase activity comprise an H898A substitution, wherein the position of the H890A substitution is defined with respect to the amino acid numbering of SEQ ID NO:26. In some embodiments, an ASDR Type II Cas protein is catalytically inactive, for example due to a D15A substitution in combination with an H898A substitution. 6.2.1.6. BCZZ Type II Cas Proteins [0089] In one aspect, the disclosure provides BCZZ Type II Cas proteins. BCZZ Type II Cas proteins can be further classified as Type IIA Cas proteins. The BCZZ Type II Cas proteins typically comprise an amino acid sequence that is at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 85%, at least 90%, or at least 95% identical to SEQ ID NO:31. In some embodiments, the BCZZ Type II Cas proteins comprise an amino acid sequence that is at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical to SEQ ID NO:31. In some embodiments, a BCZZ Type II Cas protein comprises an amino acid sequence that is identical to SEQ ID NO:31. [0090] Exemplary BCZZ Type II Cas protein sequences and nucleotide sequences encoding exemplary BCZZ Type II Cas proteins are set forth in Table 1F. TABLE 1F BCZZ Type II Cas Sequences Name Sequence SEQ ID NO BCZZ Type II EQKNCKKVDAYNIGLDIGTNSVGWAVTDLDNNILKRGNKNMWGARIFDE 31 Cas amino GSTAAQTRMFRGTRRRIERRKERINILQSLLLDDMEKEYPNFFPMLRETS acid KTKEDKNSDSINGKKYNLFSELELSDPIYYQKYPTIYHLRRELMESKEKH sequence DIRLVYLAIHHIIKYRGNFLYEGDLKNNNNEIIESINELLDFIQNTVEISFISEA TABLE 1F BCZZ Type II Cas Sequences Name Sequence SEQ ID NO (without N- EKIQEILKDKTKSKGEKKDLIIGLFDYDKDEKSVITGIVNAILGYKFDVNKIF terminal DIAVENSNISFSNEIANEEEIKENLQDQAYVYDVLLKIYNWFILQDILQGNN methionine) SISEAFINKYDKYKKDLSNIKSLYRNYLKDEYNSMFRREKDDSYSCFEKHI SACPIENLYKRIKKDLADVPDCKEKEEILKDIEDNAFLVKINTTANSAIPYQL HYQELEKILENQAQYYQTIQENKQHILELMKFRIPYYVGPLAQEGRSRFA WIVRNSNEKILPWTFDDVVNVDETAEKFIRRMTNKCTYLINEDVIPKQSIL YSEFCVLNELANIRVNNHKLTPKTKQLIIDKLFKVKKTIKQRDLKQVLTDFQ VFPEVTSITGLAEVDKFMSNMSSYIDIENILGCINDENIEMVENIIEWITIFED KKILKRKIKNTYNLEDKVVSQLIKKNYSGWSRLSKELLVGLKAADDGKSIM EKLRTTKDNFMQIINNDMYGFNKQIESRMKLQEKQITYKDVDEIPTSPAN KRAIWQSLKIVEEIKKVMKKDPKNIYIEFAREEQEKNKRKDNRAKALLKIY EKYEEEIKMLKDYNPNVYKELKKKQNEKDFNERLYLYFIQNGRCMYSSR PLNIDTLYLYEVDHILPQSYVKDDSLSNKALVYKEENQRKSGSLLLDEKIIN RQETWWKQLRKNGLIDEKKYYNLTRRKMFETDKEKVKFVSRQLIETRQS TKYITNLLVNQYSNTDVFAIRSNLTHNFRKFFEVYKNRNVNDYHHAQDAY IISVIGNVIDQKLQYKDEYKYTEYVKKYIKKSEEEANKVWIIMGMVSNNINK EKVKKALEYKDCFITHKLEEQTGAFYNQTLYSPKDKNIEIPLKEGLDVKKY GGYSGEIKAYFTIYSYIAQKGKMQIEMIGIPVKTKYDIENGKTTLIKYIQSKN SEASKIEIIRPKILKYQEYIDENNEPMMLLSDSEIRTNRQLIINNEMSKLIYIM NQEKIADDEKEKVQEKMEEIYNYLLGKLKNEYKTFNSIYQKLSNEETKEIF QKLEYKDKVSAINGIIDLMHRGQGNLTKLKQGDRAGRKSGKSFKTEALN KMIFIDKSVTGMYERRYKVNGMENCSSK BCZZ Type II MEQKNCKKVDAYNIGLDIGTNSVGWAVTDLDNNILKRGNKNMWGARIFD 32 Cas amino EGSTAAQTRMFRGTRRRIERRKERINILQSLLLDDMEKEYPNFFPMLRET acid SKTKEDKNSDSINGKKYNLFSELELSDPIYYQKYPTIYHLRRELMESKEKH sequence DIRLVYLAIHHIIKYRGNFLYEGDLKNNNNEIIESINELLDFIQNTVEISFISEA EKIQEILKDKTKSKGEKKDLIIGLFDYDKDEKSVITGIVNAILGYKFDVNKIF DIAVENSNISFSNEIANEEEIKENLQDQAYVYDVLLKIYNWFILQDILQGNN SISEAFINKYDKYKKDLSNIKSLYRNYLKDEYNSMFRREKDDSYSCFEKHI SACPIENLYKRIKKDLADVPDCKEKEEILKDIEDNAFLVKINTTANSAIPYQL HYQELEKILENQAQYYQTIQENKQHILELMKFRIPYYVGPLAQEGRSRFA WIVRNSNEKILPWTFDDVVNVDETAEKFIRRMTNKCTYLINEDVIPKQSIL YSEFCVLNELANIRVNNHKLTPKTKQLIIDKLFKVKKTIKQRDLKQVLTDFQ VFPEVTSITGLAEVDKFMSNMSSYIDIENILGCINDENIEMVENIIEWITIFED KKILKRKIKNTYNLEDKVVSQLIKKNYSGWSRLSKELLVGLKAADDGKSIM EKLRTTKDNFMQIINNDMYGFNKQIESRMKLQEKQITYKDVDEIPTSPAN KRAIWQSLKIVEEIKKVMKKDPKNIYIEFAREEQEKNKRKDNRAKALLKIY EKYEEEIKMLKDYNPNVYKELKKKQNEKDFNERLYLYFIQNGRCMYSSR PLNIDTLYLYEVDHILPQSYVKDDSLSNKALVYKEENQRKSGSLLLDEKIIN RQETWWKQLRKNGLIDEKKYYNLTRRKMFETDKEKVKFVSRQLIETRQS TKYITNLLVNQYSNTDVFAIRSNLTHNFRKFFEVYKNRNVNDYHHAQDAY IISVIGNVIDQKLQYKDEYKYTEYVKKYIKKSEEEANKVWIIMGMVSNNINK EKVKKALEYKDCFITHKLEEQTGAFYNQTLYSPKDKNIEIPLKEGLDVKKY GGYSGEIKAYFTIYSYIAQKGKMQIEMIGIPVKTKYDIENGKTTLIKYIQSKN SEASKIEIIRPKILKYQEYIDENNEPMMLLSDSEIRTNRQLIINNEMSKLIYIM NQEKIADDEKEKVQEKMEEIYNYLLGKLKNEYKTFNSIYQKLSNEETKEIF QKLEYKDKVSAINGIIDLMHRGQGNLTKLKQGDRAGRKSGKSFKTEALN KMIFIDKSVTGMYERRYKVNGMENCSSK BCZZ Type II MGKPIPNPLLGLDSTKRTADGSEFESPKKKRKVGSEQKNCKKVDAYNIG 33 Cas LDIGTNSVGWAVTDLDNNILKRGNKNMWGARIFDEGSTAAQTRMFRGT mammalian RRRIERRKERINILQSLLLDDMEKEYPNFFPMLRETSKTKEDKNSDSINGK expression KYNLFSELELSDPIYYQKYPTIYHLRRELMESKEKHDIRLVYLAIHHIIKYR construct GNFLYEGDLKNNNNEIIESINELLDFIQNTVEISFISEAEKIQEILKDKTKSK (with N- GEKKDLIIGLFDYDKDEKSVITGIVNAILGYKFDVNKIFDIAVENSNISFSNEI terminal ANEEEIKENLQDQAYVYDVLLKIYNWFILQDILQGNNSISEAFINKYDKYKK methionine, DLSNIKSLYRNYLKDEYNSMFRREKDDSYSCFEKHISACPIENLYKRIKKD stop codon, LADVPDCKEKEEILKDIEDNAFLVKINTTANSAIPYQLHYQELEKILENQAQ TABLE 1F BCZZ Type II Cas Sequences Name Sequence SEQ ID NO V5 tag, N- YYQTIQENKQHILELMKFRIPYYVGPLAQEGRSRFAWIVRNSNEKILPWT terminal NLS FDDVVNVDETAEKFIRRMTNKCTYLINEDVIPKQSILYSEFCVLNELANIRV and C- NNHKLTPKTKQLIIDKLFKVKKTIKQRDLKQVLTDFQVFPEVTSITGLAEVD terminal NLS) KFMSNMSSYIDIENILGCINDENIEMVENIIEWITIFEDKKILKRKIKNTYNLE (aa) DKVVSQLIKKNYSGWSRLSKELLVGLKAADDGKSIMEKLRTTKDNFMQII NNDMYGFNKQIESRMKLQEKQITYKDVDEIPTSPANKRAIWQSLKIVEEIK KVMKKDPKNIYIEFAREEQEKNKRKDNRAKALLKIYEKYEEEIKMLKDYNP NVYKELKKKQNEKDFNERLYLYFIQNGRCMYSSRPLNIDTLYLYEVDHIL PQSYVKDDSLSNKALVYKEENQRKSGSLLLDEKIINRQETWWKQLRKNG LIDEKKYYNLTRRKMFETDKEKVKFVSRQLIETRQSTKYITNLLVNQYSNT DVFAIRSNLTHNFRKFFEVYKNRNVNDYHHAQDAYIISVIGNVIDQKLQYK DEYKYTEYVKKYIKKSEEEANKVWIIMGMVSNNINKEKVKKALEYKDCFIT HKLEEQTGAFYNQTLYSPKDKNIEIPLKEGLDVKKYGGYSGEIKAYFTIYS YIAQKGKMQIEMIGIPVKTKYDIENGKTTLIKYIQSKNSEASKIEIIRPKILKY QEYIDENNEPMMLLSDSEIRTNRQLIINNEMSKLIYIMNQEKIADDEKEKV QEKMEEIYNYLLGKLKNEYKTFNSIYQKLSNEETKEIFQKLEYKDKVSAIN GIIDLMHRGQGNLTKLKQGDRAGRKSGKSFKTEALNKMIFIDKSVTGMYE RRYKVNGMENCSSKSRKRTADGSEFESPKKKRKV BCZZ Type II ATGGAACAAAAAAATTGCAAAAAAGTTGACGCTTATAATATAGGATTG 34 Cas coding GACATTGGAACAAATTCAGTTGGATGGGCTGTAACAGATTTGGATAAT sequence (nt) AATATTTTAAAAAGAGGCAACAAGAACATGTGGGGTGCACGCATATTT (not codon GATGAAGGAAGTACTGCAGCACAAACAAGAATGTTTAGAGGAACAAG optimized) AAGAAGGATTGAAAGAAGAAAAGAGAGAATAAATATACTTCAAAGTTT ACTATTAGATGACATGGAAAAGGAATATCCAAATTTCTTTCCAATGTTA AGGGAAACATCTAAGACTAAAGAAGATAAAAATAGTGACTCAATTAAT GGAAAAAAATATAATTTATTTTCAGAATTGGAATTATCAGATCCAATAT ATTATCAAAAATATCCAACAATATATCATTTAAGACGAGAGCTAATGGA AAGTAAAGAAAAGCATGACATTAGGCTTGTTTATCTTGCAATTCATCAT ATAATAAAATATCGTGGAAACTTCCTATATGAAGGAGACTTAAAAAATA ATAACAATGAAATAATTGAATCTATAAATGAGCTATTAGATTTTATACA AAATACAGTGGAAATAAGCTTTATTTCAGAAGCGGAAAAAATTCAAGA AATATTAAAAGATAAAACGAAATCAAAGGGAGAAAAAAAGGATTTAAT AATTGGACTTTTTGATTATGATAAGGATGAGAAGTCTGTAATTACTGG AATTGTTAATGCAATTTTGGGGTATAAATTTGACGTTAATAAGATATTT GATATTGCAGTAGAAAATTCAAATATTTCTTTTAGTAATGAAATAGCAA ATGAAGAAGAAATAAAAGAAAACTTGCAAGACCAAGCTTATGTGTATG ATGTATTACTAAAAATATATAATTGGTTTATATTACAAGATATTCTACAA GGTAATAATAGTATTTCAGAAGCTTTCATAAATAAGTATGACAAGTATA AAAAGGATTTAAGCAATATAAAAAGCTTATATAGAAATTATTTAAAAGA TGAATATAATAGTATGTTCAGAAGAGAAAAAGATGATAGCTATTCATGT TTCGAAAAGCATATATCAGCGTGTCCAATAGAAAACTTGTATAAAAGA ATAAAAAAAGATTTAGCAGATGTTCCAGATTGTAAGGAAAAAGAAGAA ATATTAAAAGATATAGAAGATAACGCTTTTTTAGTAAAAATAAATACAA CAGCAAATTCGGCAATTCCATATCAATTACATTATCAAGAATTGGAAAA AATACTTGAAAATCAAGCACAATATTATCAAACAATACAAGAAAATAAA CAACATATATTAGAACTAATGAAATTCAGAATTCCATATTATGTTGGAC CTTTGGCACAAGAGGGAAGAAGTAGATTTGCTTGGATTGTTAGAAATT CAAACGAAAAAATATTGCCATGGACATTTGATGACGTGGTCAATGTAG ATGAAACGGCAGAAAAGTTTATAAGAAGGATGACCAATAAATGTACAT ACCTAATAAATGAAGATGTAATTCCAAAACAATCAATTTTATACTCTGA GTTTTGTGTATTAAATGAACTTGCAAATATAAGAGTAAATAATCATAAA CTAACTCCAAAGACTAAACAACTTATAATAGACAAGTTATTTAAAGTCA AAAAAACAATTAAGCAAAGAGATTTAAAACAAGTATTAACAGATTTTCA AGTTTTTCCTGAAGTCACATCAATTACTGGATTAGCAGAAGTAGATAA ATTTATGTCAAACATGTCATCATATATTGATATTGAAAATATACTTGGC TGTATAAATGATGAAAACATTGAAATGGTTGAAAACATAATTGAATGGA TTACAATATTTGAAGATAAAAAAATATTAAAAAGAAAAATAAAAAATACA TABLE 1F BCZZ Type II Cas Sequences Name Sequence SEQ ID NO TACAATTTAGAAGATAAAGTAGTAAGTCAACTGATAAAGAAAAATTATA GTGGATGGTCGAGATTGTCAAAAGAATTACTTGTTGGATTAAAGGCTG CTGATGATGGAAAAAGCATAATGGAAAAGCTAAGGACAACAAAAGAC AATTTTATGCAAATTATAAATAATGATATGTATGGTTTTAATAAACAAAT TGAAAGTAGAATGAAACTACAAGAAAAACAAATAACATATAAAGATGT AGATGAAATTCCAACATCTCCAGCTAATAAAAGGGCCATATGGCAAAG CCTAAAAATAGTTGAAGAAATTAAAAAAGTAATGAAAAAAGATCCTAAA AACATTTATATTGAATTTGCAAGAGAAGAGCAGGAAAAAAATAAGAGA AAAGATAATAGAGCAAAAGCACTTCTTAAAATATACGAAAAATATGAA GAAGAAATCAAAATGCTAAAAGACTATAATCCAAATGTATATAAAGAG CTAAAGAAGAAACAAAATGAAAAAGATTTTAATGAAAGATTGTACTTAT ACTTTATTCAAAATGGAAGATGTATGTATAGTTCAAGACCATTAAATAT TGATACGCTTTACTTATATGAGGTGGATCATATATTGCCTCAAAGTTAT GTTAAAGATGATAGTTTATCAAACAAAGCGTTGGTATATAAAGAAGAA AATCAAAGAAAAAGTGGAAGTTTATTGTTAGATGAAAAAATAATAAATA GACAAGAAACATGGTGGAAGCAATTACGAAAAAATGGTTTGATTGATG AGAAAAAATATTATAACTTGACAAGAAGAAAAATGTTTGAAACTGATAA AGAAAAAGTGAAATTTGTGTCTAGACAATTGATAGAAACAAGGCAGAG CACAAAATATATAACAAATTTATTAGTTAATCAATATTCAAACACAGAT GTATTTGCAATAAGGTCAAATCTTACTCATAATTTTAGAAAATTCTTTG AAGTATACAAAAACAGAAATGTTAATGATTATCATCATGCACAGGATG CATATATTATAAGTGTAATAGGCAATGTAATAGATCAGAAATTACAATA CAAAGATGAATACAAATATACAGAGTATGTAAAAAAATATATAAAAAAG AGTGAAGAAGAAGCAAATAAAGTTTGGATAATAATGGGAATGGTTTCT AACAATATAAACAAAGAAAAAGTTAAAAAGGCTCTTGAATATAAAGATT GTTTTATAACACATAAATTAGAAGAACAAACAGGTGCATTTTATAACCA AACACTATATAGTCCAAAAGATAAAAACATAGAAATACCTTTAAAAGAG GGATTAGATGTTAAAAAATATGGAGGCTATTCTGGAGAAATTAAGGCA TATTTTACAATTTACAGTTATATTGCTCAAAAGGGAAAAATGCAAATAG AAATGATAGGAATTCCTGTAAAAACTAAATATGATATAGAAAATGGAAA AACTACACTAATTAAATATATACAAAGCAAAAATAGTGAAGCTTCTAAA ATAGAGATTATTAGACCTAAAATTTTAAAATATCAAGAATATATTGATG AAAATAACGAACCAATGATGCTATTAAGTGATAGTGAGATTAGAACTA ATAGGCAACTAATTATAAATAACGAAATGTCTAAACTAATATATATAAT GAATCAAGAAAAAATAGCAGATGATGAAAAAGAAAAAGTGCAAGAAAA AATGGAAGAAATTTATAATTATTTATTAGGTAAATTAAAAAATGAATATA AAACATTTAACTCTATATATCAAAAATTAAGTAATGAAGAGACAAAAGA AATTTTTCAAAAATTAGAATATAAAGACAAGGTGTCTGCTATAAATGGA ATTATTGATCTTATGCACAGAGGACAAGGCAATTTAACAAAATTAAAAC AAGGTGATAGAGCTGGAAGAAAAAGTGGAAAAAGTTTTAAAACAGAG GCATTAAATAAAATGATATTTATAGATAAATCTGTAACAGGAATGTACG AAAGAAGGTATAAAGTAAATGGGATGGAGAACTGTAGTAGTAAATAA BCZZ Type II GAGCAGAAGAACTGCAAGAAAGTGGATGCGTATAACATCGGCCTGGA 35 Cas coding CATAGGCACCAATAGCGTGGGCTGGGCCGTGACCGACTTGGACAAC sequence (nt) AACATCCTGAAGCGGGGCAACAAGAACATGTGGGGCGCCAGAATCTT (human TGACGAGGGAAGCACCGCAGCCCAGACAAGAATGTTCAGAGGGACC codon- CGGCGGAGAATCGAAAGACGGAAGGAGAGAATCAACATCCTGCAGT optimized; CCCTGCTGCTGGACGACATGGAAAAGGAATACCCTAACTTCTTCCCT lacking N- ATGCTGCGGGAAACCAGCAAGACAAAAGAAGATAAGAACAGCGATTC terminal TATCAACGGCAAAAAATACAATCTGTTTTCTGAACTAGAACTCAGCGA methionine; CCCTATCTACTACCAGAAGTATCCTACCATCTATCACCTGAGAAGAGA no stop GCTGATGGAAAGCAAGGAGAAGCACGACATCAGACTGGTCTACCTAG codon) CCATTCACCATATCATCAAGTATAGAGGCAATTTCCTGTACGAGGGCG ACCTGAAAAACAACAACAACGAAATCATCGAGAGCATCAACGAGCTG CTGGATTTCATCCAGAACACCGTGGAAATCAGCTTCATAAGCGAGGC CGAGAAGATCCAGGAGATACTGAAGGACAAAACCAAGAGCAAAGGC GAGAAGAAAGACCTGATCATCGGCCTGTTTGACTATGATAAGGACGA TABLE 1F BCZZ Type II Cas Sequences Name Sequence SEQ ID NO GAAAAGCGTGATCACAGGCATCGTGAACGCCATCCTGGGCTACAAGT TTGACGTGAACAAGATTTTCGATATCGCTGTTGAAAACAGCAATATCA GCTTCAGCAATGAGATCGCCAACGAGGAAGAAATCAAGGAAAACCTG CAGGACCAGGCCTACGTGTACGATGTGCTGCTGAAAATTTACAACTG GTTCATACTGCAGGATATCCTGCAGGGAAACAACTCCATCAGCGAGG CCTTTATTAACAAGTACGACAAGTACAAAAAGGATCTGAGCAACATCA AAAGCCTGTACAGAAATTACCTGAAGGATGAGTACAACAGCATGTTTC GAAGAGAGAAGGACGACTCCTACTCATGCTTCGAGAAGCACATTAGC GCTTGTCCAATCGAGAACTTATACAAGCGTATCAAGAAGGACCTGGC TGACGTTCCCGACTGCAAGGAGAAGGAAGAAATCCTGAAAGACATCG AAGATAATGCCTTTCTGGTGAAGATCAATACCACAGCTAACAGTGCCA TCCCCTACCAACTGCACTACCAGGAACTGGAAAAGATCCTTGAAAAC CAGGCCCAGTACTACCAGACCATCCAGGAAAACAAACAACACATCCT GGAGCTGATGAAGTTCAGAATCCCTTACTACGTCGGACCTCTGGCTC AGGAGGGCAGAAGCAGATTCGCCTGGATCGTGCGGAACAGCAACGA GAAGATACTGCCTTGGACCTTCGATGATGTGGTGAATGTGGACGAGA CAGCCGAGAAATTTATCCGGAGAATGACCAACAAATGTACCTACCTG ATTAACGAGGACGTTATCCCCAAACAGAGCATCCTGTACAGCGAGTT CTGCGTGCTGAACGAGCTGGCCAACATTCGGGTGAACAATCACAAGC TGACTCCTAAGACCAAGCAACTGATCATCGACAAACTGTTCAAGGTGA AGAAAACCATTAAGCAGAGAGATCTAAAGCAGGTGCTGACAGATTTC CAAGTGTTCCCAGAGGTCACCAGCATCACCGGCCTCGCTGAAGTGG ATAAATTCATGAGCAACATGAGCAGCTACATTGACATCGAGAACATCC TGGGCTGCATCAACGACGAGAATATCGAGATGGTGGAGAACATCATT GAGTGGATCACTATCTTTGAGGACAAGAAGATCCTGAAGCGGAAAAT CAAGAACACCTATAACCTGGAAGATAAGGTGGTGAGCCAGTTGATCA AGAAGAACTACTCTGGCTGGAGCCGGCTGTCTAAGGAGCTGCTCGT GGGCCTCAAGGCCGCCGATGACGGCAAGAGCATCATGGAGAAGCTG AGAACCACCAAGGATAATTTCATGCAGATCATCAACAATGACATGTAC GGCTTCAACAAGCAGATCGAGAGCAGGATGAAACTGCAGGAGAAGC AGATCACCTACAAGGACGTGGATGAGATCCCCACAAGCCCCGCTAAC AAGCGGGCCATCTGGCAGAGCCTGAAGATCGTCGAAGAGATCAAGA AAGTCATGAAAAAAGACCCCAAGAACATTTATATCGAGTTCGCCAGG GAAGAACAGGAGAAGAACAAGAGGAAGGACAACAGAGCCAAGGCTC TGTTGAAGATCTACGAGAAGTACGAAGAAGAGATCAAGATGCTGAAG GACTACAACCCCAACGTGTACAAGGAGCTTAAGAAGAAGCAAAATGA GAAAGACTTCAATGAGCGGCTGTACCTGTACTTCATCCAAAACGGCC GGTGCATGTACTCCAGCCGGCCTCTGAATATCGACACCCTGTACTTA TACGAAGTGGACCACATCCTGCCTCAGTCCTACGTGAAGGACGACAG CCTGAGCAACAAGGCCCTGGTGTACAAGGAAGAAAATCAAAGAAAGA GCGGCTCTCTGCTGCTGGACGAGAAGATCATCAACCGGCAGGAAAC CTGGTGGAAGCAGCTGAGAAAAAATGGACTGATCGACGAGAAGAAGT ACTACAACCTGACAAGACGCAAAATGTTCGAAACCGACAAAGAAAAG GTGAAGTTCGTGAGCAGACAGCTGATCGAGACAAGACAGTCTACCAA ATACATCACCAACCTCCTCGTGAACCAGTACAGCAACACCGATGTGTT CGCCATCCGGAGCAACCTGACCCATAACTTCAGAAAGTTCTTCGAGG TCTACAAGAATAGAAACGTGAACGATTACCACCACGCCCAGGACGCC TACATCATCAGCGTGATCGGCAATGTGATCGATCAGAAGCTGCAGTA CAAGGACGAATACAAGTACACAGAGTACGTGAAGAAGTACATCAAGA AGTCTGAAGAGGAAGCCAACAAAGTGTGGATCATCATGGGCATGGTG AGCAATAACATCAATAAAGAAAAGGTTAAGAAGGCCCTGGAATACAAG GATTGTTTTATCACCCACAAGCTGGAGGAACAGACCGGGGCCTTCTA CAACCAGACACTGTACAGTCCTAAGGATAAGAACATCGAAATCCCACT GAAGGAAGGCCTGGATGTGAAGAAGTACGGCGGATACAGCGGAGAG ATCAAGGCTTATTTCACCATCTACTCTTATATCGCCCAGAAGGGCAAG ATGCAGATCGAGATGATCGGAATCCCTGTGAAAACCAAATATGACATC GAGAACGGCAAGACCACACTCATCAAGTACATCCAGAGCAAGAACTC TABLE 1F BCZZ Type II Cas Sequences Name Sequence SEQ ID NO CGAAGCCAGCAAGATCGAGATCATTAGACCTAAGATCCTGAAGTACC AGGAGTACATAGACGAGAACAACGAGCCTATGATGCTGCTGTCTGAC TCCGAAATCAGAACCAATAGACAGCTGATCATCAACAATGAGATGAG CAAGCTCATCTACATCATGAACCAGGAGAAGATCGCCGACGACGAGA AAGAGAAAGTGCAGGAGAAGATGGAGGAGATCTACAACTACCTTCTG GGCAAGCTGAAGAATGAGTACAAAACCTTCAACTCCATCTACCAGAAA CTGTCCAACGAGGAAACAAAGGAGATCTTCCAGAAGCTGGAGTATAA GGACAAGGTGTCAGCTATCAACGGCATCATCGATCTGATGCACCGAG GACAGGGCAACCTGACCAAGCTGAAGCAGGGCGATCGGGCCGGCA GAAAATCAGGCAAGAGTTTTAAGACAGAGGCTCTGAACAAAATGATCT TCATAGACAAAAGCGTGACAGGAATGTACGAGAGAAGATACAAGGTG AACGGCATGGAGAACTGCAGCTCCAAG BCZZ Type II ATGGGCAAACCTATCCCCAATCCCCTGCTGGGCCTGGACAGCACCAA 36 Cas GCGGACTGCCGACGGCAGTGAGTTCGAGTCTCCCAAGAAAAAGAGG mammalian AAAGTGGGATCCGAGCAGAAGAACTGCAAGAAAGTGGATGCGTATAA expression CATCGGCCTGGACATAGGCACCAATAGCGTGGGCTGGGCCGTGACC construct GACTTGGACAACAACATCCTGAAGCGGGGCAACAAGAACATGTGGG (with N- GCGCCAGAATCTTTGACGAGGGAAGCACCGCAGCCCAGACAAGAAT terminal GTTCAGAGGGACCCGGCGGAGAATCGAAAGACGGAAGGAGAGAATC methionine, AACATCCTGCAGTCCCTGCTGCTGGACGACATGGAAAAGGAATACCC stop codon, TAACTTCTTCCCTATGCTGCGGGAAACCAGCAAGACAAAAGAAGATAA V5 tag, N- GAACAGCGATTCTATCAACGGCAAAAAATACAATCTGTTTTCTGAACT terminal NLS AGAACTCAGCGACCCTATCTACTACCAGAAGTATCCTACCATCTATCA and C- CCTGAGAAGAGAGCTGATGGAAAGCAAGGAGAAGCACGACATCAGA terminal NLS) CTGGTCTACCTAGCCATTCACCATATCATCAAGTATAGAGGCAATTTC (nt) CTGTACGAGGGCGACCTGAAAAACAACAACAACGAAATCATCGAGAG CATCAACGAGCTGCTGGATTTCATCCAGAACACCGTGGAAATCAGCT TCATAAGCGAGGCCGAGAAGATCCAGGAGATACTGAAGGACAAAACC AAGAGCAAAGGCGAGAAGAAAGACCTGATCATCGGCCTGTTTGACTA TGATAAGGACGAGAAAAGCGTGATCACAGGCATCGTGAACGCCATCC TGGGCTACAAGTTTGACGTGAACAAGATTTTCGATATCGCTGTTGAAA ACAGCAATATCAGCTTCAGCAATGAGATCGCCAACGAGGAAGAAATC AAGGAAAACCTGCAGGACCAGGCCTACGTGTACGATGTGCTGCTGAA AATTTACAACTGGTTCATACTGCAGGATATCCTGCAGGGAAACAACTC CATCAGCGAGGCCTTTATTAACAAGTACGACAAGTACAAAAAGGATCT GAGCAACATCAAAAGCCTGTACAGAAATTACCTGAAGGATGAGTACA ACAGCATGTTTCGAAGAGAGAAGGACGACTCCTACTCATGCTTCGAG AAGCACATTAGCGCTTGTCCAATCGAGAACTTATACAAGCGTATCAAG AAGGACCTGGCTGACGTTCCCGACTGCAAGGAGAAGGAAGAAATCCT GAAAGACATCGAAGATAATGCCTTTCTGGTGAAGATCAATACCACAGC TAACAGTGCCATCCCCTACCAACTGCACTACCAGGAACTGGAAAAGA TCCTTGAAAACCAGGCCCAGTACTACCAGACCATCCAGGAAAACAAA CAACACATCCTGGAGCTGATGAAGTTCAGAATCCCTTACTACGTCGG ACCTCTGGCTCAGGAGGGCAGAAGCAGATTCGCCTGGATCGTGCGG AACAGCAACGAGAAGATACTGCCTTGGACCTTCGATGATGTGGTGAA TGTGGACGAGACAGCCGAGAAATTTATCCGGAGAATGACCAACAAAT GTACCTACCTGATTAACGAGGACGTTATCCCCAAACAGAGCATCCTG TACAGCGAGTTCTGCGTGCTGAACGAGCTGGCCAACATTCGGGTGAA CAATCACAAGCTGACTCCTAAGACCAAGCAACTGATCATCGACAAACT GTTCAAGGTGAAGAAAACCATTAAGCAGAGAGATCTAAAGCAGGTGC TGACAGATTTCCAAGTGTTCCCAGAGGTCACCAGCATCACCGGCCTC GCTGAAGTGGATAAATTCATGAGCAACATGAGCAGCTACATTGACATC GAGAACATCCTGGGCTGCATCAACGACGAGAATATCGAGATGGTGGA GAACATCATTGAGTGGATCACTATCTTTGAGGACAAGAAGATCCTGAA GCGGAAAATCAAGAACACCTATAACCTGGAAGATAAGGTGGTGAGCC AGTTGATCAAGAAGAACTACTCTGGCTGGAGCCGGCTGTCTAAGGAG CTGCTCGTGGGCCTCAAGGCCGCCGATGACGGCAAGAGCATCATGG TABLE 1F BCZZ Type II Cas Sequences Name Sequence SEQ ID NO AGAAGCTGAGAACCACCAAGGATAATTTCATGCAGATCATCAACAATG ACATGTACGGCTTCAACAAGCAGATCGAGAGCAGGATGAAACTGCAG GAGAAGCAGATCACCTACAAGGACGTGGATGAGATCCCCACAAGCC CCGCTAACAAGCGGGCCATCTGGCAGAGCCTGAAGATCGTCGAAGA GATCAAGAAAGTCATGAAAAAAGACCCCAAGAACATTTATATCGAGTT CGCCAGGGAAGAACAGGAGAAGAACAAGAGGAAGGACAACAGAGCC AAGGCTCTGTTGAAGATCTACGAGAAGTACGAAGAAGAGATCAAGAT GCTGAAGGACTACAACCCCAACGTGTACAAGGAGCTTAAGAAGAAGC AAAATGAGAAAGACTTCAATGAGCGGCTGTACCTGTACTTCATCCAAA ACGGCCGGTGCATGTACTCCAGCCGGCCTCTGAATATCGACACCCTG TACTTATACGAAGTGGACCACATCCTGCCTCAGTCCTACGTGAAGGA CGACAGCCTGAGCAACAAGGCCCTGGTGTACAAGGAAGAAAATCAAA GAAAGAGCGGCTCTCTGCTGCTGGACGAGAAGATCATCAACCGGCA GGAAACCTGGTGGAAGCAGCTGAGAAAAAATGGACTGATCGACGAG AAGAAGTACTACAACCTGACAAGACGCAAAATGTTCGAAACCGACAA AGAAAAGGTGAAGTTCGTGAGCAGACAGCTGATCGAGACAAGACAGT CTACCAAATACATCACCAACCTCCTCGTGAACCAGTACAGCAACACC GATGTGTTCGCCATCCGGAGCAACCTGACCCATAACTTCAGAAAGTT CTTCGAGGTCTACAAGAATAGAAACGTGAACGATTACCACCACGCCC AGGACGCCTACATCATCAGCGTGATCGGCAATGTGATCGATCAGAAG CTGCAGTACAAGGACGAATACAAGTACACAGAGTACGTGAAGAAGTA CATCAAGAAGTCTGAAGAGGAAGCCAACAAAGTGTGGATCATCATGG GCATGGTGAGCAATAACATCAATAAAGAAAAGGTTAAGAAGGCCCTG GAATACAAGGATTGTTTTATCACCCACAAGCTGGAGGAACAGACCGG GGCCTTCTACAACCAGACACTGTACAGTCCTAAGGATAAGAACATCG AAATCCCACTGAAGGAAGGCCTGGATGTGAAGAAGTACGGCGGATAC AGCGGAGAGATCAAGGCTTATTTCACCATCTACTCTTATATCGCCCAG AAGGGCAAGATGCAGATCGAGATGATCGGAATCCCTGTGAAAACCAA ATATGACATCGAGAACGGCAAGACCACACTCATCAAGTACATCCAGA GCAAGAACTCCGAAGCCAGCAAGATCGAGATCATTAGACCTAAGATC CTGAAGTACCAGGAGTACATAGACGAGAACAACGAGCCTATGATGCT GCTGTCTGACTCCGAAATCAGAACCAATAGACAGCTGATCATCAACAA TGAGATGAGCAAGCTCATCTACATCATGAACCAGGAGAAGATCGCCG ACGACGAGAAAGAGAAAGTGCAGGAGAAGATGGAGGAGATCTACAA CTACCTTCTGGGCAAGCTGAAGAATGAGTACAAAACCTTCAACTCCAT CTACCAGAAACTGTCCAACGAGGAAACAAAGGAGATCTTCCAGAAGC TGGAGTATAAGGACAAGGTGTCAGCTATCAACGGCATCATCGATCTG ATGCACCGAGGACAGGGCAACCTGACCAAGCTGAAGCAGGGCGATC GGGCCGGCAGAAAATCAGGCAAGAGTTTTAAGACAGAGGCTCTGAAC AAAATGATCTTCATAGACAAAAGCGTGACAGGAATGTACGAGAGAAG ATACAAGGTGAACGGCATGGAGAACTGCAGCTCCAAGTCTAGAAAGC GGACAGCAGACGGCTCCGAATTTGAAAGCCCTAAGAAAAAGAGAAAG GTGTGA [0091] In some embodiments a BCZZ Type II Cas protein comprises an amino acid sequence of SEQ ID NO:31, SEQ ID NO:32, or SEQ ID NO:33. In some embodiments, a BCZZ Type II Cas protein has nickase activity, for example resulting from one or more amino acid substitutions relative to the sequence of SEQ ID NO:31, SEQ ID NO:32, or SEQ ID NO:33. In some embodiments, the one or more amino acid substitutions providing nickase activity comprise a D17A substitution, wherein the position of the D17A substitution is defined with respect to the amino acid numbering of SEQ ID NO:32. In some embodiments, the one or more amino acid substitutions providing nickase activity comprise an H836A substitution, wherein the position of the H836A substitution is defined with respect to the amino acid numbering of SEQ ID NO:32. In some embodiments, a BCZZ Type II Cas protein is catalytically inactive, for example due to a D17A substitution in combination with a H836A substitution. 6.2.1.7. DRCY Type II Cas Proteins [0092] In one aspect, the disclosure provides DRCY Type II Cas proteins. DRCY Type II Cas proteins can be further classified as Type IIA Cas proteins. The DRCY Type II Cas proteins typically comprise an amino acid sequence that is at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 85%, at least 90%, or at least 95% identical to SEQ ID NO:37. In some embodiments, the DRCY Type II Cas proteins comprise an amino acid sequence that is at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical to SEQ ID NO:37. In some embodiments, a DRCY Type II Cas protein comprises an amino acid sequence that is identical to SEQ ID NO:37. [0093] Exemplary DRCY Type II Cas protein sequences and nucleotide sequences encoding exemplary DRCY Type II Cas proteins are set forth in Table 1G. TABLE 1G DRCY Type II Cas Sequences Name Sequence SEQ ID NO DRCY Type II NLRNFKGDYSVGLDMGTGSVGWSVTDEDGNLLHFKKQPTWGSRLFDS 37 Cas amino AQTAAEARTPRGQRRRYVRRRWRLDLLQSLLQEEIERVDPDFFLRLRQS acid RLVNTDPNKTTSDYRWPLFNGSDFTEVDYYNKFPTIYHLRKWLMETDEQ sequence ADIRLVYLALHNIVKHRGNFLREGQKLSSENANPDDAVSEFCTALKEWCE (without N- GQEGDCGALKEADIVAVLTARGMRPSDCAADLLELLPVSFGDGDKQAD terminal KKLKKALASAMVGLKAEFKDIFGDFATEGTKISLSKDEESEALLEVCPDE methionine) GKVLFESLQRVHSAYVLQGLLSYAPGESISANMIAKYDQYAKDLRMLKLL VREYAPTSYGDFFRGPLCPNKGGYDKFDASVKGYTRYNLGVTSYDDFA KEVKKLFVGTGAEDDERYESMMDAFDDQRFLRRLKTSDNGSICYQLHLE EMETILDNQGRFYPTLANEKDKLLSLVTFRIPYYVGPLTQRNAREDAHGK LRFAWSERKPGTEGVTITPWNWEDVIDKGASAEKFITRMTGICTYLQGE DVLPKSSLLYEEFCVLNELNGMRWTSDGDEERRFDAEQREGMMRDLFR KTRTVSNKRIEDWLVQNGYAHSPKVSGGQNETGLESKLGSYIFFAKDIFG TDEIAKVDYPMIEEIILWNTLFEDRSILKEKLREKYGDAGEGRLSAEQIKKI CKKRFTGWGRLSKKFLAGIKVDTQLGEKSIMDVLREGDPDADIRRGRTM VLMEVIRDDNLGFQKKIDSFNRAYFAEAGSALGVNDLPGSPAIRRSLNQA IRIVDEIAHIAGHAPTNVFIEVTRDDDMGKKGHRTARRYDQLKEALAAFKK DDPQIWKEIQAASPTDLDERLTLYFVQRGKCMYSGKPIDINQLSNAGLYE VDHIIPRSYIKDDSFENKALVLREFNQRKTDSMLLDDGIRRNMAGYWRSL HDAKLIGDKKFNNLMRTHVGENAMRGFIARQLVETSQMVKLAQSLLSAK YPDTKITPVKASMSHDLREAAGFVKCREANDFHHAHDAFLACRVGLFIRK WYPDMYDKPILYTRAMRKYVREQAEEFKKTRRAPGSSGFVVGRFISSFV DADTGELWEGAEEVESIRRVLNYRQCHITRMPMEDSGAFWNATIYSPRD PKMGSKLSLRLKGDLDPQIYGGYSSQQFAYFFIYEARDKKGRPAFRFSQ VPVWLASRIASDEWALEEYAREQAASEQLEFVRIQRSKILKKQLVEIDGE RLIITGAEEVRNARQFAFSLDELSLLGRASARGDGEAIDEETISNGYNVLL KVMIERGELVGKGLMKKLSLGDRYEQFMQLDCAAKKSLILQLAAIVNASN RQIDLSKIGGAKTAGQYKVTYNKLLNDPNTDFYIVDQSVTGMFERRTRVG L DRCY Type II MNLRNFKGDYSVGLDMGTGSVGWSVTDEDGNLLHFKKQPTWGSRLFD 38 Cas amino SAQTAAEARTPRGQRRRYVRRRWRLDLLQSLLQEEIERVDPDFFLRLRQ acid SRLVNTDPNKTTSDYRWPLFNGSDFTEVDYYNKFPTIYHLRKWLMETDE sequence QADIRLVYLALHNIVKHRGNFLREGQKLSSENANPDDAVSEFCTALKEW CEGQEGDCGALKEADIVAVLTARGMRPSDCAADLLELLPVSFGDGDKQ ADKKLKKALASAMVGLKAEFKDIFGDFATEGTKISLSKDEESEALLEVCP DEGKVLFESLQRVHSAYVLQGLLSYAPGESISANMIAKYDQYAKDLRML KLLVREYAPTSYGDFFRGPLCPNKGGYDKFDASVKGYTRYNLGVTSYD DFAKEVKKLFVGTGAEDDERYESMMDAFDDQRFLRRLKTSDNGSICYQL HLEEMETILDNQGRFYPTLANEKDKLLSLVTFRIPYYVGPLTQRNAREDA HGKLRFAWSERKPGTEGVTITPWNWEDVIDKGASAEKFITRMTGICTYL QGEDVLPKSSLLYEEFCVLNELNGMRWTSDGDEERRFDAEQREGMMR TABLE 1G DRCY Type II Cas Sequences Name Sequence SEQ ID NO DLFRKTRTVSNKRIEDWLVQNGYAHSPKVSGGQNETGLESKLGSYIFFA KDIFGTDEIAKVDYPMIEEIILWNTLFEDRSILKEKLREKYGDAGEGRLSAE QIKKICKKRFTGWGRLSKKFLAGIKVDTQLGEKSIMDVLREGDPDADIRR GRTMVLMEVIRDDNLGFQKKIDSFNRAYFAEAGSALGVNDLPGSPAIRR SLNQAIRIVDEIAHIAGHAPTNVFIEVTRDDDMGKKGHRTARRYDQLKEAL AAFKKDDPQIWKEIQAASPTDLDERLTLYFVQRGKCMYSGKPIDINQLSN AGLYEVDHIIPRSYIKDDSFENKALVLREFNQRKTDSMLLDDGIRRNMAG YWRSLHDAKLIGDKKFNNLMRTHVGENAMRGFIARQLVETSQMVKLAQ SLLSAKYPDTKITPVKASMSHDLREAAGFVKCREANDFHHAHDAFLACR VGLFIRKWYPDMYDKPILYTRAMRKYVREQAEEFKKTRRAPGSSGFVVG RFISSFVDADTGELWEGAEEVESIRRVLNYRQCHITRMPMEDSGAFWNA TIYSPRDPKMGSKLSLRLKGDLDPQIYGGYSSQQFAYFFIYEARDKKGRP AFRFSQVPVWLASRIASDEWALEEYAREQAASEQLEFVRIQRSKILKKQL VEIDGERLIITGAEEVRNARQFAFSLDELSLLGRASARGDGEAIDEETISN GYNVLLKVMIERGELVGKGLMKKLSLGDRYEQFMQLDCAAKKSLILQLA AIVNASNRQIDLSKIGGAKTAGQYKVTYNKLLNDPNTDFYIVDQSVTGMF ERRTRVGL DRCY Type II MGKPIPNPLLGLDSTKRTADGSEFESPKKKRKVGSNLRNFKGDYSVGLD 39 Cas MGTGSVGWSVTDEDGNLLHFKKQPTWGSRLFDSAQTAAEARTPRGQR mammalian RRYVRRRWRLDLLQSLLQEEIERVDPDFFLRLRQSRLVNTDPNKTTSDY expression RWPLFNGSDFTEVDYYNKFPTIYHLRKWLETDEQADIRLVYLALHNIVKH construct RGNFLREGQKLSSENANPDDAVSEFCTALKEWCEGQEGDCGALKEADI (with N- VAVLTARGMRPSDCAADLLELLPVSFGDGDKQADKKLKKALASAMVGLK terminal AEFKDIFGDFATEGTKISLSKDEESEALLEVCPDEGKVLFESLQRVHSAY methionine, VLQGLLSYAPGESISANMIAKYDQYAKDLRMLKLLVREYAPTSYGDFFRG stop codon, PLCPNKGGYDKFDASVKGYTRYNLGVTSYDDFAKEVKKLFVGTGAEDD V5 tag, N- ERYESMMDAFDDQRFLRRLKTSDNGSICYQLHLEEMETILDNQGRFYPT terminal NLS LANEKDKLLSLVTFRIPYYVGPLTQRNAREDAHGKLRFAWSERKPGTEG and C- VTITPWNWEDVIDKGASAEKFITRMTGICTYLQGEDVLPKSSLLYEEFCVL terminal NLS) NELNGMRWTSDGDEERRFDAEQREGMMRDLFRKTRTVSNKRIEDWLV (aa) QNGYAHSPKVSGGQNETGLESKLGSYIFFAKDIFGTDEIAKVDYPMIEEIIL WNTLFEDRSILKEKLREKYGDAGEGRLSAEQIKKICKKRFTGWGRLSKKF LAGIKVDTQLGEKSIMDVLREGDPDADIRRGRTMVLMEVIRDDNLGFQKK IDSFNRAYFAEAGSALGVNDLPGSPAIRRSLNQAIRIVDEIAHIAGHAPTNV FIEVTRDDDMGKKGHRTARRYDQLKEALAAFKKDDPQIWKEIQAASPTD LDERLTLYFVQRGKCMYSGKPIDINQLSNAGLYEVDHIIPRSYIKDDSFEN KALVLREFNQRKTDSMLLDDGIRRNMAGYWRSLHDAKLIGDKKFNNLMR THVGENAMRGFIARQLVETSQMVKLAQSLLSAKYPDTKITPVKASMSHD LREAAGFVKCREANDFHHAHDAFLACRVGLFIRKWYPDMYDKPILYTRA MRKYVREQAEEFKKTRRAPGSSGFVVGRFISSFVDADTGELWEGAEEV ESIRRVLNYRQCHITRMPMEDSGAFWNATIYSPRDPKMGSKLSLRLKGD LDPQIYGGYSSQQFAYFFIYEARDKKGRPAFRFSQVPVWLASRIASDEW ALEEYAREQAASEQLEFVRIQRSKILKKQLVEIDGERLIITGAEEVRNARQ FAFSLDELSLLGRASARGDGEAIDEETISNGYNVLLKVMIERGELVGKGL MKKLSLGDRYEQFMQLDCAAKKSLILQLAAIVNASNRQIDLSKIGGAKTA GQYKVTYNKLLNDPNTDFYIVDQSVTGMFERRTRVGLSRKRTADGSEFE SPKKKRKV DRCY Type II GTTAATTTGCGGAATTTCAAGGGCGACTACAGCGTTGGTCTCGATAT 40 Cas coding GGGAACAGGCTCCGTTGGCTGGTCGGTGACGGACGAAGACGGCAAC sequence (nt) CTCCTTCACTTTAAAAAGCAACCAACATGGGGAAGCCGTTTGTTCGAT (not codon AGTGCTCAAACGGCAGCCGAGGCACGCACCCCTCGTGGGCAACGAA optimized) GGCGTTATGTTCGACGTCGTTGGCGACTCGACCTGCTGCAGAGTCTA CTCCAGGAAGAAATTGAACGTGTCGACCCAGATTTCTTCCTGCGTCTT CGCCAATCGCGTCTCGTAAATACGGACCCCAATAAGACGACCTCGGA TTATCGCTGGCCACTATTTAACGGAAGCGATTTTACCGAGGTCGATTA CTATAACAAGTTTCCAACTATCTATCATCTGCGTAAATGGCTGATGGA AACCGATGAGCAAGCTGATATCCGTCTTGTTTATTTGGCGCTTCACAA TABLE 1G DRCY Type II Cas Sequences Name Sequence SEQ ID NO CATAGTGAAGCATCGTGGTAACTTCTTGCGCGAAGGGCAGAAACTTT CTTCGGAAAATGCCAACCCTGATGACGCGGTTTCGGAATTTTGCACT GCGCTGAAAGAGTGGTGCGAAGGTCAAGAAGGTGACTGTGGGGCAC TCAAAGAGGCAGATATCGTTGCGGTTCTTACGGCGAGAGGCATGCGT CCGTCTGATTGCGCCGCTGATCTTTTGGAGCTACTCCCAGTTTCTTTT GGCGATGGTGACAAGCAGGCTGATAAGAAGCTTAAGAAAGCGCTCG CTAGCGCTATGGTTGGCTTGAAAGCTGAGTTCAAAGACATATTCGGT GATTTCGCTACTGAAGGAACGAAGATATCGCTCTCGAAGGACGAAGA ATCAGAAGCGCTGCTTGAAGTGTGCCCTGATGAGGGTAAGGTGCTTT TCGAGTCGCTGCAACGAGTTCATTCCGCATATGTACTCCAAGGGTTG CTATCTTATGCTCCTGGTGAATCAATTTCGGCAAATATGATTGCGAAG TACGATCAGTATGCTAAAGACTTGAGGATGCTGAAGTTACTGGTTCGC GAATATGCGCCAACGTCGTATGGCGACTTCTTCCGTGGTCCGCTTTG TCCGAATAAGGGCGGCTATGACAAGTTTGATGCTAGTGTTAAGGGCT ACACGCGCTACAACCTTGGAGTCACTTCATACGATGACTTTGCCAAA GAAGTAAAAAAGCTATTCGTAGGCACGGGTGCCGAGGACGATGAGC GCTACGAGTCTATGATGGACGCCTTCGATGACCAGCGATTCCTTCGT CGATTGAAAACGAGCGACAATGGAAGCATCTGTTATCAGCTGCACCT CGAAGAGATGGAGACGATCCTCGATAATCAGGGACGCTTCTATCCGA CCCTCGCGAATGAGAAGGATAAGCTATTGTCGCTGGTCACCTTCCGC ATTCCGTATTATGTGGGGCCGCTTACGCAGCGCAATGCGCGTGAGGA CGCACATGGAAAGCTTCGCTTCGCGTGGTCCGAACGCAAGCCGGGG ACCGAAGGTGTGACGATTACGCCTTGGAACTGGGAAGATGTTATCGA CAAGGGCGCGAGTGCCGAGAAATTCATCACGCGGATGACGGGTATA TGTACTTACTTGCAGGGCGAGGACGTGCTGCCGAAGTCGTCTCTTCT CTATGAGGAATTTTGCGTTCTGAATGAACTGAACGGTATGCGCTGGA CGTCCGATGGCGACGAAGAACGGCGATTCGATGCCGAGCAGCGTGA AGGGATGATGCGCGACTTGTTCCGCAAGACGCGCACGGTTTCGAATA AGAGGATTGAAGATTGGCTCGTTCAGAACGGGTACGCGCATTCTCCG AAGGTGTCGGGCGGGCAGAACGAAACAGGTTTGGAATCTAAACTCG GATCCTACATTTTCTTCGCTAAGGATATCTTCGGAACAGATGAAATTG CCAAAGTCGATTATCCGATGATTGAAGAGATCATTCTATGGAACACCT TGTTCGAAGATCGTTCGATCCTGAAAGAGAAGCTGCGAGAAAAGTAT GGCGATGCAGGTGAAGGCCGTTTGAGCGCTGAACAAATCAAGAAGAT TTGCAAAAAACGCTTCACGGGGTGGGGTCGCTTGTCGAAGAAGTTCT TGGCGGGGATTAAAGTCGATACGCAGCTTGGTGAGAAGTCGATTATG GACGTGCTGCGTGAAGGCGATCCGGATGCTGATATTCGACGTGGTC GCACGATGGTACTTATGGAAGTAATTCGCGACGATAACCTGGGTTTC CAAAAGAAGATCGACTCATTTAACCGTGCGTACTTTGCCGAGGCTGG CAGTGCGCTTGGCGTGAACGATCTTCCAGGGTCGCCCGCAATTCGTC GCAGTCTCAACCAGGCGATTCGCATTGTCGACGAGATTGCTCATATC GCAGGTCATGCTCCGACAAATGTCTTCATCGAGGTCACGCGTGATGA CGACATGGGCAAGAAGGGCCATCGCACTGCCCGTCGCTACGATCAG TTGAAAGAAGCGCTTGCTGCCTTTAAGAAGGACGATCCTCAAATATG GAAGGAGATACAGGCGGCCAGTCCAACCGATTTGGACGAACGTCTC ACACTGTATTTTGTGCAGCGAGGCAAATGCATGTATTCGGGTAAGCC CATCGATATCAATCAGCTTTCGAACGCTGGACTGTACGAGGTTGACC ATATCATCCCCCGATCCTATATTAAGGACGATAGTTTCGAAAATAAGG CGCTGGTGCTTCGCGAATTCAACCAGCGTAAAACGGATTCGATGTTG CTTGACGACGGCATCCGCCGCAACATGGCTGGCTATTGGCGCAGTTT GCATGATGCGAAACTTATTGGCGACAAGAAGTTCAATAATCTCATGCG CACGCACGTCGGCGAAAACGCTATGCGCGGATTTATCGCTCGTCAGT TGGTGGAGACAAGCCAAATGGTGAAGCTTGCCCAGTCGCTTCTTTCC GCCAAATATCCTGACACGAAGATCACTCCAGTGAAGGCGAGCATGTC TCATGATTTGCGCGAAGCGGCAGGGTTTGTGAAATGCCGCGAGGCA AATGATTTTCATCATGCACACGATGCCTTTTTGGCGTGCAGAGTGGGT CTTTTTATTCGGAAGTGGTACCCCGACATGTATGACAAGCCCATTCTC TABLE 1G DRCY Type II Cas Sequences Name Sequence SEQ ID NO TATACGCGTGCGATGAGAAAATACGTACGAGAGCAGGCGGAGGAGT TCAAAAAGACTCGTCGGGCACCAGGCTCGAGTGGATTCGTGGTTGG GCGCTTTATAAGTTCGTTCGTTGACGCCGATACTGGTGAACTGTGGG AGGGCGCCGAAGAGGTTGAGAGTATTCGTCGTGTGCTGAATTACCGC CAGTGCCACATCACGCGCATGCCGATGGAAGACTCAGGTGCGTTTTG GAACGCAACTATTTATTCGCCGCGCGACCCGAAGATGGGGAGTAAGC TTTCGTTGCGCTTGAAGGGCGATCTGGACCCACAGATTTATGGTGGC TATTCAAGCCAGCAATTTGCATATTTCTTCATTTACGAGGCGCGCGAT AAGAAGGGGCGCCCTGCGTTCCGTTTCTCCCAGGTACCCGTGTGGC TGGCGTCGCGGATTGCGAGCGACGAATGGGCACTCGAAGAATACGC GCGTGAGCAGGCCGCAAGCGAGCAGCTCGAGTTTGTGCGTATTCAG CGTTCGAAGATCCTGAAGAAGCAGCTGGTAGAGATCGACGGAGAGC GGTTGATTATTACAGGAGCTGAAGAGGTTCGGAATGCTAGGCAGTTT GCGTTTAGTCTGGATGAACTCTCGCTTCTTGGAAGGGCGTCAGCGCG AGGCGATGGCGAGGCAATTGACGAGGAAACTATATCAAATGGATATA ACGTGCTATTGAAAGTAATGATTGAACGTGGCGAGCTGGTTGGAAAG GGCCTTATGAAGAAGCTGTCTCTCGGTGATCGCTACGAGCAATTTAT GCAATTGGACTGCGCGGCAAAGAAATCGCTAATCCTTCAGCTCGCCG CTATTGTTAATGCTTCTAATAGGCAGATTGACTTGTCGAAGATTGGAG GGGCTAAAACGGCCGGTCAGTACAAGGTAACTTATAACAAGCTTCTC AATGACCCTAACACCGATTTCTACATTGTCGATCAGTCGGTGACGGG GATGTTCGAGCGGAGGACGCGCGTTGGCCTTTAG DRCY Type II AACCTGCGCAACTTCAAGGGCGACTACAGCGTCGGACTGGACATGG 41 Cas coding GCACAGGCTCTGTGGGCTGGTCCGTGACCGATGAGGACGGCAATCT sequence (nt) GCTGCACTTCAAAAAGCAGCCTACATGGGGAAGCAGACTGTTCGACA (human GCGCCCAGACCGCCGCTGAAGCCAGAACCCCTAGAGGCCAGCGGC codon- GGCGGTACGTGCGGAGAAGGTGGCGGCTCGACCTGCTGCAAAGCCT optimized; GCTGCAGGAGGAAATCGAGAGAGTGGACCCTGATTTCTTCCTGCGG lacking N- CTGAGACAGAGCAGACTGGTGAATACCGACCCCAACAAGACCACCA terminal GCGATTATAGATGGCCTCTGTTTAACGGCAGCGACTTTACCGAGGTG methionine; GACTACTACAACAAGTTCCCCACAATCTACCACCTGAGAAAGTGGCT no stop GGAAACCGACGAGCAGGCTGACATCCGGCTGGTGTACCTGGCCCTG codon) CACAACATCGTGAAGCACAGAGGCAACTTCCTGCGCGAGGGCCAGA AGCTCAGCTCAGAGAACGCCAATCCTGACGATGCCGTGTCCGAGTTC TGCACCGCCCTGAAGGAGTGGTGTGAAGGCCAGGAGGGAGATTGCG GCGCCCTGAAAGAGGCCGACATCGTGGCCGTGCTTACAGCCCGGGG AATGCGACCCAGCGACTGCGCCGCTGACCTGCTGGAATTGCTGCCT GTGAGCTTCGGCGACGGCGATAAGCAGGCAGACAAGAAACTTAAGA AGGCCCTGGCCAGCGCTATGGTGGGCCTGAAAGCCGAGTTCAAGGA CATCTTCGGCGATTTCGCCACAGAGGGCACTAAGATCAGCCTGTCTA AGGACGAGGAAAGCGAGGCTCTGCTGGAAGTTTGTCCTGACGAGGG CAAGGTCCTGTTCGAGAGCCTGCAGAGAGTCCACTCTGCCTACGTGC TGCAAGGCCTGCTGTCTTATGCCCCTGGCGAGTCTATCTCCGCCAAT ATGATCGCCAAGTACGATCAGTACGCCAAAGATCTGCGGATGCTGAA GCTGCTGGTGCGGGAGTACGCCCCTACCAGCTACGGCGACTTCTTC AGGGGACCTCTGTGCCCCAACAAGGGCGGATACGACAAGTTCGACG CCAGCGTGAAAGGCTACACCAGATACAACCTGGGCGTGACCTCCTAC GACGACTTCGCCAAGGAGGTGAAGAAGCTGTTCGTGGGCACCGGAG CTGAGGACGACGAGAGATACGAGAGCATGATGGATGCGTTCGACGA CCAACGGTTCCTGCGGAGACTGAAAACAAGTGATAACGGATCTATCT GCTACCAGCTCCATCTGGAAGAGATGGAGACAATCCTGGACAACCAG GGCAGATTCTACCCTACGCTGGCCAACGAGAAGGACAAGCTGCTGA GCCTCGTGACCTTCCGCATCCCCTACTACGTGGGCCCTCTGACCCAG AGAAACGCTAGAGAGGATGCCCACGGCAAGCTGCGGTTCGCTTGGA GCGAGAGAAAACCTGGCACCGAGGGCGTGACAATTACACCCTGGAA CTGGGAGGATGTGATCGACAAGGGCGCCAGCGCCGAGAAGTTCATC ACCAGAATGACCGGAATCTGCACCTACCTGCAGGGAGAAGATGTTCT TABLE 1G DRCY Type II Cas Sequences Name Sequence SEQ ID NO GCCTAAGAGCTCTCTGCTGTATGAGGAGTTCTGCGTGCTGAACGAGC TGAATGGTATGAGATGGACCAGCGACGGCGACGAGGAGAGAAGATT TGATGCCGAGCAACGGGAGGGCATGATGCGGGACCTGTTCAGAAAG ACCAGAACTGTGTCCAACAAGAGAATTGAGGACTGGCTGGTGCAGAA CGGCTACGCCCACAGCCCTAAGGTGAGCGGCGGACAAAACGAGACA GGCCTAGAAAGCAAGCTCGGATCTTACATCTTCTTTGCCAAGGACAT CTTTGGCACTGACGAGATTGCCAAGGTGGATTACCCTATGATCGAGG AGATCATCCTGTGGAACACCCTGTTTGAGGATCGGAGCATCCTCAAA GAGAAGCTAAGAGAGAAGTACGGCGACGCCGGCGAGGGCAGACTGT CAGCCGAGCAGATCAAGAAGATCTGTAAAAAAAGATTCACCGGCTGG GGCAGACTGAGCAAGAAGTTCCTCGCCGGCATCAAGGTGGATACCC AGCTGGGCGAGAAAAGCATCATGGACGTGCTGCGGGAAGGCGATCC CGACGCGGATATCAGACGGGGCCGGACAATGGTGCTGATGGAAGTG ATCCGGGACGACAATCTGGGATTTCAGAAAAAAATCGACAGCTTCAAT AGAGCATATTTCGCCGAGGCCGGTAGCGCTCTGGGCGTTAACGACC TGCCAGGCAGCCCTGCTATCCGGAGAAGCCTGAACCAGGCCATCCG GATCGTGGACGAAATCGCCCATATCGCAGGCCACGCCCCTACAAAC GTGTTCATCGAGGTGACACGGGATGACGACATGGGCAAGAAAGGCC ACAGAACTGCCCGGAGATACGACCAGCTGAAGGAGGCCCTGGCCGC CTTTAAAAAGGACGATCCCCAGATTTGGAAGGAAATCCAGGCCGCCA GTCCTACCGACCTGGACGAGCGGCTGACACTGTACTTCGTCCAGAGA GGAAAATGCATGTACAGCGGAAAGCCTATCGATATTAACCAGCTGAG CAACGCCGGCCTGTATGAAGTGGACCACATCATCCCTAGATCTTACA TCAAGGACGATTCTTTTGAAAATAAGGCCCTGGTGCTGCGGGAGTTC AACCAGAGAAAGACCGATAGCATGCTACTGGATGACGGCATCAGAAG AAACATGGCCGGCTACTGGAGAAGCCTGCACGACGCTAAGCTGATC GGCGATAAGAAGTTCAACAACCTGATGAGAACACACGTGGGTGAGAA CGCCATGAGAGGCTTTATCGCTAGGCAGCTGGTCGAAACCAGCCAG ATGGTCAAGCTGGCCCAGAGCCTGCTGTCCGCTAAGTACCCCGACA CCAAGATCACACCTGTGAAGGCCTCTATGAGCCACGACCTGAGAGAG GCCGCCGGCTTCGTGAAGTGCCGGGAAGCTAACGACTTCCACCACG CCCACGATGCCTTTCTGGCCTGCAGAGTGGGACTGTTTATTAGAAAG TGGTATCCTGACATGTACGACAAGCCCATCCTGTACACCAGGGCCAT GAGAAAGTATGTGCGAGAGCAGGCCGAAGAGTTCAAGAAAACCCGG AGAGCTCCAGGCTCCTCCGGATTCGTGGTGGGCCGGTTTATCAGCA GCTTCGTGGATGCCGACACCGGCGAGCTGTGGGAAGGCGCCGAAGA GGTGGAAAGTATCAGAAGAGTGCTGAACTACAGACAGTGCCACATCA CCCGGATGCCTATGGAGGATAGCGGCGCCTTCTGGAATGCCACAAT CTACAGCCCCAGAGATCCGAAAATGGGCAGCAAATTATCACTGCGCC TGAAGGGCGATCTGGACCCACAGATCTACGGGGGCTATAGCTCTCA GCAGTTTGCCTACTTCTTCATCTACGAGGCCCGGGATAAAAAGGGCA GACCTGCCTTCAGATTCAGCCAAGTGCCAGTGTGGCTGGCAAGCAGA ATCGCCTCCGACGAATGGGCGCTGGAGGAATATGCCAGAGAGCAGG CCGCCAGCGAACAACTGGAGTTCGTGAGAATCCAGCGGTCCAAGAT CCTGAAGAAGCAGCTGGTCGAGATCGATGGTGAGCGGCTCATCATCA CAGGCGCCGAGGAAGTGCGCAATGCCCGGCAGTTCGCTTTTAGCTT GGATGAACTGAGCCTCCTCGGCAGGGCCAGCGCTAGAGGCGACGG CGAAGCAATTGACGAAGAAACCATCTCTAATGGCTACAACGTGCTGC TGAAGGTGATGATCGAACGGGGCGAGCTGGTGGGCAAAGGCCTGAT GAAGAAACTGTCTCTGGGAGACAGATACGAGCAGTTCATGCAGCTGG ATTGTGCCGCCAAGAAGTCCCTGATCCTCCAGCTCGCAGCCATCGTG AACGCCAGCAACAGACAGATCGATCTGAGCAAAATTGGCGGCGCCAA GACCGCCGGCCAGTACAAGGTAACCTACAACAAGCTGCTGAACGAC CCTAATACCGATTTCTACATCGTGGACCAAAGCGTGACCGGCATGTT CGAGCGGAGAACCAGAGTCGGACTG DRCY Type II ATGGGCAAACCTATCCCCAATCCCCTGCTGGGCCTGGACAGCACCAA 42 Cas GCGGACTGCCGACGGCAGTGAGTTCGAGTCTCCCAAGAAAAAGAGG TABLE 1G DRCY Type II Cas Sequences Name Sequence SEQ ID NO mammalian AAAGTGGGATCCAACCTGCGCAACTTCAAGGGCGACTACAGCGTCG expression GACTGGACATGGGCACAGGCTCTGTGGGCTGGTCCGTGACCGATGA construct GGACGGCAATCTGCTGCACTTCAAAAAGCAGCCTACATGGGGAAGCA (with N- GACTGTTCGACAGCGCCCAGACCGCCGCTGAAGCCAGAACCCCTAG terminal AGGCCAGCGGCGGCGGTACGTGCGGAGAAGGTGGCGGCTCGACCT methionine, GCTGCAAAGCCTGCTGCAGGAGGAAATCGAGAGAGTGGACCCTGAT stop codon, TTCTTCCTGCGGCTGAGACAGAGCAGACTGGTGAATACCGACCCCAA V5 tag, N- CAAGACCACCAGCGATTATAGATGGCCTCTGTTTAACGGCAGCGACT terminal NLS TTACCGAGGTGGACTACTACAACAAGTTCCCCACAATCTACCACCTGA and C- GAAAGTGGCTGGAAACCGACGAGCAGGCTGACATCCGGCTGGTGTA terminal NLS) CCTGGCCCTGCACAACATCGTGAAGCACAGAGGCAACTTCCTGCGC (nt) GAGGGCCAGAAGCTCAGCTCAGAGAACGCCAATCCTGACGATGCCG TGTCCGAGTTCTGCACCGCCCTGAAGGAGTGGTGTGAAGGCCAGGA GGGAGATTGCGGCGCCCTGAAAGAGGCCGACATCGTGGCCGTGCTT ACAGCCCGGGGAATGCGACCCAGCGACTGCGCCGCTGACCTGCTGG AATTGCTGCCTGTGAGCTTCGGCGACGGCGATAAGCAGGCAGACAA GAAACTTAAGAAGGCCCTGGCCAGCGCTATGGTGGGCCTGAAAGCC GAGTTCAAGGACATCTTCGGCGATTTCGCCACAGAGGGCACTAAGAT CAGCCTGTCTAAGGACGAGGAAAGCGAGGCTCTGCTGGAAGTTTGTC CTGACGAGGGCAAGGTCCTGTTCGAGAGCCTGCAGAGAGTCCACTC TGCCTACGTGCTGCAAGGCCTGCTGTCTTATGCCCCTGGCGAGTCTA TCTCCGCCAATATGATCGCCAAGTACGATCAGTACGCCAAAGATCTG CGGATGCTGAAGCTGCTGGTGCGGGAGTACGCCCCTACCAGCTACG GCGACTTCTTCAGGGGACCTCTGTGCCCCAACAAGGGCGGATACGA CAAGTTCGACGCCAGCGTGAAAGGCTACACCAGATACAACCTGGGC GTGACCTCCTACGACGACTTCGCCAAGGAGGTGAAGAAGCTGTTCGT GGGCACCGGAGCTGAGGACGACGAGAGATACGAGAGCATGATGGAT GCGTTCGACGACCAACGGTTCCTGCGGAGACTGAAAACAAGTGATAA CGGATCTATCTGCTACCAGCTCCATCTGGAAGAGATGGAGACAATCC TGGACAACCAGGGCAGATTCTACCCTACGCTGGCCAACGAGAAGGA CAAGCTGCTGAGCCTCGTGACCTTCCGCATCCCCTACTACGTGGGCC CTCTGACCCAGAGAAACGCTAGAGAGGATGCCCACGGCAAGCTGCG GTTCGCTTGGAGCGAGAGAAAACCTGGCACCGAGGGCGTGACAATT ACACCCTGGAACTGGGAGGATGTGATCGACAAGGGCGCCAGCGCCG AGAAGTTCATCACCAGAATGACCGGAATCTGCACCTACCTGCAGGGA GAAGATGTTCTGCCTAAGAGCTCTCTGCTGTATGAGGAGTTCTGCGT GCTGAACGAGCTGAATGGTATGAGATGGACCAGCGACGGCGACGAG GAGAGAAGATTTGATGCCGAGCAACGGGAGGGCATGATGCGGGACC TGTTCAGAAAGACCAGAACTGTGTCCAACAAGAGAATTGAGGACTGG CTGGTGCAGAACGGCTACGCCCACAGCCCTAAGGTGAGCGGCGGAC AAAACGAGACAGGCCTAGAAAGCAAGCTCGGATCTTACATCTTCTTTG CCAAGGACATCTTTGGCACTGACGAGATTGCCAAGGTGGATTACCCT ATGATCGAGGAGATCATCCTGTGGAACACCCTGTTTGAGGATCGGAG CATCCTCAAAGAGAAGCTAAGAGAGAAGTACGGCGACGCCGGCGAG GGCAGACTGTCAGCCGAGCAGATCAAGAAGATCTGTAAAAAAAGATT CACCGGCTGGGGCAGACTGAGCAAGAAGTTCCTCGCCGGCATCAAG GTGGATACCCAGCTGGGCGAGAAAAGCATCATGGACGTGCTGCGGG AAGGCGATCCCGACGCGGATATCAGACGGGGCCGGACAATGGTGCT GATGGAAGTGATCCGGGACGACAATCTGGGATTTCAGAAAAAAATCG ACAGCTTCAATAGAGCATATTTCGCCGAGGCCGGTAGCGCTCTGGGC GTTAACGACCTGCCAGGCAGCCCTGCTATCCGGAGAAGCCTGAACC AGGCCATCCGGATCGTGGACGAAATCGCCCATATCGCAGGCCACGC CCCTACAAACGTGTTCATCGAGGTGACACGGGATGACGACATGGGCA AGAAAGGCCACAGAACTGCCCGGAGATACGACCAGCTGAAGGAGGC CCTGGCCGCCTTTAAAAAGGACGATCCCCAGATTTGGAAGGAAATCC AGGCCGCCAGTCCTACCGACCTGGACGAGCGGCTGACACTGTACTT CGTCCAGAGAGGAAAATGCATGTACAGCGGAAAGCCTATCGATATTA TABLE 1G DRCY Type II Cas Sequences Name Sequence SEQ ID NO ACCAGCTGAGCAACGCCGGCCTGTATGAAGTGGACCACATCATCCCT AGATCTTACATCAAGGACGATTCTTTTGAAAATAAGGCCCTGGTGCTG CGGGAGTTCAACCAGAGAAAGACCGATAGCATGCTACTGGATGACG GCATCAGAAGAAACATGGCCGGCTACTGGAGAAGCCTGCACGACGC TAAGCTGATCGGCGATAAGAAGTTCAACAACCTGATGAGAACACACG TGGGTGAGAACGCCATGAGAGGCTTTATCGCTAGGCAGCTGGTCGA AACCAGCCAGATGGTCAAGCTGGCCCAGAGCCTGCTGTCCGCTAAG TACCCCGACACCAAGATCACACCTGTGAAGGCCTCTATGAGCCACGA CCTGAGAGAGGCCGCCGGCTTCGTGAAGTGCCGGGAAGCTAACGAC TTCCACCACGCCCACGATGCCTTTCTGGCCTGCAGAGTGGGACTGTT TATTAGAAAGTGGTATCCTGACATGTACGACAAGCCCATCCTGTACAC CAGGGCCATGAGAAAGTATGTGCGAGAGCAGGCCGAAGAGTTCAAG AAAACCCGGAGAGCTCCAGGCTCCTCCGGATTCGTGGTGGGCCGGT TTATCAGCAGCTTCGTGGATGCCGACACCGGCGAGCTGTGGGAAGG CGCCGAAGAGGTGGAAAGTATCAGAAGAGTGCTGAACTACAGACAGT GCCACATCACCCGGATGCCTATGGAGGATAGCGGCGCCTTCTGGAA TGCCACAATCTACAGCCCCAGAGATCCGAAAATGGGCAGCAAATTAT CACTGCGCCTGAAGGGCGATCTGGACCCACAGATCTACGGGGGCTA TAGCTCTCAGCAGTTTGCCTACTTCTTCATCTACGAGGCCCGGGATAA AAAGGGCAGACCTGCCTTCAGATTCAGCCAAGTGCCAGTGTGGCTG GCAAGCAGAATCGCCTCCGACGAATGGGCGCTGGAGGAATATGCCA GAGAGCAGGCCGCCAGCGAACAACTGGAGTTCGTGAGAATCCAGCG GTCCAAGATCCTGAAGAAGCAGCTGGTCGAGATCGATGGTGAGCGG CTCATCATCACAGGCGCCGAGGAAGTGCGCAATGCCCGGCAGTTCG CTTTTAGCTTGGATGAACTGAGCCTCCTCGGCAGGGCCAGCGCTAGA GGCGACGGCGAAGCAATTGACGAAGAAACCATCTCTAATGGCTACAA CGTGCTGCTGAAGGTGATGATCGAACGGGGCGAGCTGGTGGGCAAA GGCCTGATGAAGAAACTGTCTCTGGGAGACAGATACGAGCAGTTCAT GCAGCTGGATTGTGCCGCCAAGAAGTCCCTGATCCTCCAGCTCGCA GCCATCGTGAACGCCAGCAACAGACAGATCGATCTGAGCAAAATTGG CGGCGCCAAGACCGCCGGCCAGTACAAGGTAACCTACAACAAGCTG CTGAACGACCCTAATACCGATTTCTACATCGTGGACCAAAGCGTGAC CGGCATGTTCGAGCGGAGAACCAGAGTCGGACTGTCTAGAAAGCGG ACAGCAGACGGCTCCGAATTTGAAAGCCCTAAGAAAAAGAGAAAGGT GTGA [0094] In some embodiments a DRCY Type II Cas protein comprises an amino acid sequence of SEQ ID NO:37, SEQ ID NO:38, or SEQ ID NO:39. In some embodiments, a DRCY Type II Cas protein has nickase activity, for example resulting from one or more amino acid substitutions relative to the sequence of SEQ ID NO:37, SEQ ID NO:38, or SEQ ID NO:39. In some embodiments, the one or more amino acid substitutions providing nickase activity comprise a D15A substitution, wherein the position of the D15A substitution is defined with respect to the amino acid numbering of SEQ ID NO:38. In some embodiments, the one or more amino acid substitutions providing nickase activity comprise an H893A substitution, wherein the position of the H893A substitution is defined with respect to the amino acid numbering of SEQ ID NO:38. In some embodiments, a DRCY Type II Cas protein is catalytically inactive, for example due to a D15A substitution in combination with a H893A substitution. 6.2.1.8. DRPY Type II Cas Proteins [0095] In one aspect, the disclosure provides DRPY Type II Cas proteins. DRPY Type II Cas proteins can be further classified as Type IIA Cas proteins. The DRPY Type II Cas proteins typically comprise an amino acid sequence that is at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 85%, at least 90%, or at least 95% identical to SEQ ID NO:43. In some embodiments, the DRPY Type II Cas proteins comprise an amino acid sequence that is at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical to SEQ ID NO:43. In some embodiments, a DRPY Type II Cas protein comprises an amino acid sequence that is identical to SEQ ID NO:43. [0096] Exemplary DRPY Type II Cas protein sequences and nucleotide sequences encoding exemplary DRPY Type II Cas proteins are set forth in Table 1H. TABLE 1H DRPY Type II Cas Sequences Name Sequence SEQ ID NO DRPY Type II KKRAIEEYYLGLDIGTNSVGYAVTDPQYKILKYHGEPMWGSHVFEEGSQ 43 Cas amino CAERRGFRTARRRLNRRQQRVRLVQEIFAHEIEKVDSRFFLRLKESALYR acid EDANGNDPFILFNDNGYTDKEYYLKYPTIHHLIMDLIDDENPHDVRLVYLA sequence VAWLVAHRGHFLSDVEKTNVEKVLDFSTSYNALKEMYSGIGQDLPWIDE (without N- EETFKNILLKKCGVKDKEKAFKDKLLKNNSNKKNSDITEESEVLGISAVITL terminal LSGGTVAADKLFVQAEFQDKISISFKKNEDEFEQILTELDEYAEYLLKIRAV methionine) YDWAVLYEASKGQVYISKAKIEVYEQHKKDLAGLKAFIRKYCPDKYNEIF RDAGDNNYVAYSYNFNSLNLTDGEKRPKRKASQEDFCTYIRKIVKDIKCE MEDEEFYDDMWQRLELGTFMPKQVNTDNRVLPYQLYYFELARLLGRVS AYLPFINEKDNDGYITKEKLLSIMEFRIPYYVGPLHRENKDNNSFAWIKRK AQGRIYPWNYEEKIDLDASEQAFINKMTNQCSYLPGEDVLPKYSLLYCKY EVLNEINNIKINGQKITVECKKGIFGLFKKNKKVTVKKIKDFLISNNYMGTS DEITGLDISVKSSLKSYHDFRKMLENKIVNEEDVEEIIKRLTYTEDKKRINR YLEENYPNISEEDRRYISKLKYKDFGRLSEKFLTGIKGVVKETGEILSVIQI MLETNDNLMQIIFSDHYMIKDALEREKKDYYSEHPASVEQILSDMYISNAV KRPIYRTLDIISDVTKACQNAPRKIFVEMARGGGEKGKRTISRRDKIKELY KNMDKQEVREISKEISELLGELDKKTDNELQSEVLFLYFMQLGICMYSGD PIDITKLKSDAYVNVDHIYPQAYVKDDSLNNKVLVKSKLNGEKSDNYPINK DIRDKMSSIWKHYRKEGLISEEKYNRLTRKTPFTDEEKQGFVNRQLVETR QSTKAVAEILKMMFPETEVVYVKAGLASEFRHVFGIIKSRQINDLHHAKDA YLNVVCGNVYHSTFTKNFFLKHQTYSIKTETVYKRRIEADGKLIWNGQES LSFVKKMLGKNNIHYTRYAFCRKGGFFDQMPVTAKEGLVSRKKDLPSEK YGGYNKPTASYFIMAKYTEKVKKEKQDIMIVPVELMVGEKVLNDKNFAVG YIQQQIAQISNRKVEEIVNVSFPLGLRPLKVNTRFSFDGFEACVTGKDSG GKNISMSSLTSLKVSQANEKYIKKVERFAEKKEVNNKITLDEKYDEINENL NFEIYRMLQQKITKSCFGVAFGNLSNVFEKGENKFVALKTDMQCEVLLNII NVFKTGRSGTCNLKSIGGVEKAASYRISAKVSNWKKNYTCVKIIDESASGI FRKESENILEWL DRPY Type II MKKRAIEEYYLGLDIGTNSVGYAVTDPQYKILKYHGEPMWGSHVFEEGS 44 Cas amino QCAERRGFRTARRRLNRRQQRVRLVQEIFAHEIEKVDSRFFLRLKESAL acid YREDANGNDPFILFNDNGYTDKEYYLKYPTIHHLIMDLIDDENPHDVRLVY sequence LAVAWLVAHRGHFLSDVEKTNVEKVLDFSTSYNALKEMYSGIGQDLPWI DEEETFKNILLKKCGVKDKEKAFKDKLLKNNSNKKNSDITEESEVLGISAV ITLLSGGTVAADKLFVQAEFQDKISISFKKNEDEFEQILTELDEYAEYLLKI RAVYDWAVLYEASKGQVYISKAKIEVYEQHKKDLAGLKAFIRKYCPDKYN EIFRDAGDNNYVAYSYNFNSLNLTDGEKRPKRKASQEDFCTYIRKIVKDIK CEMEDEEFYDDMWQRLELGTFMPKQVNTDNRVLPYQLYYFELARLLGR VSAYLPFINEKDNDGYITKEKLLSIMEFRIPYYVGPLHRENKDNNSFAWIK RKAQGRIYPWNYEEKIDLDASEQAFINKMTNQCSYLPGEDVLPKYSLLYC KYEVLNEINNIKINGQKITVECKKGIFGLFKKNKKVTVKKIKDFLISNNYMG TSDEITGLDISVKSSLKSYHDFRKMLENKIVNEEDVEEIIKRLTYTEDKKRI NRYLEENYPNISEEDRRYISKLKYKDFGRLSEKFLTGIKGVVKETGEILSVI QIMLETNDNLMQIIFSDHYMIKDALEREKKDYYSEHPASVEQILSDMYISN AVKRPIYRTLDIISDVTKACQNAPRKIFVEMARGGGEKGKRTISRRDKIKE LYKNMDKQEVREISKEISELLGELDKKTDNELQSEVLFLYFMQLGICMYS GDPIDITKLKSDAYVNVDHIYPQAYVKDDSLNNKVLVKSKLNGEKSDNYPI NKDIRDKMSSIWKHYRKEGLISEEKYNRLTRKTPFTDEEKQGFVNRQLVE TABLE 1H DRPY Type II Cas Sequences Name Sequence SEQ ID NO TRQSTKAVAEILKMMFPETEVVYVKAGLASEFRHVFGIIKSRQINDLHHAK DAYLNVVCGNVYHSTFTKNFFLKHQTYSIKTETVYKRRIEADGKLIWNGQ ESLSFVKKMLGKNNIHYTRYAFCRKGGFFDQMPVTAKEGLVSRKKDLPS EKYGGYNKPTASYFIMAKYTEKVKKEKQDIMIVPVELMVGEKVLNDKNFA VGYIQQQIAQISNRKVEEIVNVSFPLGLRPLKVNTRFSFDGFEACVTGKD SGGKNISMSSLTSLKVSQANEKYIKKVERFAEKKEVNNKITLDEKYDEINE NLNFEIYRMLQQKITKSCFGVAFGNLSNVFEKGENKFVALKTDMQCEVLL NIINVFKTGRSGTCNLKSIGGVEKAASYRISAKVSNWKKNYTCVKIIDESA SGIFRKESENILEWL DRPY Type II MGKPIPNPLLGLDSTKRTADGSEFESPKKKRKVGSKKRAIEEYYLGLDIG 45 Cas TNSVGYAVTDPQYKILKYHGEPMWGSHVFEEGSQCAERRGFRTARRRL mammalian NRRQQRVRLVQEIFAHEIEKVDSRFFLRLKESALYREDANGNDPFILFND expression NGYTDKEYYLKYPTIHHLIMDLIDDENPHDVRLVYLAVAWLVAHRGHFLS construct DVEKTNVEKVLDFSTSYNALKEMYSGIGQDLPWIDEEETFKNILLKKCGV (with N- KDKEKAFKDKLLKNNSNKKNSDITEESEVLGISAVITLLSGGTVAADKLFV terminal QAEFQDKISISFKKNEDEFEQILTELDEYAEYLLKIRAVYDWAVLYEASKG methionine, QVYISKAKIEVYEQHKKDLAGLKAFIRKYCPDKYNEIFRDAGDNNYVAYS stop codon, YNFNSLNLTDGEKRPKRKASQEDFCTYIRKIVKDIKCEMEDEEFYDDMW V5 tag, N- QRLELGTFMPKQVNTDNRVLPYQLYYFELARLLGRVSAYLPFINEKDND terminal NLS GYITKEKLLSIMEFRIPYYVGPLHRENKDNNSFAWIKRKAQGRIYPWNYE and C- EKIDLDASEQAFINKMTNQCSYLPGEDVLPKYSLLYCKYEVLNEINNIKIN terminal NLS) GQKITVECKKGIFGLFKKNKKVTVKKIKDFLISNNYMGTSDEITGLDISVKS (aa) SLKSYHDFRKMLENKIVNEEDVEEIIKRLTYTEDKKRINRYLEENYPNISEE DRRYISKLKYKDFGRLSEKFLTGIKGVVKETGEILSVIQIMLETNDNLMQIIF SDHYMIKDALEREKKDYYSEHPASVEQILSDMYISNAVKRPIYRTLDIISDV TKACQNAPRKIFVEMARGGGEKGKRTISRRDKIKELYKNMDKQEVREISK EISELLGELDKKTDNELQSEVLFLYFMQLGICMYSGDPIDITKLKSDAYVN VDHIYPQAYVKDDSLNNKVLVKSKLNGEKSDNYPINKDIRDKMSSIWKHY RKEGLISEEKYNRLTRKTPFTDEEKQGFVNRQLVETRQSTKAVAEILKMM FPETEVVYVKAGLASEFRHVFGIIKSRQINDLHHAKDAYLNVVCGNVYHS TFTKNFFLKHQTYSIKTETVYKRRIEADGKLIWNGQESLSFVKKMLGKNNI HYTRYAFCRKGGFFDQMPVTAKEGLVSRKKDLPSEKYGGYNKPTASYFI MAKYTEKVKKEKQDIMIVPVELMVGEKVLNDKNFAVGYIQQQIAQISNRK VEEIVNVSFPLGLRPLKVNTRFSFDGFEACVTGKDSGGKNISMSSLTSLK VSQANEKYIKKVERFAEKKEVNNKITLDEKYDEINENLNFEIYRMLQQKIT KSCFGVAFGNLSNVFEKGENKFVALKTDMQCEVLLNIINVFKTGRSGTCN LKSIGGVEKAASYRISAKVSNWKKNYTCVKIIDESASGIFRKESENILEWL SRKRTADGSEFESPKKKRKV DRPY Type II ATGAAAAAAAGAGCAATTGAAGAATATTATTTAGGACTTGATATTGGAA 46 Cas coding CAAATTCTGTGGGATATGCAGTTACTGATCCACAGTATAAAATTTTAAA sequence (nt) ATATCACGGAGAGCCGATGTGGGGATCTCATGTGTTTGAGGAGGGCA (not codon GCCAGTGTGCGGAAAGGCGTGGCTTTAGAACTGCCAGAAGGAGATT optimized) AAACAGACGTCAGCAAAGAGTACGCTTGGTTCAGGAGATATTTGCCC ACGAGATTGAAAAAGTTGATAGCAGATTTTTCCTTCGCCTAAAGGAAA GTGCTCTGTATCGTGAAGACGCAAACGGGAATGATCCATTTATATTGT TTAATGACAACGGATACACTGACAAAGAATATTATTTAAAATATCCAAC GATACATCATCTTATCATGGATCTGATTGATGATGAAAATCCGCACGA TGTCCGCCTGGTTTATCTTGCGGTGGCATGGCTTGTTGCACACAGAG GTCATTTTTTAAGTGATGTTGAAAAAACAAATGTAGAAAAGGTGTTAGA TTTTTCCACTTCATATAATGCACTTAAGGAAATGTATTCTGGTATTGGA CAGGATTTACCTTGGATTGACGAAGAGGAAACATTTAAAAACATTTTA TTGAAAAAATGTGGGGTGAAAGATAAAGAAAAAGCATTTAAGGATAAA TTATTGAAAAATAACTCTAATAAAAAGAATTCAGATATAACGGAGGAGA GCGAAGTACTGGGAATAAGTGCGGTCATAACCTTACTTTCTGGAGGA ACGGTTGCAGCAGATAAGCTTTTTGTGCAAGCCGAGTTTCAGGACAA AATTAGCATTAGTTTTAAAAAGAATGAAGATGAGTTTGAACAGATTTTA ACAGAATTAGATGAATATGCGGAATATTTATTAAAGATCAGAGCTGTA TABLE 1H DRPY Type II Cas Sequences Name Sequence SEQ ID NO TATGACTGGGCTGTATTATATGAGGCATCAAAGGGACAAGTATACATT TCAAAGGCCAAAATAGAGGTGTATGAACAGCACAAAAAAGATTTGGC AGGGCTCAAAGCCTTTATAAGAAAATATTGTCCGGATAAATACAATGA AATTTTCCGTGATGCAGGCGATAACAATTATGTGGCGTATTCGTATAA TTTTAATTCTTTGAATCTTACTGATGGGGAAAAGCGACCGAAGAGAAA AGCTTCACAGGAAGATTTCTGCACTTATATACGGAAAATTGTTAAAGA TATTAAGTGCGAAATGGAGGATGAAGAATTTTATGACGATATGTGGCA GCGGTTAGAACTTGGAACATTTATGCCGAAACAGGTTAATACGGATAA TCGTGTCCTTCCGTATCAGTTATATTATTTTGAACTTGCCCGCCTTTTA GGAAGGGTATCGGCATATCTTCCATTTATCAATGAAAAGGATAATGAT GGATACATTACGAAAGAAAAATTGCTTTCTATTATGGAATTTCGGATTC CTTATTATGTAGGTCCATTACATAGAGAGAATAAAGATAATAATTCATT TGCCTGGATTAAGAGAAAGGCTCAAGGACGTATTTATCCATGGAACTA TGAAGAAAAAATTGATCTTGATGCAAGTGAGCAGGCATTTATCAACAA AATGACAAATCAGTGTAGTTACCTGCCGGGAGAGGATGTACTGCCGA AATATTCCCTGTTGTACTGTAAATATGAAGTGTTAAATGAAATTAATAA TATTAAAATAAATGGTCAAAAAATTACAGTGGAATGTAAAAAAGGAATC TTTGGTCTTTTTAAAAAGAACAAAAAAGTTACAGTAAAGAAGATTAAAG ATTTTTTGATCAGTAATAACTATATGGGGACTTCAGATGAGATAACGG GACTTGACATTTCTGTAAAATCATCTTTGAAATCATACCATGATTTTCG AAAAATGTTGGAAAATAAAATTGTCAATGAGGAGGACGTGGAAGAAAT TATCAAGCGTCTTACCTATACGGAAGACAAAAAAAGAATTAATAGGTA TTTAGAAGAAAATTATCCTAATATTTCGGAGGAAGACAGGCGTTATAT TTCAAAATTGAAATATAAGGATTTTGGAAGATTATCGGAAAAATTTTTG ACAGGAATTAAGGGGGTTGTAAAAGAAACGGGAGAAATCTTAAGTGT TATACAGATTATGTTGGAAACAAATGACAACCTGATGCAGATTATTTTT TCTGATCATTATATGATTAAAGATGCACTTGAAAGAGAAAAGAAAGATT ATTATAGTGAACATCCGGCAAGTGTGGAACAGATATTATCGGATATGT ACATTTCGAATGCGGTGAAACGTCCGATTTATAGAACATTGGATATTA TAAGTGATGTTACAAAAGCATGTCAGAATGCACCACGCAAGATATTTG TCGAGATGGCACGCGGAGGTGGTGAAAAGGGGAAAAGAACCATTTC GCGTCGCGATAAAATTAAGGAATTGTACAAGAATATGGACAAACAGG AAGTAAGAGAAATATCAAAAGAGATCTCCGAGCTTCTTGGTGAGTTGG ATAAAAAGACAGACAATGAGCTGCAGAGTGAGGTGTTGTTCTTATATT TTATGCAGTTAGGAATATGTATGTATTCGGGAGATCCGATTGACATTA CCAAACTGAAATCGGACGCATATGTGAATGTAGATCATATTTATCCGC AGGCGTATGTGAAAGATGACAGCTTGAATAATAAAGTATTGGTAAAAT CAAAATTAAATGGAGAAAAGTCAGATAATTATCCGATAAACAAGGATA TCCGAGATAAAATGTCATCAATTTGGAAGCACTATAGAAAAGAAGGGT TAATTTCTGAGGAAAAATATAACCGCTTAACAAGAAAAACGCCATTTA CGGATGAGGAAAAACAAGGATTTGTGAATCGTCAGTTAGTAGAAACA AGACAATCCACAAAGGCTGTGGCTGAAATTTTAAAAATGATGTTTCCG GAAACGGAAGTTGTATATGTAAAAGCAGGGTTAGCGTCAGAGTTTCG ACATGTATTTGGAATTATAAAATCTCGTCAGATTAATGACCTGCATCAT GCGAAGGACGCTTATTTAAACGTAGTATGTGGAAATGTTTATCATTCT ACATTTACAAAAAACTTTTTCTTGAAGCATCAGACGTATTCGATAAAAA CAGAAACGGTATATAAACGAAGAATTGAGGCAGATGGAAAGCTGATA TGGAATGGACAGGAGTCGTTATCTTTTGTTAAAAAGATGCTTGGCAAA AATAATATACATTATACCAGATATGCATTTTGCAGGAAGGGCGGATTT TTTGATCAGATGCCGGTTACAGCAAAAGAGGGGCTGGTTTCGAGAAA GAAAGATTTACCATCAGAAAAATATGGTGGATATAATAAACCAACAGC GTCCTATTTCATAATGGCAAAATATACGGAAAAAGTGAAGAAGGAAAA ACAGGATATTATGATTGTACCTGTAGAGCTAATGGTGGGTGAAAAAGT ACTGAATGATAAAAACTTTGCGGTTGGTTATATTCAGCAACAGATTGC ACAGATTTCAAATAGGAAAGTAGAAGAAATTGTTAATGTTTCGTTTCC GTTGGGATTGAGACCACTTAAGGTAAATACGAGATTTTCTTTTGATGG GTTTGAAGCATGTGTAACGGGAAAAGATAGTGGAGGAAAAAATATAA TABLE 1H DRPY Type II Cas Sequences Name Sequence SEQ ID NO GTATGAGTTCACTTACTTCTTTAAAAGTGAGTCAAGCAAATGAAAAATA TATAAAGAAAGTAGAAAGATTTGCAGAAAAGAAGGAGGTAAACAACAA AATAACTTTAGATGAAAAATATGATGAAATAAATGAAAACTTAAATTTT GAAATATATAGAATGTTGCAACAAAAAATAACAAAAAGTTGCTTTGGG GTTGCATTTGGTAATTTATCTAATGTGTTTGAAAAAGGAGAAAACAAAT TTGTTGCTTTAAAGACAGATATGCAATGTGAGGTGTTACTAAACATAAT TAATGTTTTTAAAACAGGTAGAAGTGGTACATGTAATTTGAAAAGCATT GGTGGAGTGGAAAAGGCTGCATCATATAGAATTAGTGCAAAAGTTTC AAATTGGAAAAAGAACTATACATGTGTAAAGATAATCGACGAATCAGC ATCAGGAATTTTCCGTAAAGAATCGGAAAATATATTGGAGTGGTTATG A DRPY Type II AAGAAAAGAGCCATCGAGGAATACTACCTGGGCCTGGATATTGGCAC 47 Cas coding CAACAGCGTGGGGTACGCCGTGACCGATCCTCAGTACAAAATCCTGA sequence (nt) AGTACCACGGCGAGCCTATGTGGGGTAGCCACGTGTTTGAGGAAGG (human CAGCCAGTGCGCTGAGAGAAGGGGCTTCAGAACCGCCAGACGGAGA codon- TTGAATAGGAGGCAGCAAAGAGTCAGACTGGTCCAGGAGATCTTTGC optimized; CCACGAGATCGAGAAGGTGGACAGCAGATTCTTCCTGAGACTGAAGG lacking N- AGAGCGCCCTGTATAGAGAGGACGCCAACGGCAACGACCCTTTCATC terminal CTGTTTAACGACAACGGCTACACAGACAAGGAGTACTACCTGAAGTA methionine; CCCCACCATCCACCACCTGATCATGGACCTGATCGACGATGAAAACC no stop CCCACGACGTGAGACTGGTGTACCTCGCTGTGGCCTGGCTGGTGGC codon) TCACAGAGGCCACTTCCTGTCTGATGTGGAGAAAACCAATGTGGAAA AAGTGCTGGACTTCTCCACCTCGTACAATGCTCTGAAGGAGATGTAC TCCGGAATCGGCCAAGACCTGCCTTGGATCGACGAAGAGGAAACCTT CAAGAACATCCTCCTGAAGAAGTGCGGCGTCAAGGACAAGGAGAAG GCTTTTAAGGACAAGCTGCTGAAGAACAACAGTAACAAGAAAAACAG CGACATTACAGAAGAATCTGAAGTCCTGGGCATCTCAGCCGTGATCA CCCTGCTGTCTGGTGGCACCGTGGCTGCCGACAAGCTGTTCGTGCA GGCCGAGTTCCAGGACAAGATCAGCATCTCTTTCAAGAAGAATGAGG ATGAGTTCGAACAGATCCTGACAGAGCTGGACGAGTACGCGGAGTAC CTGCTGAAGATCAGGGCCGTTTATGACTGGGCCGTGCTGTACGAAGC CAGCAAAGGCCAGGTGTACATCAGCAAGGCTAAGATCGAGGTTTACG AGCAACACAAGAAGGACCTGGCCGGACTGAAGGCCTTTATCAGAAAG TATTGTCCTGATAAGTACAACGAAATCTTCCGCGACGCAGGCGACAA CAATTACGTGGCTTATTCCTACAACTTCAACAGCCTGAACCTGACCGA CGGCGAGAAGCGGCCCAAAAGAAAGGCCAGCCAGGAGGACTTCTGC ACCTACATCCGGAAGATTGTGAAGGATATAAAGTGTGAAATGGAAGAT GAGGAGTTCTATGACGACATGTGGCAGAGACTGGAACTCGGAACCTT CATGCCTAAGCAGGTGAACACCGATAACCGGGTGCTGCCGTACCAG CTGTATTACTTCGAGCTGGCACGGCTGCTGGGCAGAGTGAGCGCCT ATCTGCCTTTTATCAATGAAAAAGACAACGACGGCTACATCACCAAAG AGAAGCTGCTGAGCATCATGGAGTTCAGAATCCCATACTACGTTGGA CCTCTGCATAGAGAAAACAAGGATAACAACTCCTTTGCCTGGATCAAG CGGAAGGCCCAGGGAAGAATCTACCCTTGGAACTACGAGGAGAAGA TCGACCTGGACGCTTCCGAGCAGGCCTTCATCAACAAGATGACCAAC CAATGTAGCTACCTACCTGGCGAGGACGTGCTGCCTAAGTATTCTCT GCTGTACTGCAAGTACGAGGTCCTGAACGAGATCAACAATATTAAGAT CAACGGACAGAAAATAACCGTGGAGTGCAAGAAGGGCATCTTCGGC CTGTTCAAGAAGAACAAAAAGGTGACCGTGAAGAAGATCAAGGACTT CCTGATCTCTAACAACTACATGGGCACAAGCGACGAAATCACAGGCC TGGACATCAGCGTGAAGTCAAGCCTGAAATCTTACCATGATTTCCGG AAGATGCTGGAGAACAAGATCGTGAACGAGGAGGACGTGGAAGAAA TCATCAAGAGGCTGACATACACCGAGGACAAGAAGCGGATCAACAGA TACCTGGAGGAAAATTACCCTAACATTTCTGAGGAAGATAGACGGTAC ATCTCCAAGCTGAAATACAAGGACTTCGGCAGACTGAGCGAAAAGTT CCTGACCGGCATCAAGGGCGTGGTGAAGGAAACCGGCGAAATTCTG AGCGTCATTCAGATCATGCTGGAAACAAACGACAATCTGATGCAGAT TABLE 1H DRPY Type II Cas Sequences Name Sequence SEQ ID NO CATATTCAGCGATCACTACATGATCAAGGACGCCCTGGAAAGAGAAA AGAAGGACTACTACAGCGAGCACCCCGCGAGCGTGGAACAAATCCT GAGCGACATGTACATCTCCAACGCCGTGAAGCGGCCTATCTACCGCA CCCTGGATATCATCAGCGATGTGACCAAAGCCTGCCAGAACGCCCCT AGAAAGATCTTCGTCGAAATGGCCAGAGGCGGCGGCGAGAAGGGCA AGAGAACAATCAGCAGAAGAGACAAGATCAAAGAGCTGTATAAGAAT ATGGACAAACAGGAGGTGCGGGAAATCAGCAAGGAGATCAGCGAGC TGCTGGGCGAGCTGGACAAGAAAACCGATAACGAGCTGCAGAGCGA AGTACTGTTCTTGTACTTCATGCAGCTGGGAATCTGCATGTACAGCG GAGATCCTATCGACATCACTAAGCTGAAGAGCGACGCCTACGTGAAT GTCGACCACATTTACCCCCAGGCTTACGTGAAGGACGACAGCCTGAA CAACAAAGTGCTGGTGAAGTCTAAACTGAATGGGGAAAAAAGCGACA ATTACCCTATTAACAAGGACATCCGGGACAAGATGAGCAGCATCTGG AAGCACTACAGAAAGGAAGGCTTGATTTCAGAAGAAAAATATAACAGA CTGACCAGAAAGACACCTTTCACAGATGAAGAAAAGCAGGGCTTCGT GAATAGACAGCTGGTCGAAACCAGACAGAGTACCAAGGCGGTTGCT GAAATCCTGAAGATGATGTTCCCCGAAACCGAGGTTGTGTACGTGAA GGCCGGACTGGCCAGCGAGTTCAGACACGTGTTCGGTATCATCAAG AGCAGACAGATCAATGATCTGCATCACGCCAAAGACGCCTACCTGAA TGTGGTGTGCGGCAACGTGTACCACAGCACCTTCACCAAGAACTTTT TCCTGAAGCACCAGACTTATTCAATCAAGACCGAGACAGTGTACAAG CGCAGAATCGAGGCCGATGGCAAGCTGATCTGGAATGGCCAGGAGA GCCTGTCTTTCGTGAAGAAAATGCTGGGAAAGAACAACATCCACTAC ACCAGGTACGCGTTCTGCAGAAAGGGCGGCTTCTTCGACCAGATGC CTGTGACCGCCAAGGAGGGACTGGTGTCCCGGAAAAAGGACCTGCC CAGCGAGAAGTACGGCGGCTATAACAAGCCAACGGCCAGCTATTTTA TCATGGCCAAATACACCGAAAAAGTGAAAAAGGAAAAGCAGGATATC ATGATCGTGCCCGTGGAGCTGATGGTCGGCGAGAAAGTGCTGAACG ACAAAAACTTCGCCGTGGGCTACATCCAACAGCAGATCGCCCAGATC AGCAACCGGAAGGTGGAGGAGATTGTGAACGTGTCTTTCCCTCTGGG CCTGCGGCCTCTGAAGGTGAACACCCGGTTCAGCTTTGATGGCTTTG AGGCCTGTGTGACAGGCAAGGATTCTGGCGGCAAGAACATCTCCATG AGCTCTTTGACAAGCCTGAAAGTTAGCCAAGCTAATGAAAAGTACATC AAGAAGGTGGAGAGATTCGCCGAGAAGAAAGAGGTGAATAACAAGAT CACCCTGGACGAGAAATACGACGAGATCAATGAGAACCTGAACTTCG AGATCTACAGAATGCTGCAGCAGAAGATAACAAAGTCCTGTTTTGGC GTGGCCTTTGGCAACCTGAGCAACGTGTTCGAGAAGGGCGAGAACA AGTTCGTGGCCCTGAAAACCGATATGCAGTGCGAGGTGCTGCTCAAC ATCATCAACGTGTTCAAGACTGGAAGAAGCGGCACATGCAACCTGAA GAGCATCGGAGGAGTGGAAAAGGCCGCCAGCTACCGGATCTCTGCC AAAGTGTCCAACTGGAAGAAGAATTACACCTGCGTGAAAATTATCGAC GAGAGCGCCAGCGGCATCTTTAGAAAGGAGTCTGAGAACATCCTGGA ATGGCTG DRPY Type II ATGGGCAAACCTATCCCCAATCCCCTGCTGGGCCTGGACAGCACCAA 48 Cas GCGGACTGCCGACGGCAGTGAGTTCGAGTCTCCCAAGAAAAAGAGG mammalian AAAGTGGGATCCAAGAAAAGAGCCATCGAGGAATACTACCTGGGCCT expression GGATATTGGCACCAACAGCGTGGGGTACGCCGTGACCGATCCTCAG construct TACAAAATCCTGAAGTACCACGGCGAGCCTATGTGGGGTAGCCACGT (with N- GTTTGAGGAAGGCAGCCAGTGCGCTGAGAGAAGGGGCTTCAGAACC terminal GCCAGACGGAGATTGAATAGGAGGCAGCAAAGAGTCAGACTGGTCC methionine, AGGAGATCTTTGCCCACGAGATCGAGAAGGTGGACAGCAGATTCTTC stop codon, CTGAGACTGAAGGAGAGCGCCCTGTATAGAGAGGACGCCAACGGCA V5 tag, N- ACGACCCTTTCATCCTGTTTAACGACAACGGCTACACAGACAAGGAG terminal NLS TACTACCTGAAGTACCCCACCATCCACCACCTGATCATGGACCTGAT and C- CGACGATGAAAACCCCCACGACGTGAGACTGGTGTACCTCGCTGTG terminal NLS) GCCTGGCTGGTGGCTCACAGAGGCCACTTCCTGTCTGATGTGGAGA (nt) AAACCAATGTGGAAAAAGTGCTGGACTTCTCCACCTCGTACAATGCTC TABLE 1H DRPY Type II Cas Sequences Name Sequence SEQ ID NO TGAAGGAGATGTACTCCGGAATCGGCCAAGACCTGCCTTGGATCGAC GAAGAGGAAACCTTCAAGAACATCCTCCTGAAGAAGTGCGGCGTCAA GGACAAGGAGAAGGCTTTTAAGGACAAGCTGCTGAAGAACAACAGTA ACAAGAAAAACAGCGACATTACAGAAGAATCTGAAGTCCTGGGCATC TCAGCCGTGATCACCCTGCTGTCTGGTGGCACCGTGGCTGCCGACA AGCTGTTCGTGCAGGCCGAGTTCCAGGACAAGATCAGCATCTCTTTC AAGAAGAATGAGGATGAGTTCGAACAGATCCTGACAGAGCTGGACGA GTACGCGGAGTACCTGCTGAAGATCAGGGCCGTTTATGACTGGGCC GTGCTGTACGAAGCCAGCAAAGGCCAGGTGTACATCAGCAAGGCTAA GATCGAGGTTTACGAGCAACACAAGAAGGACCTGGCCGGACTGAAG GCCTTTATCAGAAAGTATTGTCCTGATAAGTACAACGAAATCTTCCGC GACGCAGGCGACAACAATTACGTGGCTTATTCCTACAACTTCAACAG CCTGAACCTGACCGACGGCGAGAAGCGGCCCAAAAGAAAGGCCAGC CAGGAGGACTTCTGCACCTACATCCGGAAGATTGTGAAGGATATAAA GTGTGAAATGGAAGATGAGGAGTTCTATGACGACATGTGGCAGAGAC TGGAACTCGGAACCTTCATGCCTAAGCAGGTGAACACCGATAACCGG GTGCTGCCGTACCAGCTGTATTACTTCGAGCTGGCACGGCTGCTGG GCAGAGTGAGCGCCTATCTGCCTTTTATCAATGAAAAAGACAACGAC GGCTACATCACCAAAGAGAAGCTGCTGAGCATCATGGAGTTCAGAAT CCCATACTACGTTGGACCTCTGCATAGAGAAAACAAGGATAACAACTC CTTTGCCTGGATCAAGCGGAAGGCCCAGGGAAGAATCTACCCTTGGA ACTACGAGGAGAAGATCGACCTGGACGCTTCCGAGCAGGCCTTCATC AACAAGATGACCAACCAATGTAGCTACCTACCTGGCGAGGACGTGCT GCCTAAGTATTCTCTGCTGTACTGCAAGTACGAGGTCCTGAACGAGA TCAACAATATTAAGATCAACGGACAGAAAATAACCGTGGAGTGCAAGA AGGGCATCTTCGGCCTGTTCAAGAAGAACAAAAAGGTGACCGTGAAG AAGATCAAGGACTTCCTGATCTCTAACAACTACATGGGCACAAGCGA CGAAATCACAGGCCTGGACATCAGCGTGAAGTCAAGCCTGAAATCTT ACCATGATTTCCGGAAGATGCTGGAGAACAAGATCGTGAACGAGGAG GACGTGGAAGAAATCATCAAGAGGCTGACATACACCGAGGACAAGAA GCGGATCAACAGATACCTGGAGGAAAATTACCCTAACATTTCTGAGG AAGATAGACGGTACATCTCCAAGCTGAAATACAAGGACTTCGGCAGA CTGAGCGAAAAGTTCCTGACCGGCATCAAGGGCGTGGTGAAGGAAA CCGGCGAAATTCTGAGCGTCATTCAGATCATGCTGGAAACAAACGAC AATCTGATGCAGATCATATTCAGCGATCACTACATGATCAAGGACGCC CTGGAAAGAGAAAAGAAGGACTACTACAGCGAGCACCCCGCGAGCG TGGAACAAATCCTGAGCGACATGTACATCTCCAACGCCGTGAAGCGG CCTATCTACCGCACCCTGGATATCATCAGCGATGTGACCAAAGCCTG CCAGAACGCCCCTAGAAAGATCTTCGTCGAAATGGCCAGAGGCGGC GGCGAGAAGGGCAAGAGAACAATCAGCAGAAGAGACAAGATCAAAG AGCTGTATAAGAATATGGACAAACAGGAGGTGCGGGAAATCAGCAAG GAGATCAGCGAGCTGCTGGGCGAGCTGGACAAGAAAACCGATAACG AGCTGCAGAGCGAAGTACTGTTCTTGTACTTCATGCAGCTGGGAATC TGCATGTACAGCGGAGATCCTATCGACATCACTAAGCTGAAGAGCGA CGCCTACGTGAATGTCGACCACATTTACCCCCAGGCTTACGTGAAGG ACGACAGCCTGAACAACAAAGTGCTGGTGAAGTCTAAACTGAATGGG GAAAAAAGCGACAATTACCCTATTAACAAGGACATCCGGGACAAGAT GAGCAGCATCTGGAAGCACTACAGAAAGGAAGGCTTGATTTCAGAAG AAAAATATAACAGACTGACCAGAAAGACACCTTTCACAGATGAAGAAA AGCAGGGCTTCGTGAATAGACAGCTGGTCGAAACCAGACAGAGTACC AAGGCGGTTGCTGAAATCCTGAAGATGATGTTCCCCGAAACCGAGGT TGTGTACGTGAAGGCCGGACTGGCCAGCGAGTTCAGACACGTGTTC GGTATCATCAAGAGCAGACAGATCAATGATCTGCATCACGCCAAAGA CGCCTACCTGAATGTGGTGTGCGGCAACGTGTACCACAGCACCTTCA CCAAGAACTTTTTCCTGAAGCACCAGACTTATTCAATCAAGACCGAGA CAGTGTACAAGCGCAGAATCGAGGCCGATGGCAAGCTGATCTGGAAT GGCCAGGAGAGCCTGTCTTTCGTGAAGAAAATGCTGGGAAAGAACAA TABLE 1H DRPY Type II Cas Sequences Name Sequence SEQ ID NO CATCCACTACACCAGGTACGCGTTCTGCAGAAAGGGCGGCTTCTTCG ACCAGATGCCTGTGACCGCCAAGGAGGGACTGGTGTCCCGGAAAAA GGACCTGCCCAGCGAGAAGTACGGCGGCTATAACAAGCCAACGGCC AGCTATTTTATCATGGCCAAATACACCGAAAAAGTGAAAAAGGAAAAG CAGGATATCATGATCGTGCCCGTGGAGCTGATGGTCGGCGAGAAAG TGCTGAACGACAAAAACTTCGCCGTGGGCTACATCCAACAGCAGATC GCCCAGATCAGCAACCGGAAGGTGGAGGAGATTGTGAACGTGTCTTT CCCTCTGGGCCTGCGGCCTCTGAAGGTGAACACCCGGTTCAGCTTT GATGGCTTTGAGGCCTGTGTGACAGGCAAGGATTCTGGCGGCAAGA ACATCTCCATGAGCTCTTTGACAAGCCTGAAAGTTAGCCAAGCTAATG AAAAGTACATCAAGAAGGTGGAGAGATTCGCCGAGAAGAAAGAGGTG AATAACAAGATCACCCTGGACGAGAAATACGACGAGATCAATGAGAA CCTGAACTTCGAGATCTACAGAATGCTGCAGCAGAAGATAACAAAGT CCTGTTTTGGCGTGGCCTTTGGCAACCTGAGCAACGTGTTCGAGAAG GGCGAGAACAAGTTCGTGGCCCTGAAAACCGATATGCAGTGCGAGG TGCTGCTCAACATCATCAACGTGTTCAAGACTGGAAGAAGCGGCACA TGCAACCTGAAGAGCATCGGAGGAGTGGAAAAGGCCGCCAGCTACC GGATCTCTGCCAAAGTGTCCAACTGGAAGAAGAATTACACCTGCGTG AAAATTATCGACGAGAGCGCCAGCGGCATCTTTAGAAAGGAGTCTGA GAACATCCTGGAATGGCTGTCTAGAAAGCGGACAGCAGACGGCTCC GAATTTGAAAGCCCTAAGAAAAAGAGAAAGGTGTGA [0097] In some embodiments a DRPY Type II Cas protein comprises an amino acid sequence of SEQ ID NO:43, SEQ ID NO:44, or SEQ ID NO:45. In some embodiments, a DRPY Type II Cas protein has nickase activity, for example resulting from one or more amino acid substitutions relative to the sequence of SEQ ID NO:43, SEQ ID NO:44, or SEQ ID NO:45. In some embodiments, the one or more amino acid substitutions providing nickase activity comprise a D14A substitution, wherein the position of the D14A substitution is defined with respect to the amino acid numbering of SEQ ID NO:44. In some embodiments, the one or more amino acid substitutions providing nickase activity comprise an H878A substitution, wherein the position of the H878A substitution is defined with respect to the amino acid numbering of SEQ ID NO:44. In some embodiments, a DRPY Type II Cas protein is catalytically inactive, for example due to a D14A substitution in combination with an H878A substitution. 6.2.1.9. EAJR Type II Cas Proteins [0098] In one aspect, the disclosure provides EAJR Type II Cas proteins. EAJR Type II Cas proteins can be further classified as Type IIA Cas proteins. The EAJR Type II Cas proteins typically comprise an amino acid sequence that is at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 85%, at least 90%, or at least 95% identical to SEQ ID NO:49. In some embodiments, the EAJR Type II Cas proteins comprise an amino acid sequence that is at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical to SEQ ID NO:49. In some embodiments, an EAJR Type II Cas protein comprises an amino acid sequence that is identical to SEQ ID NO:49. [0099] Exemplary EAJR Type II Cas protein sequences and nucleotide sequences encoding exemplary EAJR Type II Cas proteins are set forth in Table 1J. TABLE 1J EAJR Type II Cas Sequences Name Sequence SEQ ID NO EAJR Type II EEKKTIRSHSPYYIGLDIGTNSVGWAVTDPEYNVLRFKGNSMWGARLFE 49 Cas amino EAKPAADRRAARIARRRLERRKQRLLYLEEFFSAEIGKSDPLFFRRLHES acid ALFPDDKTTGTGKYTLFNDPDFTDRDYLRRYPTVYHLRSELVHSSEPHD sequence PRLVFLALHHIIKHRGHFLYETSENGDECPTVDAAMQELTEYLSAEMNVE (without N- LGLEPEYFTILTDRNMRISQKKAALRAALGRSVQTEESGADIQALSDLLA terminal GAKVKLAALYDDESLKNADPASIALSDDLDAVFDALSSALGERVELLLAA methionine) KKVFDAARLSEMRGGEAYLCDAKVRQYEKNHADLRALKDYVREAAPDK YRNIFYKNAKFPNNYAAYSRYRSESGDKTCKQEDFCAFLKSVLPKPAAE HSFAPQYVQIEEKTFLPRLTGSENGIIPYQLHRQELRKILDNASAYLPFLLE EDEDGISVREKIEAVFSFRVPYYVGPLKGRWAERTEEAKRTGEKAYPWN FEKVVDLDASAAAFLKDLIGKCTYTGAPVLPKDSLLYSEFMLLNELNPLRL NGNELPVSVKQQLIEDLFKKENRRVTKKRILDYLVMHGYAAKTDEISGIDD PVKTRLRSYHDFAPLIPVLGEEAVEKIIEGTLVYSDDKRMLRRWLKKNFP ALNEDDVRKICRLKYADWGRLSKEFLTELYSGADEQGEAQNILYFLRNTN LNLMQLLNDPEHANLPAAAEERRRARFGRDSLRSQLDEMYISPAVRRGI WQTLKISEEIAHIRGGAPAKIFVEVARGEDKEKKGTRTKSRKEQLLELYKA CERDSGELYEKLAATEESALRRDKLFLYYTQFGKCMYSGEPIDLETALRD DTTYDIDHIFPRSRIKDDSIENRVLVKSVLNREKTNTYPISQDIRSRMLPFW SMLHDKKMIGDKKYDRLKRNTELTEDELASFVQRQLTETQQSTKALAQL LKERYPESEIVYSKAGNISEFRQKFEMLKCREVNDLHHAKDAYLNIVVGN VYKTEFTDRFFLNIRKENYSLKKVFDGDTPGAWKADGTSIATVKKYMAKN NPIITRMPREEKGQLFNLNIMPAGLGQLPIKAGKPIEKYGGYNKLAGSYY CVVEHTEKKKRVRSIETVLLYARGLYESDPVKYCREILGLVDPVVVAPKIR INSLLQLDGFRLYPSGRTGNRIIYKHAYQLAIDAERERYIRNLTKYIERCAKI EYPATAFDGISAEENVALYNWFVERLESSAYGKLMQALLANLKASDETF KNMDILSQAKTLMEVLKAFKCDAQSSNLKDLCGKGSVGTILKNASLQGV QSAYLIHQSPTGLYEYRTDLLH EAJR Type II MEEKKTIRSHSPYYIGLDIGTNSVGWAVTDPEYNVLRFKGNSMWGARLF 50 Cas amino EEAKPAADRRAARIARRRLERRKQRLLYLEEFFSAEIGKSDPLFFRRLHE acid SALFPDDKTTGTGKYTLFNDPDFTDRDYLRRYPTVYHLRSELVHSSEPH sequence DPRLVFLALHHIIKHRGHFLYETSENGDECPTVDAAMQELTEYLSAEMNV ELGLEPEYFTILTDRNMRISQKKAALRAALGRSVQTEESGADIQALSDLLA GAKVKLAALYDDESLKNADPASIALSDDLDAVFDALSSALGERVELLLAA KKVFDAARLSEMRGGEAYLCDAKVRQYEKNHADLRALKDYVREAAPDK YRNIFYKNAKFPNNYAAYSRYRSESGDKTCKQEDFCAFLKSVLPKPAAE HSFAPQYVQIEEKTFLPRLTGSENGIIPYQLHRQELRKILDNASAYLPFLLE EDEDGISVREKIEAVFSFRVPYYVGPLKGRWAERTEEAKRTGEKAYPWN FEKVVDLDASAAAFLKDLIGKCTYTGAPVLPKDSLLYSEFMLLNELNPLRL NGNELPVSVKQQLIEDLFKKENRRVTKKRILDYLVMHGYAAKTDEISGIDD PVKTRLRSYHDFAPLIPVLGEEAVEKIIEGTLVYSDDKRMLRRWLKKNFP ALNEDDVRKICRLKYADWGRLSKEFLTELYSGADEQGEAQNILYFLRNTN LNLMQLLNDPEHANLPAAAEERRRARFGRDSLRSQLDEMYISPAVRRGI WQTLKISEEIAHIRGGAPAKIFVEVARGEDKEKKGTRTKSRKEQLLELYKA CERDSGELYEKLAATEESALRRDKLFLYYTQFGKCMYSGEPIDLETALRD DTTYDIDHIFPRSRIKDDSIENRVLVKSVLNREKTNTYPISQDIRSRMLPFW SMLHDKKMIGDKKYDRLKRNTELTEDELASFVQRQLTETQQSTKALAQL LKERYPESEIVYSKAGNISEFRQKFEMLKCREVNDLHHAKDAYLNIVVGN VYKTEFTDRFFLNIRKENYSLKKVFDGDTPGAWKADGTSIATVKKYMAKN NPIITRMPREEKGQLFNLNIMPAGLGQLPIKAGKPIEKYGGYNKLAGSYY CVVEHTEKKKRVRSIETVLLYARGLYESDPVKYCREILGLVDPVVVAPKIR INSLLQLDGFRLYPSGRTGNRIIYKHAYQLAIDAERERYIRNLTKYIERCAKI EYPATAFDGISAEENVALYNWFVERLESSAYGKLMQALLANLKASDETF KNMDILSQAKTLMEVLKAFKCDAQSSNLKDLCGKGSVGTILKNASLQGV QSAYLIHQSPTGLYEYRTDLLH EAJR Type II MGKPIPNPLLGLDSTKRTADGSEFESPKKKRKVGSEEKKTIRSHSPYYIG 51 Cas LDIGTNSVGWAVTDPEYNVLRFKGNSMWGARLFEEAKPAADRRAARIA mammalian RRRLERRKQRLLYLEEFFSAEIGKSDPLFFRRLHESALFPDDKTTGTGKY TABLE 1J EAJR Type II Cas Sequences Name Sequence SEQ ID NO expression TLFNDPDFTDRDYLRRYPTVYHLRSELVHSSEPHDPRLVFLALHHIIKHR construct GHFLYETSENGDECPTVDAAMQELTEYLSAEMNVELGLEPEYFTILTDR (with N- NMRISQKKAALRAALGRSVQTEESGADIQALSDLLAGAKVKLAALYDDES terminal LKNADPASIALSDDLDAVFDALSSALGERVELLLAAKKVFDAARLSEMRG methionine, GEAYLCDAKVRQYEKNHADLRALKDYVREAAPDKYRNIFYKNAKFPNNY stop codon, AAYSRYRSESGDKTCKQEDFCAFLKSVLPKPAAEHSFAPQYVQIEEKTF V5 tag, N- LPRLTGSENGIIPYQLHRQELRKILDNASAYLPFLLEEDEDGISVREKIEAV terminal NLS FSFRVPYYVGPLKGRWAERTEEAKRTGEKAYPWNFEKVVDLDASAAAF and C- LKDLIGKCTYTGAPVLPKDSLLYSEFMLLNELNPLRLNGNELPVSVKQQLI terminal NLS) EDLFKKENRRVTKKRILDYLVMHGYAAKTDEISGIDDPVKTRLRSYHDFA (aa) PLIPVLGEEAVEKIIEGTLVYSDDKRMLRRWLKKNFPALNEDDVRKICRLK YADWGRLSKEFLTELYSGADEQGEAQNILYFLRNTNLNLMQLLNDPEHA NLPAAAEERRRARFGRDSLRSQLDEMYISPAVRRGIWQTLKISEEIAHIR GGAPAKIFVEVARGEDKEKKGTRTKSRKEQLLELYKACERDSGELYEKL AATEESALRRDKLFLYYTQFGKCMYSGEPIDLETALRDDTTYDIDHIFPRS RIKDDSIENRVLVKSVLNREKTNTYPISQDIRSRMLPFWSMLHDKKMIGD KKYDRLKRNTELTEDELASFVQRQLTETQQSTKALAQLLKERYPESEIVY SKAGNISEFRQKFEMLKCREVNDLHHAKDAYLNIVVGNVYKTEFTDRFFL NIRKENYSLKKVFDGDTPGAWKADGTSIATVKKYMAKNNPIITRMPREEK GQLFNLNIMPAGLGQLPIKAGKPIEKYGGYNKLAGSYYCVVEHTEKKKRV RSIETVLLYARGLYESDPVKYCREILGLVDPVVVAPKIRINSLLQLDGFRLY PSGRTGNRIIYKHAYQLAIDAERERYIRNLTKYIERCAKIEYPATAFDGISA EENVALYNWFVERLESSAYGKLMQALLANLKASDETFKNMDILSQAKTL MEVLKAFKCDAQSSNLKDLCGKGSVGTILKNASLQGVQSAYLIHQSPTG LYEYRTDLLHSRKRTADGSEFESPKKKRKV EAJR Type II ATGGAAGAGAAAAAAACGATCCGATCGCATTCTCCGTATTATATCGGC 52 Cas coding CTTGACATAGGCACAAATTCCGTCGGCTGGGCGGTGACCGATCCAGA sequence (nt) ATACAACGTTCTGCGCTTCAAAGGAAACTCCATGTGGGGCGCGAGAT (not codon TGTTTGAGGAGGCAAAACCGGCGGCGGATCGGCGCGCGGCGCGCA optimized) TTGCCCGCCGGCGTCTGGAGCGGAGAAAACAGAGACTTTTGTATCTG GAAGAATTCTTCTCTGCGGAGATCGGGAAGTCTGACCCGCTCTTTTT CCGACGGCTGCATGAAAGCGCGCTCTTTCCGGACGATAAGACAACC GGAACCGGGAAATACACGCTTTTCAATGACCCGGATTTTACGGATCG GGATTATCTGCGCCGGTACCCCACGGTGTACCACCTGCGCAGCGAA CTGGTGCATTCCTCCGAGCCGCACGACCCGCGGCTCGTATTTCTGG CGCTGCACCATATCATCAAGCACCGCGGGCATTTCCTTTATGAAACAT CGGAGAACGGCGACGAATGCCCGACGGTAGACGCTGCGATGCAGGA GCTGACAGAGTATCTCTCAGCGGAAATGAATGTGGAGCTGGGATTGG AGCCGGAGTACTTCACGATCCTTACCGACCGCAATATGCGCATATCC CAGAAGAAGGCGGCGCTGCGCGCCGCTCTGGGGAGGTCGGTGCAG ACGGAGGAGAGCGGCGCGGATATTCAGGCCCTGTCCGATCTGCTTG CGGGGGCCAAAGTCAAGCTGGCTGCGCTCTATGACGATGAGTCGCT CAAAAATGCCGACCCGGCGAGCATCGCGCTGAGCGACGATCTGGAC GCCGTGTTTGACGCTCTCTCGTCGGCGTTGGGCGAGCGGGTGGAGC TGCTTCTTGCGGCAAAAAAGGTGTTTGATGCGGCACGGCTGTCGGAA ATGCGCGGCGGCGAAGCGTATCTCTGCGATGCAAAGGTCCGTCAGT ATGAGAAAAACCATGCCGACCTTCGTGCGCTGAAAGACTATGTGAGA GAAGCGGCGCCGGACAAATACCGGAACATATTCTATAAAAACGCGAA GTTCCCGAATAACTATGCCGCCTACAGCCGCTACCGTTCGGAGAGCG GGGATAAGACCTGCAAGCAGGAAGACTTTTGCGCGTTCCTCAAGTCT GTGCTGCCGAAGCCGGCGGCGGAGCATTCGTTCGCGCCGCAGTATG TGCAGATCGAAGAGAAGACGTTTCTTCCGCGGCTGACCGGCAGCGA AAACGGCATCATCCCGTATCAGCTCCACCGGCAGGAGCTGCGGAAA ATTCTGGATAACGCATCGGCGTATCTGCCGTTCCTGCTCGAAGAGGA CGAGGACGGCATCTCCGTGCGCGAGAAGATCGAAGCGGTGTTTTCC TTCCGTGTGCCATATTATGTCGGGCCGCTGAAAGGCCGTTGGGCAGA GCGGACTGAGGAAGCAAAGAGAACCGGCGAGAAAGCCTACCCTTGG TABLE 1J EAJR Type II Cas Sequences Name Sequence SEQ ID NO AATTTTGAAAAAGTTGTGGATCTCGACGCGAGCGCGGCGGCCTTTCT GAAAGACCTGATCGGAAAATGCACCTACACCGGCGCGCCGGTGCTG CCGAAGGACTCTCTGCTGTACAGCGAATTTATGCTTCTCAACGAGCT GAACCCGCTGCGTCTCAACGGAAACGAGCTCCCGGTCAGCGTAAAG CAGCAGTTGATCGAAGACCTGTTCAAAAAAGAAAACCGCCGCGTGAC GAAAAAACGCATTCTGGACTATCTGGTCATGCACGGCTATGCCGCAA AAACCGATGAGATCAGCGGCATCGACGACCCGGTGAAGACCAGACT GCGTTCGTACCACGATTTTGCGCCGCTCATCCCCGTGCTCGGCGAG GAAGCGGTGGAAAAGATCATCGAAGGAACGCTGGTATACAGCGACG ACAAGCGTATGCTGCGCCGCTGGCTGAAAAAGAATTTCCCTGCGCTG AACGAGGACGATGTGCGGAAGATCTGCCGTCTGAAATATGCCGACTG GGGACGCCTTTCCAAAGAGTTTTTGACGGAACTCTATTCCGGCGCGG ATGAGCAGGGCGAGGCGCAGAACATTCTGTATTTTCTGCGAAACACC AATCTCAATCTCATGCAGCTTCTCAATGATCCGGAGCATGCCAATCTT CCGGCCGCGGCGGAAGAGCGGCGGCGCGCTCGTTTCGGAAGAGAC AGTCTCCGCTCACAGCTGGACGAAATGTACATTTCTCCCGCCGTGCG CCGCGGCATATGGCAGACGCTGAAGATCTCCGAAGAGATCGCCCAT ATCCGCGGCGGCGCACCGGCGAAGATCTTTGTCGAGGTGGCGCGC GGCGAGGATAAGGAGAAGAAGGGAACGCGAACCAAATCCAGAAAAG AACAGCTTCTGGAGCTCTATAAGGCCTGCGAGCGGGACAGCGGCGA GCTGTACGAAAAGCTTGCGGCGACGGAGGAGAGCGCGCTGCGGAG AGACAAGCTCTTCCTGTACTATACACAGTTCGGGAAATGCATGTATTC CGGCGAGCCCATCGATCTGGAAACGGCTCTGCGCGATGATACAACG TATGATATCGACCATATTTTCCCGCGCTCCCGCATCAAGGACGACAG CATTGAGAACCGCGTGCTCGTAAAGAGCGTTCTCAACCGCGAAAAAA CGAACACCTATCCGATCTCGCAGGATATCCGCAGCCGGATGCTGCC GTTCTGGTCGATGCTGCACGATAAGAAAATGATCGGCGATAAAAAGT ACGACCGCCTGAAACGGAATACAGAACTGACGGAGGACGAACTGGC GTCCTTTGTACAGCGCCAACTGACGGAAACGCAGCAGTCCACCAAGG CGCTTGCCCAGCTTTTAAAGGAACGGTACCCGGAGTCGGAGATCGTA TACTCCAAGGCGGGCAACATTTCGGAGTTCCGGCAGAAGTTTGAAAT GCTCAAATGCCGCGAGGTGAACGACCTGCACCACGCCAAGGACGCC TATCTGAACATCGTTGTCGGCAACGTATATAAGACAGAGTTTACCGAC CGGTTTTTCCTGAATATCCGAAAAGAGAATTACAGTCTCAAAAAAGTC TTTGACGGGGATACGCCGGGCGCGTGGAAAGCGGACGGAACGAGC ATCGCCACGGTGAAAAAGTACATGGCGAAGAATAACCCCATCATCAC CCGCATGCCGCGAGAGGAGAAGGGACAGCTCTTCAACCTGAACATC ATGCCTGCCGGACTGGGCCAGCTTCCGATCAAAGCCGGAAAACCCA TTGAGAAATACGGCGGATACAACAAGCTGGCCGGTTCGTATTACTGT GTCGTGGAGCATACGGAAAAGAAGAAACGTGTTCGTTCCATCGAAAC GGTACTGCTTTACGCACGGGGACTGTATGAGAGCGACCCGGTAAAAT ACTGCCGTGAGATACTTGGCCTTGTTGACCCGGTCGTTGTTGCACCG AAGATCCGAATAAACTCCCTTTTGCAATTGGATGGTTTCCGCTTGTAC CCGTCCGGAAGAACCGGAAACAGAATCATTTACAAACACGCCTATCA GCTTGCCATTGACGCCGAAAGAGAACGCTACATCCGTAATCTGACAA AATACATAGAGCGCTGTGCCAAAATCGAATATCCGGCAACAGCCTTT GACGGGATCAGTGCGGAAGAGAATGTGGCGCTGTACAACTGGTTTGT CGAAAGACTGGAAAGCAGCGCATATGGAAAACTCATGCAGGCTCTTC TTGCCAATCTGAAAGCAAGTGATGAGACGTTTAAAAACATGGATATCC TTTCACAGGCAAAGACGCTGATGGAAGTTTTAAAAGCATTTAAGTGCG ATGCGCAAAGCTCAAATTTAAAGGACCTCTGCGGCAAAGGGTCGGTC GGAACAATCCTGAAAAATGCTTCACTGCAAGGCGTCCAATCGGCCTA TCTTATCCACCAGTCGCCGACCGGTCTGTACGAGTATCGTACCGACC TTCTGCACTGA EAJR Type II GAAGAAAAGAAAACCATCCGGAGTCACAGTCCTTACTACATCGGCCT 53 Cas coding CGACATCGGCACCAACAGCGTGGGCTGGGCCGTGACCGATCCTGAA sequence (nt) TACAACGTGCTGAGATTCAAGGGAAACAGCATGTGGGGCGCTAGGCT TABLE 1J EAJR Type II Cas Sequences Name Sequence SEQ ID NO (human GTTCGAAGAGGCCAAACCCGCCGCTGATAGACGGGCCGCCAGGATC codon- GCCAGGCGCCGGCTGGAAAGACGCAAGCAGAGACTGCTCTACCTGG optimized; AAGAGTTTTTCAGCGCTGAGATCGGCAAGTCTGACCCTCTGTTTTTCA lacking N- GACGGCTGCACGAGTCCGCCCTGTTCCCCGACGATAAGACAACCGG terminal CACCGGCAAGTACACCCTGTTCAACGACCCTGACTTTACCGACCGGG methionine; ACTACCTTAGAAGATACCCTACCGTGTACCACCTGAGATCTGAGCTG no stop GTGCACAGCAGCGAACCCCACGATCCTAGACTGGTGTTCCTGGCCCT codon) GCACCACATCATCAAGCATAGAGGTCACTTCCTCTACGAAACCAGCG AGAACGGCGATGAGTGCCCCACCGTTGATGCCGCCATGCAGGAGCT GACAGAGTACCTATCTGCCGAGATGAACGTGGAACTGGGCCTGGAA CCTGAGTACTTCACTATCCTGACCGACCGTAACATGCGAATCTCACA GAAGAAGGCCGCCCTGAGAGCCGCTCTGGGTAGAAGCGTGCAGACC GAAGAGTCTGGAGCCGATATACAGGCCCTGAGCGATCTGCTGGCCG GAGCCAAAGTGAAGCTCGCCGCCCTTTACGACGACGAATCTCTGAAG AACGCCGACCCTGCCAGCATCGCCCTGTCCGACGACCTGGATGCTG TGTTTGACGCTCTGAGCAGCGCCTTGGGCGAGAGAGTCGAACTGCT GCTGGCTGCCAAAAAAGTGTTCGACGCCGCCAGACTGAGCGAGATG AGAGGAGGCGAGGCTTACCTGTGTGATGCCAAGGTGCGGCAGTACG AGAAGAACCACGCCGACCTGAGAGCCCTGAAAGATTACGTGAGAGA GGCCGCCCCTGATAAGTACCGGAATATCTTCTACAAGAATGCTAAGTT CCCAAACAATTACGCTGCGTATAGCAGATACAGAAGCGAGTCTGGAG ACAAAACATGTAAGCAGGAGGATTTCTGCGCCTTCCTAAAGAGCGTG CTGCCCAAGCCTGCCGCTGAGCACAGCTTCGCCCCTCAGTACGTGC AGATCGAGGAAAAGACATTTCTGCCCAGGCTCACCGGGAGCGAGAA CGGCATCATCCCTTACCAGCTGCACAGACAGGAGCTGCGGAAGATC CTGGACAACGCCAGCGCCTACCTGCCCTTCCTCCTGGAGGAGGACG AGGACGGCATCAGCGTGCGGGAAAAGATCGAGGCAGTGTTCAGCTT TCGGGTCCCCTACTACGTGGGCCCTCTGAAGGGCAGATGGGCCGAA AGAACCGAGGAAGCCAAGAGAACCGGCGAAAAGGCCTATCCTTGGA ATTTCGAGAAGGTGGTGGATCTGGATGCCAGCGCCGCCGCTTTTCTA AAGGACCTGATCGGCAAGTGTACCTACACCGGCGCCCCAGTGCTGC CTAAGGACAGCCTGCTGTACTCTGAGTTCATGCTGCTGAATGAGCTG AACCCTCTGCGACTGAACGGCAATGAGCTCCCCGTGTCCGTGAAGCA ACAACTCATCGAGGACCTGTTCAAGAAAGAAAACAGAAGGGTGACAA AGAAGAGAATCCTGGACTACCTGGTGATGCACGGCTACGCCGCCAA GACCGATGAGATCAGCGGCATCGACGACCCTGTGAAAACCAGACTG CGAAGCTACCACGACTTCGCCCCGCTGATCCCCGTGCTGGGCGAAG AAGCTGTGGAGAAAATCATTGAAGGCACCCTGGTGTACAGCGATGAC AAGCGGATGCTGAGAAGATGGCTGAAGAAGAATTTCCCTGCTTTAAA TGAGGACGACGTGCGGAAGATCTGCAGACTGAAATATGCCGACTGG GGTAGACTGAGCAAGGAGTTCCTGACAGAACTGTACAGCGGCGCAG ATGAGCAGGGCGAGGCCCAGAACATCCTGTATTTTTTGAGAAACACC AACCTGAATCTGATGCAGCTGCTGAACGACCCCGAACACGCCAACCT GCCTGCGGCTGCTGAAGAGAGAAGAAGAGCCAGATTTGGCAGAGAT TCCCTGAGATCCCAACTCGACGAGATGTACATCAGCCCCGCCGTGAG AAGAGGCATTTGGCAGACCCTGAAGATCAGCGAAGAAATCGCCCACA TCCGGGGAGGAGCCCCTGCCAAGATCTTTGTGGAAGTGGCCCGGGG AGAGGACAAAGAAAAGAAGGGCACAAGAACCAAGTCCAGAAAGGAA CAGCTGCTCGAACTGTATAAGGCCTGCGAGCGCGACAGCGGCGAGC TGTACGAGAAGCTGGCCGCTACAGAAGAATCTGCTCTGAGGCGGGA CAAGCTGTTCCTGTACTACACCCAGTTCGGCAAGTGCATGTACAGCG GCGAACCTATCGATCTGGAGACAGCCCTGCGGGACGACACCACCTA CGACATCGACCACATCTTCCCTAGAAGCAGAATTAAGGATGATTCAAT CGAAAACCGGGTGCTGGTCAAGAGCGTGCTGAACCGGGAAAAGACC AACACCTACCCTATCTCCCAGGACATCCGGAGCAGAATGCTGCCTTT CTGGAGCATGCTGCACGACAAAAAGATGATCGGCGACAAGAAGTATG ACAGACTGAAGAGAAACACCGAGCTCACCGAGGACGAGCTGGCCAG TABLE 1J EAJR Type II Cas Sequences Name Sequence SEQ ID NO CTTCGTGCAAAGACAGCTGACCGAGACACAGCAGAGCACCAAGGCC CTCGCTCAGTTACTGAAAGAAAGATACCCAGAGTCCGAGATCGTTTA CAGCAAGGCCGGAAACATCTCTGAGTTCCGGCAGAAATTCGAGATGC TGAAATGTAGAGAGGTGAACGACCTGCATCATGCCAAAGACGCCTAC CTGAATATCGTGGTAGGCAACGTGTACAAGACCGAGTTCACAGATAG ATTCTTCCTGAATATTAGGAAGGAGAACTACTCTCTGAAGAAGGTGTT CGATGGAGATACCCCTGGCGCCTGGAAGGCCGACGGAACGAGCATC GCCACCGTGAAGAAGTATATGGCCAAGAACAACCCCATCATCACAAG AATGCCTAGAGAAGAGAAAGGGCAGCTGTTTAACCTGAACATCATGC CTGCCGGCCTGGGCCAGCTGCCAATCAAGGCCGGCAAACCTATCGA GAAGTATGGCGGCTACAACAAGCTGGCAGGCTCTTACTACTGCGTGG TCGAGCACACCGAAAAAAAGAAGCGGGTGAGAAGCATCGAAACCGT GCTGCTGTACGCTAGAGGCCTGTACGAATCCGATCCAGTGAAGTACT GCAGAGAGATTCTGGGCCTGGTGGACCCTGTCGTGGTGGCCCCTAA GATCAGAATCAACAGCCTGTTGCAGCTGGACGGCTTCAGACTCTACC CCAGCGGAAGAACAGGAAACCGGATCATCTACAAGCACGCCTACCA GCTGGCTATCGACGCTGAGAGAGAGAGATACATTCGGAACCTGACAA AGTACATCGAGCGGTGCGCCAAGATCGAGTACCCAGCTACTGCCTTT GACGGCATCAGCGCCGAGGAGAACGTCGCCCTGTACAATTGGTTCG TGGAACGGCTGGAGAGCAGCGCATACGGCAAACTGATGCAAGCCCT GCTGGCCAACCTGAAGGCCAGCGACGAGACATTCAAAAATATGGACA TCTTGAGCCAGGCCAAAACCCTGATGGAAGTGCTGAAGGCCTTCAAG TGCGATGCCCAGAGCAGCAACCTGAAAGACCTGTGCGGCAAGGGCA GCGTGGGAACCATCCTGAAAAACGCCTCTCTGCAGGGCGTGCAATCT GCCTACCTGATCCACCAGAGCCCAACAGGCCTGTACGAGTACAGAAC AGATCTGCTGCAC EAJR Type II ATGGGCAAACCTATCCCCAATCCCCTGCTGGGCCTGGACAGCACCAA 54 Cas GCGGACTGCCGACGGCAGTGAGTTCGAGTCTCCCAAGAAAAAGAGG mammalian AAAGTGGGATCCGAAGAAAAGAAAACCATCCGGAGTCACAGTCCTTA expression CTACATCGGCCTCGACATCGGCACCAACAGCGTGGGCTGGGCCGTG construct ACCGATCCTGAATACAACGTGCTGAGATTCAAGGGAAACAGCATGTG (with N- GGGCGCTAGGCTGTTCGAAGAGGCCAAACCCGCCGCTGATAGACGG terminal GCCGCCAGGATCGCCAGGCGCCGGCTGGAAAGACGCAAGCAGAGA methionine, CTGCTCTACCTGGAAGAGTTTTTCAGCGCTGAGATCGGCAAGTCTGA stop codon, CCCTCTGTTTTTCAGACGGCTGCACGAGTCCGCCCTGTTCCCCGACG V5 tag, N- ATAAGACAACCGGCACCGGCAAGTACACCCTGTTCAACGACCCTGAC terminal NLS TTTACCGACCGGGACTACCTTAGAAGATACCCTACCGTGTACCACCT and C- GAGATCTGAGCTGGTGCACAGCAGCGAACCCCACGATCCTAGACTG terminal NLS) GTGTTCCTGGCCCTGCACCACATCATCAAGCATAGAGGTCACTTCCT (nt) CTACGAAACCAGCGAGAACGGCGATGAGTGCCCCACCGTTGATGCC GCCATGCAGGAGCTGACAGAGTACCTATCTGCCGAGATGAACGTGG AACTGGGCCTGGAACCTGAGTACTTCACTATCCTGACCGACCGTAAC ATGCGAATCTCACAGAAGAAGGCCGCCCTGAGAGCCGCTCTGGGTA GAAGCGTGCAGACCGAAGAGTCTGGAGCCGATATACAGGCCCTGAG CGATCTGCTGGCCGGAGCCAAAGTGAAGCTCGCCGCCCTTTACGAC GACGAATCTCTGAAGAACGCCGACCCTGCCAGCATCGCCCTGTCCG ACGACCTGGATGCTGTGTTTGACGCTCTGAGCAGCGCCTTGGGCGA GAGAGTCGAACTGCTGCTGGCTGCCAAAAAAGTGTTCGACGCCGCC AGACTGAGCGAGATGAGAGGAGGCGAGGCTTACCTGTGTGATGCCA AGGTGCGGCAGTACGAGAAGAACCACGCCGACCTGAGAGCCCTGAA AGATTACGTGAGAGAGGCCGCCCCTGATAAGTACCGGAATATCTTCT ACAAGAATGCTAAGTTCCCAAACAATTACGCTGCGTATAGCAGATACA GAAGCGAGTCTGGAGACAAAACATGTAAGCAGGAGGATTTCTGCGCC TTCCTAAAGAGCGTGCTGCCCAAGCCTGCCGCTGAGCACAGCTTCGC CCCTCAGTACGTGCAGATCGAGGAAAAGACATTTCTGCCCAGGCTCA CCGGGAGCGAGAACGGCATCATCCCTTACCAGCTGCACAGACAGGA GCTGCGGAAGATCCTGGACAACGCCAGCGCCTACCTGCCCTTCCTC TABLE 1J EAJR Type II Cas Sequences Name Sequence SEQ ID NO CTGGAGGAGGACGAGGACGGCATCAGCGTGCGGGAAAAGATCGAG GCAGTGTTCAGCTTTCGGGTCCCCTACTACGTGGGCCCTCTGAAGGG CAGATGGGCCGAAAGAACCGAGGAAGCCAAGAGAACCGGCGAAAAG GCCTATCCTTGGAATTTCGAGAAGGTGGTGGATCTGGATGCCAGCGC CGCCGCTTTTCTAAAGGACCTGATCGGCAAGTGTACCTACACCGGCG CCCCAGTGCTGCCTAAGGACAGCCTGCTGTACTCTGAGTTCATGCTG CTGAATGAGCTGAACCCTCTGCGACTGAACGGCAATGAGCTCCCCGT GTCCGTGAAGCAACAACTCATCGAGGACCTGTTCAAGAAAGAAAACA GAAGGGTGACAAAGAAGAGAATCCTGGACTACCTGGTGATGCACGG CTACGCCGCCAAGACCGATGAGATCAGCGGCATCGACGACCCTGTG AAAACCAGACTGCGAAGCTACCACGACTTCGCCCCGCTGATCCCCGT GCTGGGCGAAGAAGCTGTGGAGAAAATCATTGAAGGCACCCTGGTG TACAGCGATGACAAGCGGATGCTGAGAAGATGGCTGAAGAAGAATTT CCCTGCTTTAAATGAGGACGACGTGCGGAAGATCTGCAGACTGAAAT ATGCCGACTGGGGTAGACTGAGCAAGGAGTTCCTGACAGAACTGTAC AGCGGCGCAGATGAGCAGGGCGAGGCCCAGAACATCCTGTATTTTTT GAGAAACACCAACCTGAATCTGATGCAGCTGCTGAACGACCCCGAAC ACGCCAACCTGCCTGCGGCTGCTGAAGAGAGAAGAAGAGCCAGATT TGGCAGAGATTCCCTGAGATCCCAACTCGACGAGATGTACATCAGCC CCGCCGTGAGAAGAGGCATTTGGCAGACCCTGAAGATCAGCGAAGA AATCGCCCACATCCGGGGAGGAGCCCCTGCCAAGATCTTTGTGGAA GTGGCCCGGGGAGAGGACAAAGAAAAGAAGGGCACAAGAACCAAGT CCAGAAAGGAACAGCTGCTCGAACTGTATAAGGCCTGCGAGCGCGA CAGCGGCGAGCTGTACGAGAAGCTGGCCGCTACAGAAGAATCTGCT CTGAGGCGGGACAAGCTGTTCCTGTACTACACCCAGTTCGGCAAGTG CATGTACAGCGGCGAACCTATCGATCTGGAGACAGCCCTGCGGGAC GACACCACCTACGACATCGACCACATCTTCCCTAGAAGCAGAATTAA GGATGATTCAATCGAAAACCGGGTGCTGGTCAAGAGCGTGCTGAACC GGGAAAAGACCAACACCTACCCTATCTCCCAGGACATCCGGAGCAGA ATGCTGCCTTTCTGGAGCATGCTGCACGACAAAAAGATGATCGGCGA CAAGAAGTATGACAGACTGAAGAGAAACACCGAGCTCACCGAGGAC GAGCTGGCCAGCTTCGTGCAAAGACAGCTGACCGAGACACAGCAGA GCACCAAGGCCCTCGCTCAGTTACTGAAAGAAAGATACCCAGAGTCC GAGATCGTTTACAGCAAGGCCGGAAACATCTCTGAGTTCCGGCAGAA ATTCGAGATGCTGAAATGTAGAGAGGTGAACGACCTGCATCATGCCA AAGACGCCTACCTGAATATCGTGGTAGGCAACGTGTACAAGACCGAG TTCACAGATAGATTCTTCCTGAATATTAGGAAGGAGAACTACTCTCTG AAGAAGGTGTTCGATGGAGATACCCCTGGCGCCTGGAAGGCCGACG GAACGAGCATCGCCACCGTGAAGAAGTATATGGCCAAGAACAACCCC ATCATCACAAGAATGCCTAGAGAAGAGAAAGGGCAGCTGTTTAACCT GAACATCATGCCTGCCGGCCTGGGCCAGCTGCCAATCAAGGCCGGC AAACCTATCGAGAAGTATGGCGGCTACAACAAGCTGGCAGGCTCTTA CTACTGCGTGGTCGAGCACACCGAAAAAAAGAAGCGGGTGAGAAGC ATCGAAACCGTGCTGCTGTACGCTAGAGGCCTGTACGAATCCGATCC AGTGAAGTACTGCAGAGAGATTCTGGGCCTGGTGGACCCTGTCGTG GTGGCCCCTAAGATCAGAATCAACAGCCTGTTGCAGCTGGACGGCTT CAGACTCTACCCCAGCGGAAGAACAGGAAACCGGATCATCTACAAGC ACGCCTACCAGCTGGCTATCGACGCTGAGAGAGAGAGATACATTCGG AACCTGACAAAGTACATCGAGCGGTGCGCCAAGATCGAGTACCCAGC TACTGCCTTTGACGGCATCAGCGCCGAGGAGAACGTCGCCCTGTACA ATTGGTTCGTGGAACGGCTGGAGAGCAGCGCATACGGCAAACTGAT GCAAGCCCTGCTGGCCAACCTGAAGGCCAGCGACGAGACATTCAAA AATATGGACATCTTGAGCCAGGCCAAAACCCTGATGGAAGTGCTGAA GGCCTTCAAGTGCGATGCCCAGAGCAGCAACCTGAAAGACCTGTGC GGCAAGGGCAGCGTGGGAACCATCCTGAAAAACGCCTCTCTGCAGG GCGTGCAATCTGCCTACCTGATCCACCAGAGCCCAACAGGCCTGTAC TABLE 1J EAJR Type II Cas Sequences Name Sequence SEQ ID NO GAGTACAGAACAGATCTGCTGCACTCTAGAAAGCGGACAGCAGACG GCTCCGAATTTGAAAGCCCTAAGAAAAAGAGAAAGGTGTGA [0100] In some embodiments an EAJR Type II Cas protein comprises an amino acid sequence of SEQ ID NO:49, SEQ ID NO:50, or SEQ ID NO:51. In some embodiments, an EAJR Type II Cas protein has nickase activity, for example resulting from one or more amino acid substitutions relative to the sequence of SEQ ID NO:49, SEQ ID NO:50, or SEQ ID NO:51. In some embodiments, the one or more amino acid substitutions providing nickase activity comrpise a D18A substitution, wherein the position of the D18A substitution is defined with respect to the amino acid numbering of SEQ ID NO:50. In some embodiments, the one or more amino acid substitutions providing nickase activity comrpise an H587A substitution, wherein the position of the N610A substitution is defined with respect to the amino acid numbering of SEQ ID NO:50. In some embodiments, an EAJR Type II Cas protein is catalytically inactive, for example due to a D18A substitution in combination with a H587A substitution. 6.2.1.10. CFDE Type II Cas Proteins [0101] In one aspect, the disclosure provides CFDE Type II Cas proteins. CFDE Type II Cas proteins can be further classified as Type IIA Cas proteins. The CFDE Type II Cas proteins typically comprise an amino acid sequence that is at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 85%, at least 90%, or at least 95% identical to SEQ ID NO:55. In some embodiments, the CFDE Type II Cas proteins comprise an amino acid sequence that is at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical to SEQ ID NO:55. In some embodiments, a CFDE Type II Cas protein comprises an amino acid sequence that is identical to SEQ ID NO:55. [0102] Exemplary CFDE Type II Cas protein sequences and nucleotide sequences encoding exemplary CFDE Type II Cas proteins are set forth in Table 1K. TABLE 1K CFDE Type II Cas Sequences Name Sequence SEQ ID NO CFDE Type II NLRNYSGDYNIGLDMGTGSVGWAVTDAQGKLLHFKKQPTWGSRLFDSA 55 Cas amino QPASEARVHRGQRRRYVRRRWRLDLLQKLFEPAMSEIDPGFFMRLNQS acid RTIEGDPIFTKDFTKADYYDRFPTIYHLRAYLMDTDEQADLRLVYLAIHNIV sequence KHRGNFLRQGEKLTAKTAKTSDALERLNDSLQGWCESREYEVAKINKSA (without N- VAKILADERSSRSQKAKDIQALVKVTTGDAAADKKLAKGLSNAVVGLQCE terminal FKDVFGEFECEATKLALSDDEKLETLQAACPDDCVELLESVCGVYSAYV methionine) LQGLLSYAEGQTISINMIKKYEQYAEDLALLKLLVRKYALDSYEPFFRGAT YHDSNSDDPSRDYDASKATGYTAYDLHKLSYDDFAKSVKKLFAGTGAET DERYIAMMDAFDKQRFLRRLKTSDNGAIYYQLHLEELQAILKNQGRFYPF LKDEAAKIESLVTFRIPYYVGPLTSKNAAQDKHGKPRFQWSERKPGMED ATITPWNWDAVIDKNKSAENFILRMTGDCTYLAGEKVLPKQSLLYEEFCV LNELNGVRWSEDGDNWQRLDAAQREGIIHELFHKKRRVTYKMIADWLV KEGDATNPVVRGGQRESGLESKMGSYIFFAKDIFGVDELSRAEYPVIEKII LWNTLFEDRSILKEKLQDEFGPSGNGMLDAEQIKKICKKRLTGWGRLSE KFLTGIRIDTQAGRSMSIMDVLREGNPSSEHRLGETMVMMEILHDNTLGF QEKVDEHNRKYYAEHEKSLGVNELPGSPAIRRSLNQAIRIVDEIAKIAGHA PANVFVEVTRDEDEKKKGKRTTKRWDAIEDALKAFKAEGGDLAVVDDFK ELRAANVDLDERLTLYLMQNGKCLYSGRAIDLNKLMAGSGEYEVDHIIPR AYIKDDSLENKALVYREENQHKTDQLLIDKSIRWKMGETWKQLHQAKLIG TABLE 1K CFDE Type II Cas Sequences Name Sequence SEQ ID NO DKKYRNLLRSSINENAMKGFVARQLVETSQMVKMAQALLEARYADEGTK IVPVKASVSHNLREAAGLVKCREANDFHHAHDAYLACRLGLFIQMRHPG MYENPIGYSHVIKKYVREQSELFKKTHRMPGSAGFVVNSFMTAGFDPET GEVFDDAWTEDDEVKRYRGKYLPDGWNAAAEVESIRRALNYRQCYISR MPQEDTGAFWKATIYSPRDPKMGAKLELSTKQGLDAKRYGGFSSQQFA YFFIYEAKKKGKPVFRFSQVPVWLAARIEGDSAALGEYARGLAEKEGLEY VGITRPKIYKKQLIEVDGERFFITGKKEMRNATQLAFSNEQMKLLAGDTC SEDAIKLLDMFNPAASSVSGRLLRQLKIEAIIEKCATVDAEDVVAVVKRLL ALVNGTMNMVDLSPAGGSKFSGCIQPNYSKLLSDSNVTVFLIDQSVTGM FERRTRVGL CFDE Type II MNLRNYSGDYNIGLDMGTGSVGWAVTDAQGKLLHFKKQPTWGSRLFD 56 Cas amino SAQPASEARVHRGQRRRYVRRRWRLDLLQKLFEPAMSEIDPGFFMRLN acid QSRTIEGDPIFTKDFTKADYYDRFPTIYHLRAYLMDTDEQADLRLVYLAIH sequence NIVKHRGNFLRQGEKLTAKTAKTSDALERLNDSLQGWCESREYEVAKIN KSAVAKILADERSSRSQKAKDIQALVKVTTGDAAADKKLAKGLSNAVVGL QCEFKDVFGEFECEATKLALSDDEKLETLQAACPDDCVELLESVCGVYS AYVLQGLLSYAEGQTISINMIKKYEQYAEDLALLKLLVRKYALDSYEPFFR GATYHDSNSDDPSRDYDASKATGYTAYDLHKLSYDDFAKSVKKLFAGTG AETDERYIAMMDAFDKQRFLRRLKTSDNGAIYYQLHLEELQAILKNQGRF YPFLKDEAAKIESLVTFRIPYYVGPLTSKNAAQDKHGKPRFQWSERKPG MEDATITPWNWDAVIDKNKSAENFILRMTGDCTYLAGEKVLPKQSLLYEE FCVLNELNGVRWSEDGDNWQRLDAAQREGIIHELFHKKRRVTYKMIAD WLVKEGDATNPVVRGGQRESGLESKMGSYIFFAKDIFGVDELSRAEYPV IEKIILWNTLFEDRSILKEKLQDEFGPSGNGMLDAEQIKKICKKRLTGWGR LSEKFLTGIRIDTQAGRSMSIMDVLREGNPSSEHRLGETMVMMEILHDNT LGFQEKVDEHNRKYYAEHEKSLGVNELPGSPAIRRSLNQAIRIVDEIAKIA GHAPANVFVEVTRDEDEKKKGKRTTKRWDAIEDALKAFKAEGGDLAVVD DFKELRAANVDLDERLTLYLMQNGKCLYSGRAIDLNKLMAGSGEYEVDH IIPRAYIKDDSLENKALVYREENQHKTDQLLIDKSIRWKMGETWKQLHQA KLIGDKKYRNLLRSSINENAMKGFVARQLVETSQMVKMAQALLEARYAD EGTKIVPVKASVSHNLREAAGLVKCREANDFHHAHDAYLACRLGLFIQM RHPGMYENPIGYSHVIKKYVREQSELFKKTHRMPGSAGFVVNSFMTAGF DPETGEVFDDAWTEDDEVKRYRGKYLPDGWNAAAEVESIRRALNYRQC YISRMPQEDTGAFWKATIYSPRDPKMGAKLELSTKQGLDAKRYGGFSSQ QFAYFFIYEAKKKGKPVFRFSQVPVWLAARIEGDSAALGEYARGLAEKE GLEYVGITRPKIYKKQLIEVDGERFFITGKKEMRNATQLAFSNEQMKLLAG DTCSEDAIKLLDMFNPAASSVSGRLLRQLKIEAIIEKCATVDAEDVVAVVK RLLALVNGTMNMVDLSPAGGSKFSGCIQPNYSKLLSDSNVTVFLIDQSVT GMFERRTRVGL CFDE Type II MGKPIPNPLLGLDSTKRTADGSEFESPKKKRKVGSNLRNYSGDYNIGLD 57 Cas MGTGSVGWAVTDAQGKLLHFKKQPTWGSRLFDSAQPASEARVHRGQR mammalian RRYVRRRWRLDLLQKLFEPAMSEIDPGFFMRLNQSRTIEGDPIFTKDFTK expression ADYYDRFPTIYHLRAYLMDTDEQADLRLVYLAIHNIVKHRGNFLRQGEKL construct TAKTAKTSDALERLNDSLQGWCESREYEVAKINKSAVAKILADERSSRS (with N- QKAKDIQALVKVTTGDAAADKKLAKGLSNAVVGLQCEFKDVFGEFECEA terminal TKLALSDDEKLETLQAACPDDCVELLESVCGVYSAYVLQGLLSYAEGQTI methionine, SINMIKKYEQYAEDLALLKLLVRKYALDSYEPFFRGATYHDSNSDDPSRD stop codon, YDASKATGYTAYDLHKLSYDDFAKSVKKLFAGTGAETDERYIAMMDAFD V5 tag, N- KQRFLRRLKTSDNGAIYYQLHLEELQAILKNQGRFYPFLKDEAAKIESLVT terminal NLS FRIPYYVGPLTSKNAAQDKHGKPRFQWSERKPGMEDATITPWNWDAVI and C- DKNKSAENFILRMTGDCTYLAGEKVLPKQSLLYEEFCVLNELNGVRWSE terminal NLS) DGDNWQRLDAAQREGIIHELFHKKRRVTYKMIADWLVKEGDATNPVVR (aa) GGQRESGLESKMGSYIFFAKDIFGVDELSRAEYPVIEKIILWNTLFEDRSIL KEKLQDEFGPSGNGMLDAEQIKKICKKRLTGWGRLSEKFLTGIRIDTQAG RSMSIMDVLREGNPSSEHRLGETMVMMEILHDNTLGFQEKVDEHNRKY YAEHEKSLGVNELPGSPAIRRSLNQAIRIVDEIAKIAGHAPANVFVEVTRD EDEKKKGKRTTKRWDAIEDALKAFKAEGGDLAVVDDFKELRAANVDLDE TABLE 1K CFDE Type II Cas Sequences Name Sequence SEQ ID NO RLTLYLMQNGKCLYSGRAIDLNKLMAGSGEYEVDHIIPRAYIKDDSLENK ALVYREENQHKTDQLLIDKSIRWKMGETWKQLHQAKLIGDKKYRNLLRS SINENAMKGFVARQLVETSQMVKMAQALLEARYADEGTKIVPVKASVSH NLREAAGLVKCREANDFHHAHDAYLACRLGLFIQMRHPGMYENPIGYSH VIKKYVREQSELFKKTHRMPGSAGFVVNSFMTAGFDPETGEVFDDAWT EDDEVKRYRGKYLPDGWNAAAEVESIRRALNYRQCYISRMPQEDTGAF WKATIYSPRDPKMGAKLELSTKQGLDAKRYGGFSSQQFAYFFIYEAKKK GKPVFRFSQVPVWLAARIEGDSAALGEYARGLAEKEGLEYVGITRPKIYK KQLIEVDGERFFITGKKEMRNATQLAFSNEQMKLLAGDTCSEDAIKLLDM FNPAASSVSGRLLRQLKIEAIIEKCATVDAEDVVAVVKRLLALVNGTMNM VDLSPAGGSKFSGCIQPNYSKLLSDSNVTVFLIDQSVTGMFERRTRVGLS RKRTADGSEFESPKKKRKV CFDE Type II ATGAATTTAAGAAACTACTCTGGCGACTACAATATCGGCCTCGACATG 58 Cas coding GGAACCGGCTCCGTTGGTTGGGCGGTGACTGATGCCCAGGGCAAGC sequence (nt) TTCTGCATTTTAAGAAGCAGCCGACGTGGGGTAGCCGCTTGTTCGAT (not codon AGTGCGCAGCCGGCATCCGAGGCGCGCGTTCATCGTGGCCAGCGTC optimized) GTCGTTACGTGCGTCGTCGCTGGCGTCTCGACTTGCTTCAAAAGCTG TTCGAACCCGCGATGAGCGAGATCGACCCCGGCTTTTTCATGCGACT CAATCAGTCGCGAACCATCGAAGGCGACCCGATCTTTACCAAGGACT TTACGAAGGCCGATTACTACGATCGTTTCCCGACCATCTATCATCTGC GCGCTTACCTTATGGACACCGATGAACAGGCCGATCTTCGTTTGGTG TATCTTGCGATCCATAACATCGTTAAGCATCGCGGTAACTTTTTGCGC CAGGGAGAGAAGCTGACGGCAAAAACGGCAAAGACGTCGGATGCCC TCGAGCGTCTTAATGACTCCCTGCAGGGTTGGTGCGAGTCGAGGGA ATATGAGGTTGCAAAAATCAATAAAAGCGCCGTCGCAAAGATCCTCG CTGACGAGAGGTCTTCTCGCTCGCAAAAGGCAAAAGACATTCAGGCG CTTGTAAAAGTTACCACTGGAGACGCTGCCGCAGATAAAAAGCTGGC GAAGGGCTTGTCGAATGCTGTCGTCGGCCTTCAGTGCGAGTTCAAAG ACGTGTTTGGCGAGTTTGAGTGCGAGGCAACTAAGCTTGCGCTTTCG GATGACGAGAAACTCGAGACACTGCAAGCCGCGTGTCCCGACGATT GCGTCGAGCTTCTTGAATCTGTCTGCGGGGTATATTCTGCATATGTTT TGCAGGGGCTTTTGTCATATGCAGAGGGCCAAACTATTTCGATCAAC ATGATTAAAAAATACGAGCAATACGCAGAAGACCTTGCTCTGCTTAAG CTCTTGGTCCGCAAATATGCTCTCGACTCATACGAGCCCTTCTTCCGT GGCGCGACCTATCATGATTCGAATAGCGATGACCCCTCGCGCGACTA CGATGCCTCGAAAGCAACGGGGTACACCGCGTATGACCTTCATAAGC TCTCCTACGATGACTTTGCCAAATCGGTAAAGAAGCTCTTTGCGGGAA CGGGCGCGGAGACGGATGAGCGCTATATCGCCATGATGGACGCATT CGACAAACAGCGCTTTTTGCGTCGCCTGAAGACGAGCGATAACGGC GCCATCTACTACCAGCTGCATCTTGAGGAACTTCAGGCTATTCTTAAA AACCAGGGGCGCTTTTATCCGTTCCTTAAGGATGAGGCTGCAAAAAT CGAGAGTCTGGTCACGTTCCGTATTCCGTACTATGTGGGGCCGCTGA CGTCTAAAAACGCTGCCCAGGACAAACACGGCAAGCCTCGATTTCAA TGGTCCGAGCGCAAGCCCGGCATGGAAGATGCGACGATTACGCCTT GGAACTGGGATGCCGTGATCGATAAGAATAAGAGCGCCGAGAACTTT ATCCTGCGCATGACGGGTGACTGCACGTATCTTGCTGGCGAAAAGGT GCTTCCCAAGCAGTCTTTGCTCTATGAGGAGTTTTGCGTTCTGAATGA ACTCAATGGCGTTCGCTGGTCCGAAGATGGTGACAATTGGCAACGCC TGGATGCCGCGCAGCGCGAGGGCATTATCCATGAGCTTTTCCATAAA AAGCGTCGCGTTACCTACAAGATGATTGCCGACTGGCTTGTAAAAGA GGGCGACGCGACAAATCCGGTTGTCCGTGGTGGCCAGCGTGAGAGC GGTCTTGAATCGAAGATGGGCTCGTACATCTTCTTTGCTAAGGACATC TTTGGTGTGGACGAGTTGAGCCGTGCCGAGTATCCGGTTATCGAGAA GATCATTCTTTGGAACACATTGTTTGAGGACCGTTCGATCCTTAAGGA AAAACTGCAGGATGAGTTTGGTCCAAGTGGCAATGGAATGCTCGATG CCGAGCAGATTAAGAAGATTTGCAAGAAGCGTCTAACTGGCTGGGGA CGTCTGTCGGAGAAGTTCCTGACGGGTATCAGGATTGATACCCAAGC TABLE 1K CFDE Type II Cas Sequences Name Sequence SEQ ID NO TGGACGCTCTATGAGCATCATGGACGTGCTGCGAGAGGGAAACCCC AGTTCTGAGCATCGCCTGGGCGAGACCATGGTGATGATGGAGATTCT CCACGACAACACGCTGGGCTTCCAGGAAAAAGTGGATGAACACAATC GCAAATACTACGCCGAACACGAGAAGTCTCTAGGCGTAAATGAGCTC CCTGGATCTCCGGCGATTCGTCGCAGCTTGAATCAAGCGATTCGCAT TGTTGATGAAATCGCAAAGATTGCGGGACACGCGCCGGCCAACGTTT TTGTTGAGGTGACTCGCGACGAGGACGAAAAGAAGAAGGGCAAGCG GACCACGAAGCGCTGGGACGCCATCGAGGATGCGCTCAAAGCGTTT AAGGCCGAGGGCGGCGACCTGGCTGTTGTGGATGATTTCAAGGAGC TCAGGGCAGCCAATGTAGACCTTGACGAGCGTCTGACCCTCTATTTG ATGCAGAACGGAAAATGCCTGTATTCTGGCCGCGCTATCGACCTCAA TAAGCTTATGGCGGGCTCGGGCGAGTATGAGGTCGACCATATCATTC CGCGCGCCTATATTAAGGACGACAGCCTCGAAAACAAGGCGCTGGT GTACCGCGAGGAGAATCAGCACAAGACCGACCAGCTTTTGATTGATA AGAGCATTCGCTGGAAGATGGGCGAGACGTGGAAACAGTTGCATCA GGCGAAGCTCATTGGCGATAAAAAGTATCGCAACCTGCTGAGGTCTT CCATAAACGAGAATGCCATGAAGGGCTTTGTCGCGCGCCAGCTCGTG GAGACGAGCCAGATGGTCAAGATGGCGCAGGCGTTGCTTGAGGCGC GTTATGCCGACGAGGGAACGAAGATCGTTCCCGTTAAGGCGAGCGT ATCGCACAACCTTCGTGAGGCAGCCGGCTTGGTCAAATGCCGCGAA GCCAATGATTTCCATCATGCGCACGATGCCTATCTGGCATGTCGCTT GGGCCTGTTTATTCAGATGCGCCACCCCGGAATGTATGAAAACCCCA TTGGCTACTCGCACGTGATTAAGAAGTACGTGCGCGAGCAGTCTGAG CTGTTTAAGAAAACCCATCGCATGCCTGGATCTGCCGGTTTTGTCGTC AATAGCTTTATGACTGCGGGTTTTGACCCCGAGACGGGCGAGGTCTT TGACGATGCCTGGACCGAGGACGACGAGGTCAAGCGTTATAGGGGA AAGTATCTTCCGGACGGCTGGAATGCAGCAGCGGAGGTTGAGAGCA TTCGCCGTGCGCTTAACTATCGCCAGTGCTACATCTCGAGGATGCCG CAGGAGGATACGGGAGCGTTCTGGAAGGCAACGATTTACTCGCCGC GTGATCCCAAGATGGGTGCAAAGCTTGAGTTGTCGACCAAGCAGGG GCTTGATGCTAAGCGGTACGGTGGTTTTTCGAGCCAGCAGTTTGCGT ATTTCTTTATCTATGAGGCCAAAAAGAAGGGGAAGCCGGTGTTCCGG TTCTCCCAGGTTCCCGTGTGGCTTGCGGCGCGCATCGAGGGCGACT CGGCGGCACTCGGCGAATATGCAAGGGGACTTGCGGAGAAGGAAGG TTTGGAATACGTCGGTATCACGAGGCCGAAGATCTACAAGAAACAGC TGATTGAGGTGGATGGCGAGAGATTCTTTATTACGGGGAAAAAGGAG ATGAGGAATGCTACGCAACTGGCTTTTTCAAATGAGCAAATGAAACTG CTCGCAGGAGATACGTGTAGCGAAGATGCTATCAAGCTTTTGGATAT GTTTAATCCCGCGGCTTCTTCCGTCTCTGGCAGGCTGCTGCGTCAAC TCAAAATTGAGGCGATCATTGAAAAGTGCGCAACTGTGGATGCGGAA GATGTTGTTGCAGTAGTCAAGCGCCTGCTTGCCTTGGTGAATGGAAC AATGAATATGGTTGATTTGTCACCCGCGGGCGGCTCAAAGTTTTCGG GTTGTATACAGCCTAATTATTCAAAGTTGCTGAGCGATTCAAATGTGA CGGTTTTCCTTATTGATCAGTCCGTAACAGGCATGTTCGAGAGAAGG ACCCGTGTCGGGCTTTAG CFDE Type II AACCTGAGAAACTACAGCGGCGACTACAACATCGGACTCGACATGGG 59 Cas coding CACCGGCAGCGTGGGCTGGGCCGTGACAGACGCCCAAGGTAAGTTG sequence (nt) CTGCATTTCAAGAAACAACCTACCTGGGGCTCCAGACTCTTTGACAG (human CGCCCAGCCTGCCAGCGAGGCGAGAGTGCACCGGGGACAGAGAAG codon- AAGATACGTGCGTAGAAGATGGCGGTTGGACCTGCTGCAGAAGCTCT optimized; TTGAACCTGCCATGAGCGAGATCGACCCTGGCTTCTTCATGCGGCTG lacking N- AACCAGAGCAGAACTATCGAGGGAGATCCTATCTTCACCAAGGACTT terminal TACCAAAGCCGACTACTACGACAGATTCCCTACCATCTATCACCTGCG methionine; GGCCTATCTGATGGACACCGATGAGCAGGCTGATCTGCGGCTGGTG no stop TACTTAGCCATCCACAACATCGTCAAACACCGGGGCAACTTCCTGCG codon) CCAAGGGGAGAAGTTGACCGCCAAGACCGCCAAAACCAGCGACGCC CTGGAGCGTCTGAATGATTCGCTGCAAGGATGGTGCGAAAGCAGAG TABLE 1K CFDE Type II Cas Sequences Name Sequence SEQ ID NO AGTACGAGGTTGCCAAGATCAACAAGTCGGCCGTGGCGAAGATCCT GGCGGATGAACGCAGCAGCCGGTCCCAGAAAGCCAAGGACATCCAG GCCCTGGTGAAGGTGACGACTGGCGACGCCGCCGCGGATAAGAAGC TGGCTAAGGGCCTCTCTAACGCTGTGGTGGGCCTCCAGTGTGAGTTC AAGGACGTGTTCGGGGAGTTCGAGTGCGAAGCCACCAAGCTGGCCT TGAGCGACGATGAGAAACTGGAAACACTCCAGGCTGCATGCCCTGAC GATTGCGTGGAGCTGTTGGAGAGCGTTTGTGGCGTGTACAGCGCCT ATGTCCTGCAAGGCCTGCTGAGCTACGCCGAAGGCCAGACCATCAG TATCAACATGATCAAGAAGTACGAGCAATACGCCGAAGATCTGGCCC TGCTGAAGTTGCTGGTGAGAAAGTATGCTCTGGACAGCTACGAGCCC TTCTTCCGGGGAGCCACGTACCACGATAGCAACAGCGACGACCCTTC CAGGGACTACGACGCCAGCAAGGCTACAGGCTACACCGCCTACGAC CTGCACAAGCTCAGCTATGACGACTTTGCCAAGTCCGTGAAGAAACT GTTTGCTGGAACCGGTGCCGAAACCGACGAGAGATATATCGCCATGA TGGACGCTTTCGACAAACAGAGATTCCTAAGACGGCTGAAAACCTCC GATAACGGCGCCATCTACTACCAGCTTCATCTGGAGGAACTGCAGGC CATCCTGAAAAACCAGGGCAGATTCTACCCTTTTCTGAAGGACGAGG CCGCTAAGATCGAGAGCCTGGTGACCTTCAGAATCCCTTACTACGTG GGCCCCCTGACAAGCAAGAACGCCGCCCAGGATAAGCACGGCAAGC CCAGATTTCAGTGGTCTGAGCGGAAGCCTGGCATGGAAGATGCAACA ATCACCCCTTGGAACTGGGACGCCGTTATCGACAAGAACAAGAGCGC TGAAAATTTCATCCTGCGCATGACAGGCGACTGCACCTACCTGGCTG GAGAAAAGGTGCTGCCTAAGCAGAGCCTGCTGTACGAGGAATTTTGC GTGCTGAACGAGCTGAATGGCGTGCGGTGGAGCGAGGACGGCGAC AACTGGCAGCGGCTGGACGCCGCCCAGCGGGAAGGCATTATCCATG AGCTGTTCCACAAGAAGAGAAGAGTGACCTACAAGATGATCGCCGAC TGGCTGGTGAAGGAGGGCGATGCCACCAACCCCGTGGTGCGGGGC GGCCAACGCGAGAGCGGCCTGGAAAGCAAGATGGGCTCTTACATCT TTTTCGCTAAAGATATCTTCGGCGTGGATGAGCTGAGCAGGGCCGAG TACCCCGTGATCGAGAAGATCATCCTGTGGAATACCCTGTTCGAAGA TAGATCTATCCTTAAAGAAAAGCTGCAGGACGAGTTCGGCCCTAGCG GAAATGGTATGCTGGACGCCGAGCAGATTAAGAAGATTTGTAAAAAA CGGCTGACCGGCTGGGGCAGACTGAGCGAAAAGTTCCTGACAGGTA TCAGAATCGACACCCAGGCCGGCCGGTCTATGTCTATCATGGACGTG CTGCGCGAAGGCAACCCTTCTTCCGAACACCGGCTGGGCGAGACAA TGGTGATGATGGAAATCCTGCATGACAACACACTGGGCTTTCAGGAG AAAGTCGATGAACACAACAGAAAGTACTACGCCGAGCACGAAAAGTC TCTGGGCGTCAACGAGCTGCCTGGTTCTCCTGCCATCAGGAGATCTC TGAATCAGGCGATCAGAATAGTGGACGAGATCGCCAAGATTGCCGGC CACGCTCCTGCCAATGTGTTCGTGGAAGTGACCAGAGATGAAGATGA GAAGAAGAAGGGCAAGCGGACAACAAAAAGATGGGATGCCATTGAG GATGCCCTGAAGGCATTTAAGGCCGAAGGCGGCGACCTGGCAGTGG TGGACGACTTCAAGGAACTGAGAGCCGCCAACGTGGACCTGGATGA AAGACTGACACTGTACCTGATGCAAAACGGCAAGTGTCTGTACTCCG GCAGAGCTATCGACCTGAACAAGCTGATGGCCGGCTCTGGCGAGTA CGAGGTGGACCACATCATCCCAAGAGCTTACATCAAGGACGATAGCC TGGAAAATAAAGCCCTGGTGTACCGGGAGGAGAACCAGCACAAGAC AGACCAGCTACTGATCGATAAGTCTATAAGATGGAAGATGGGAGAAA CATGGAAGCAGCTGCACCAGGCTAAACTGATCGGGGATAAGAAATAC AGAAACCTGCTGAGATCTTCCATCAACGAGAATGCAATGAAGGGCTT CGTTGCAAGACAACTCGTGGAAACCAGCCAGATGGTCAAGATGGCCC AGGCCCTGCTGGAAGCCAGATACGCCGACGAGGGCACAAAGATCGT GCCTGTGAAGGCCAGCGTGTCCCACAATCTGAGAGAAGCTGCCGGC CTGGTGAAGTGCAGAGAAGCAAACGACTTCCACCACGCCCACGACG CCTATCTGGCCTGCAGACTGGGCCTGTTCATCCAAATGAGACACCCC GGCATGTATGAGAACCCAATCGGCTACAGCCACGTGATCAAGAAGTA CGTGAGAGAGCAGTCTGAACTGTTCAAGAAAACCCACCGAATGCCCG TABLE 1K CFDE Type II Cas Sequences Name Sequence SEQ ID NO GCAGCGCCGGATTCGTGGTCAACAGCTTCATGACCGCTGGCTTCGA CCCAGAAACCGGCGAGGTGTTTGACGACGCTTGGACCGAGGATGAT GAAGTTAAACGGTATAGAGGAAAGTACCTGCCTGACGGCTGGAATGC CGCCGCAGAGGTGGAAAGCATTCGTAGAGCCCTGAACTACCGGCAG TGCTACATCAGCAGAATGCCTCAGGAGGATACCGGAGCCTTCTGGAA GGCCACAATCTACAGCCCCCGGGACCCTAAGATGGGCGCTAAGTTA GAACTGAGCACCAAGCAGGGACTGGATGCTAAAAGATACGGCGGATT CAGCAGCCAGCAGTTCGCCTACTTCTTCATCTACGAGGCCAAAAAGA AAGGCAAACCCGTGTTCAGATTCAGCCAGGTGCCAGTGTGGCTGGCT GCCAGAATCGAGGGCGATTCCGCCGCTCTGGGAGAATACGCCAGGG GCCTGGCCGAGAAGGAGGGCCTGGAGTACGTGGGCATCACCCGGC CTAAGATCTACAAGAAGCAGCTGATAGAAGTGGACGGCGAGAGATTC TTCATCACCGGCAAGAAGGAAATGAGAAACGCCACCCAGCTGGCCTT CAGCAACGAGCAGATGAAGCTGCTGGCCGGCGACACCTGCAGCGAG GATGCCATCAAGCTGTTAGACATGTTCAACCCTGCAGCCTCCAGCGT GTCTGGCAGACTGCTGAGGCAGCTCAAGATCGAGGCCATTATTGAGA AGTGCGCCACCGTGGACGCCGAGGACGTGGTCGCCGTAGTGAAGC GGCTGCTGGCTCTGGTGAACGGCACCATGAACATGGTGGACCTCAG TCCTGCAGGAGGCAGTAAGTTTTCCGGATGTATCCAGCCCAACTACT CAAAGCTGCTGAGCGACTCCAATGTCACCGTGTTTCTGATCGATCAG AGCGTGACAGGAATGTTCGAGAGAAGAACAAGAGTGGGCCTG CFDE Type II ATGGGCAAACCTATCCCCAATCCCCTGCTGGGCCTGGACAGCACCAA 60 Cas GCGGACTGCCGACGGCAGTGAGTTCGAGTCTCCCAAGAAAAAGAGG mammalian AAAGTGGGATCCAACCTGAGAAACTACAGCGGCGACTACAACATCGG expression ACTCGACATGGGCACCGGCAGCGTGGGCTGGGCCGTGACAGACGC construct CCAAGGTAAGTTGCTGCATTTCAAGAAACAACCTACCTGGGGCTCCA (with N- GACTCTTTGACAGCGCCCAGCCTGCCAGCGAGGCGAGAGTGCACCG terminal GGGACAGAGAAGAAGATACGTGCGTAGAAGATGGCGGTTGGACCTG methionine, CTGCAGAAGCTCTTTGAACCTGCCATGAGCGAGATCGACCCTGGCTT stop codon, CTTCATGCGGCTGAACCAGAGCAGAACTATCGAGGGAGATCCTATCT V5 tag, N- TCACCAAGGACTTTACCAAAGCCGACTACTACGACAGATTCCCTACCA terminal NLS TCTATCACCTGCGGGCCTATCTGATGGACACCGATGAGCAGGCTGAT and C- CTGCGGCTGGTGTACTTAGCCATCCACAACATCGTCAAACACCGGGG terminal NLS) CAACTTCCTGCGCCAAGGGGAGAAGTTGACCGCCAAGACCGCCAAA (nt) ACCAGCGACGCCCTGGAGCGTCTGAATGATTCGCTGCAAGGATGGT GCGAAAGCAGAGAGTACGAGGTTGCCAAGATCAACAAGTCGGCCGT GGCGAAGATCCTGGCGGATGAACGCAGCAGCCGGTCCCAGAAAGCC AAGGACATCCAGGCCCTGGTGAAGGTGACGACTGGCGACGCCGCCG CGGATAAGAAGCTGGCTAAGGGCCTCTCTAACGCTGTGGTGGGCCT CCAGTGTGAGTTCAAGGACGTGTTCGGGGAGTTCGAGTGCGAAGCC ACCAAGCTGGCCTTGAGCGACGATGAGAAACTGGAAACACTCCAGG CTGCATGCCCTGACGATTGCGTGGAGCTGTTGGAGAGCGTTTGTGG CGTGTACAGCGCCTATGTCCTGCAAGGCCTGCTGAGCTACGCCGAA GGCCAGACCATCAGTATCAACATGATCAAGAAGTACGAGCAATACGC CGAAGATCTGGCCCTGCTGAAGTTGCTGGTGAGAAAGTATGCTCTGG ACAGCTACGAGCCCTTCTTCCGGGGAGCCACGTACCACGATAGCAAC AGCGACGACCCTTCCAGGGACTACGACGCCAGCAAGGCTACAGGCT ACACCGCCTACGACCTGCACAAGCTCAGCTATGACGACTTTGCCAAG TCCGTGAAGAAACTGTTTGCTGGAACCGGTGCCGAAACCGACGAGA GATATATCGCCATGATGGACGCTTTCGACAAACAGAGATTCCTAAGAC GGCTGAAAACCTCCGATAACGGCGCCATCTACTACCAGCTTCATCTG GAGGAACTGCAGGCCATCCTGAAAAACCAGGGCAGATTCTACCCTTT TCTGAAGGACGAGGCCGCTAAGATCGAGAGCCTGGTGACCTTCAGA ATCCCTTACTACGTGGGCCCCCTGACAAGCAAGAACGCCGCCCAGG ATAAGCACGGCAAGCCCAGATTTCAGTGGTCTGAGCGGAAGCCTGG CATGGAAGATGCAACAATCACCCCTTGGAACTGGGACGCCGTTATCG ACAAGAACAAGAGCGCTGAAAATTTCATCCTGCGCATGACAGGCGAC TABLE 1K CFDE Type II Cas Sequences Name Sequence SEQ ID NO TGCACCTACCTGGCTGGAGAAAAGGTGCTGCCTAAGCAGAGCCTGCT GTACGAGGAATTTTGCGTGCTGAACGAGCTGAATGGCGTGCGGTGG AGCGAGGACGGCGACAACTGGCAGCGGCTGGACGCCGCCCAGCGG GAAGGCATTATCCATGAGCTGTTCCACAAGAAGAGAAGAGTGACCTA CAAGATGATCGCCGACTGGCTGGTGAAGGAGGGCGATGCCACCAAC CCCGTGGTGCGGGGCGGCCAACGCGAGAGCGGCCTGGAAAGCAAG ATGGGCTCTTACATCTTTTTCGCTAAAGATATCTTCGGCGTGGATGAG CTGAGCAGGGCCGAGTACCCCGTGATCGAGAAGATCATCCTGTGGA ATACCCTGTTCGAAGATAGATCTATCCTTAAAGAAAAGCTGCAGGACG AGTTCGGCCCTAGCGGAAATGGTATGCTGGACGCCGAGCAGATTAA GAAGATTTGTAAAAAACGGCTGACCGGCTGGGGCAGACTGAGCGAA AAGTTCCTGACAGGTATCAGAATCGACACCCAGGCCGGCCGGTCTAT GTCTATCATGGACGTGCTGCGCGAAGGCAACCCTTCTTCCGAACACC GGCTGGGCGAGACAATGGTGATGATGGAAATCCTGCATGACAACACA CTGGGCTTTCAGGAGAAAGTCGATGAACACAACAGAAAGTACTACGC CGAGCACGAAAAGTCTCTGGGCGTCAACGAGCTGCCTGGTTCTCCTG CCATCAGGAGATCTCTGAATCAGGCGATCAGAATAGTGGACGAGATC GCCAAGATTGCCGGCCACGCTCCTGCCAATGTGTTCGTGGAAGTGAC CAGAGATGAAGATGAGAAGAAGAAGGGCAAGCGGACAACAAAAAGAT GGGATGCCATTGAGGATGCCCTGAAGGCATTTAAGGCCGAAGGCGG CGACCTGGCAGTGGTGGACGACTTCAAGGAACTGAGAGCCGCCAAC GTGGACCTGGATGAAAGACTGACACTGTACCTGATGCAAAACGGCAA GTGTCTGTACTCCGGCAGAGCTATCGACCTGAACAAGCTGATGGCCG GCTCTGGCGAGTACGAGGTGGACCACATCATCCCAAGAGCTTACATC AAGGACGATAGCCTGGAAAATAAAGCCCTGGTGTACCGGGAGGAGA ACCAGCACAAGACAGACCAGCTACTGATCGATAAGTCTATAAGATGG AAGATGGGAGAAACATGGAAGCAGCTGCACCAGGCTAAACTGATCG GGGATAAGAAATACAGAAACCTGCTGAGATCTTCCATCAACGAGAAT GCAATGAAGGGCTTCGTTGCAAGACAACTCGTGGAAACCAGCCAGAT GGTCAAGATGGCCCAGGCCCTGCTGGAAGCCAGATACGCCGACGAG GGCACAAAGATCGTGCCTGTGAAGGCCAGCGTGTCCCACAATCTGA GAGAAGCTGCCGGCCTGGTGAAGTGCAGAGAAGCAAACGACTTCCA CCACGCCCACGACGCCTATCTGGCCTGCAGACTGGGCCTGTTCATC CAAATGAGACACCCCGGCATGTATGAGAACCCAATCGGCTACAGCCA CGTGATCAAGAAGTACGTGAGAGAGCAGTCTGAACTGTTCAAGAAAA CCCACCGAATGCCCGGCAGCGCCGGATTCGTGGTCAACAGCTTCAT GACCGCTGGCTTCGACCCAGAAACCGGCGAGGTGTTTGACGACGCT TGGACCGAGGATGATGAAGTTAAACGGTATAGAGGAAAGTACCTGCC TGACGGCTGGAATGCCGCCGCAGAGGTGGAAAGCATTCGTAGAGCC CTGAACTACCGGCAGTGCTACATCAGCAGAATGCCTCAGGAGGATAC CGGAGCCTTCTGGAAGGCCACAATCTACAGCCCCCGGGACCCTAAG ATGGGCGCTAAGTTAGAACTGAGCACCAAGCAGGGACTGGATGCTAA AAGATACGGCGGATTCAGCAGCCAGCAGTTCGCCTACTTCTTCATCT ACGAGGCCAAAAAGAAAGGCAAACCCGTGTTCAGATTCAGCCAGGTG CCAGTGTGGCTGGCTGCCAGAATCGAGGGCGATTCCGCCGCTCTGG GAGAATACGCCAGGGGCCTGGCCGAGAAGGAGGGCCTGGAGTACG TGGGCATCACCCGGCCTAAGATCTACAAGAAGCAGCTGATAGAAGTG GACGGCGAGAGATTCTTCATCACCGGCAAGAAGGAAATGAGAAACGC CACCCAGCTGGCCTTCAGCAACGAGCAGATGAAGCTGCTGGCCGGC GACACCTGCAGCGAGGATGCCATCAAGCTGTTAGACATGTTCAACCC TGCAGCCTCCAGCGTGTCTGGCAGACTGCTGAGGCAGCTCAAGATC GAGGCCATTATTGAGAAGTGCGCCACCGTGGACGCCGAGGACGTGG TCGCCGTAGTGAAGCGGCTGCTGGCTCTGGTGAACGGCACCATGAA CATGGTGGACCTCAGTCCTGCAGGAGGCAGTAAGTTTTCCGGATGTA TCCAGCCCAACTACTCAAAGCTGCTGAGCGACTCCAATGTCACCGTG TTTCTGATCGATCAGAGCGTGACAGGAATGTTCGAGAGAAGAACAAG TABLE 1K CFDE Type II Cas Sequences Name Sequence SEQ ID NO AGTGGGCCTGTCTAGAAAGCGGACAGCAGACGGCTCCGAATTTGAAA GCCCTAAGAAAAAGAGAAAGGTGTGA [0103] In some embodiments a CFDE Type II Cas protein comprises an amino acid sequence of SEQ ID NO:55, SEQ ID NO:56, or SEQ ID NO:57. In some embodiments, a CFDE Type II Cas protein has nickase activity, for example resulting from one or more amino acid substitutions relative to the sequence of SEQ ID NO:55, SEQ ID NO:56, or SEQ ID NO:57. In some embodiments, the one or more amino acid substitutions providing nickase activity comprise a D15A substitution, wherein the position of the D15A substitution is defined with respect to the amino acid numbering of SEQ ID NO:56. In some embodiments, the one or more amino acid substitutions providing nickase activity comprise an H890A substitution, wherein the position of the H890A substitution is defined with respect to the amino acid numbering of SEQ ID NO:56. In some embodiments, a CFDE Type II Cas protein is catalytically inactive, for example due to a D15A substitution in combination with an H890A substitution. 6.2.2. Fusion and Chimeric Proteins [0104] The disclosure provides Type II Cas proteins (e.g., a DGHJ Type II Cas protein as described in Section 6.2.1.1, a CUAZ Type II Cas protein as described in Section 6.2.1.2, an AHWY Type II Cas protein as described in Section 6.2.1.3, a CBGI Type II Cas protein as described in Section 6.2.1.4, an ASDR Type II Cas protein as described in Section 6.2.1.5, a BCZZ Type II Cas protein as described in Section 6.2.1.6, a DRCY Type II Cas protein as described in Section 6.2.1.7, a DRPY Type II Cas protein as described in Section 6.2.1.8, an EAJR Type II Cas protein as described in Section 6.2.1.9, or a CFDE Type II Cas protein as described in Section 6.2.1.10) which are in the form of fusion proteins comprising a Type II Cas protein sequence fused with one or more additional amino acid sequences, such as one or more nuclear localization signals and/or one or more non-native tags. Fusion proteins can also comprise an amino acid sequence of, for example, a nucleoside deaminase, a reverse transcriptase, a transcriptional activator (e.g., VP64), a transcriptional repressor (e.g., Krüppel associated box (KRAB)), a histone-modifying protein, an integrase, or a recombinase. [0105] In some embodiments, a fusion protein of the disclosure comprises a means for localizing the Type II Cas protein to the nucleus, for example a nuclear localization signal. [0106] Non-limiting examples of nuclear localization signals include KRTADGSEFESPKKKRKV (SEQ ID NO:116), PKKKRKV (SEQ ID NO:117), PKKKRRV (SEQ ID NO:118), KRPAATKKAGQAKKKK (SEQ ID NO:119), YGRKKRRQRRR (SEQ ID NO:120), RKKRRQRRR (SEQ ID NO:121), PAAKRVKLD (SEQ ID NO:122), RQRRNELKRSP (SEQ ID NO:123), VSRKRPRP (SEQ ID NO:124), PPKKARED (SEQ ID NO:125), PQPKKKPL (SEQ ID NO:126), SALIKKKKKMAP (SEQ ID NO:127), PKQKKRK (SEQ ID NO:128), RKLKKKIKKL (SEQ ID NO:129), REKKKFLKRR (SEQ ID NO:130), KRKGDEVDGVDEVAKKKSKK (SEQ ID NO:131), RKCLQAGMNLEARKTKK (SEQ ID NO:132), NQSSNFGPMKGGNFGGRSSGPYGGGGQYFAKPRNQGGY (SEQ ID NO:133), RMRIZFKNKGKDTAELRRRRVEVSVELRKAKKDEQILKRRNV (SEQ ID NO:134) PAAKKKKLD (SEQ ID NO:582) and KRPAATKKAGQAKKKKLD (SEQ ID NO:583). In some embodiments, a nuclear localization signal comprises. PAAKKKKLD (SEQ ID NO 582) In some embodiments, a nuclear localization signal comprises KRPAATKKAGQAKKKKLD (SEQ ID NO:583) [0107] Exemplary fusion partners include protein tags (e.g., V5-tag (e.g., having the sequence GKPIPNPLLGLDST (SEQ ID NO:135) or IPNPLLGLD (SEQ ID NO:136)), FLAG-tag, myc-tag, HA-tag, GST-tag, polyHis-tag, MBP-tag), protein domains, transcription modulators, enzymes acting on small molecule substrates, DNA, RNA and protein modification enzymes (e.g., adenosine deaminase, cytidine deaminase, guanosyl transferase, DNA methyltransferase, RNA methyltransferases, DNA demethylases, RNA demethylases, dioxygenases, polyadenylate polymerases, pseudouridine synthases, acetyltransferases, deacetylase, ubiquitin-ligases, deubiquitinases, kinases, phosphatases, NEDD8- ligases, de-NEDDylases, SUMO-ligases, deSUMOylases, histone deacetylases, reverse transcriptases, histone acetyltransferases histone methyltransferases, histone demethylases), protein DNA binding domains, RNA binding proteins, polypeptide sequences with specific biological functions (e.g., nuclear localization signals, mitochondrial localization signals, plastid localization signals, subcellular localization signals, destabilizing signals, Geminin destruction box motifs), and biological tethering domains (e.g., MS2, Csy4 and lambda N protein). Various Type II Cas fusion proteins are described in Ribeiro et al., 2018, In. J. Genomics, Article ID:1652567; Jayavaradhan, et al., 2019, Nat Commun 10:2866; Xiao et al., 2019, The CRISPR Journal, 2(1):51-63; Mali et al., 2013, Nat Methods.10(10):957-63; US patent nos. 9,322,037, and 9,388,430. In some embodiments, a fusion partner is an adenosine deaminase. An exemplary adenosine deaminase is the tRNA adenosine deaminase (TadA) moiety contained in the adenine base editor ABE8e (Richter, 2020, Nature Biotechnology 38:883-891). The TadA moiety of ABE8e comprises the following amino acid sequence: SEVEFSHEYWMRHALTLAKRARDEREVPVGAVLVLNNRVIGEGWNRAIGLHDPTAHAEIMALRQGGLV MQNYRLIDATLYVTFEPCVMCAGAMIHSRIGRVVFGVRNSKRGAAGSLMNVLNYPGMNHRVEITEGILAD ECAALLCDFYRMPRQVFNAQKKAQSSIN (SEQ ID NO:137) [0108] In some embodiments, an adenosine deaminase fusion partner comprises an amino acid sequence having at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, at least 95%, at least 97%, at least 98%, at least 99% amino acid sequence identity with SEQ ID NO:137. [0109] Type II Cas proteins of the disclosure in the form of a fusion protein comprising an adenosine GHDPLQDVH^FDQ^EH^XVHG^DV^DQ^DGHQLQH^EDVH^HGLWRU^WR^FKDQJH^DQ^³$´^WR^D^³*´^LQ^'1$^^7\SH^,,^&DV^ proteins of the disclosure in the form of a fusion protein comprising a cytidine deaminase can be used as a F\WRVLQH^EDVH^HGLWRU^WR^FKDQJH^D^³&´^WR^D^³7´^LQ^'1$^ [0110] In some embodiments, a fusion protein of the disclosure comprises a means for deaminating adenosine, for example an adenosine deaminase, e.g., a TadA variant. In some embodiments, a fusion protein of the disclosure comprises a means for deaminating cytidine, for example a cytidine deaminase, e.g., cytidine deaminase 1 (CDA1) or an apolipoprotein B mRNA-editing complex (APOBEC) family deaminase (Cheng et al., 2019, Nat Commun.10(1):3612; Gehrke et al., 2018, Nat Biotechnol. 36(10):977-982). [0111] In some embodiments, a fusion protein of the disclosure comprises a means for synthesizing DNA from a single-stranded template, for example a reverse transcriptase. Type II Cas proteins of the disclosure in the form of a fusion protein comprising a reverse transcriptase (RT) can be used as a prime editor to carry out precise base editing without double-stranded DNA breaks. [0112] In some embodiments, a fusion protein of the disclosure is a prime editor, e.g., a Type II Cas protein fused to a suitable RT (e.g., Moloney murine leukemia virus (M-MLV) RT or other RT enzyme). Such fusion proteins can be used in conjunction with a prime editing guide RNA (pegRNA) that both specifies the target site and encodes the desired edit (Anzalone et al., 2019, Nature, 576(7785):149- 157). [0113] In some embodiments, a fusion protein of the disclosure comprises one or more nuclear localization signals positioned N-terminal and/or C-terminal to a Type II Cas protein sequence (e.g., a DGHJ Type II Cas protein having a sequence of SEQ ID NO:1). In some embodiments, a fusion protein of the disclosure comprises an N-terminal and a C-terminal nuclear localization signal, for example each having the sequence KRTADGSEFESPKKKRKV (SEQ ID NO:116). [0114] The disclosure provides chimeric Type II Cas proteins comprising one or more domains of a DGHJ Type II Cas protein and one or more domains of one or more different proteins (e.g., one or more different Type II Cas proteins), chimeric Type II Cas proteins comprising one or more domains of a CUAZ Type II Cas protein and one or more domains of one or more different proteins (e.g., one or more different Type II Cas proteins), chimeric Type II Cas proteins comprising one or more domains of an AHWY Type II Cas protein and one or more domains of one or more different proteins (e.g., one or more different Type II Cas proteins), chimeric Type II Cas proteins comprising one or more domains of a CBGI Type II Cas protein and one or more domains of one or more different proteins (e.g., one or more different Type II Cas proteins), chimeric Type II Cas proteins comprising one or more domains of an ASDR Type II Cas protein and one or more domains of one or more different proteins (e.g., one or more different Type II Cas proteins), chimeric Type II Cas proteins comprising one or more domains of a BCZZ Type II Cas protein and one or more domains of one or more different proteins (e.g., one or more different Type II Cas proteins), chimeric Type II Cas proteins comprising one or more domains of a DRCY Type II Cas protein and one or more domains of one or more different proteins (e.g., one or more different Type II Cas proteins), chimeric Type II Cas proteins comprising one or more domains of a DRPY Type II Cas protein and one or more domains of one or more different proteins (e.g., one or more different Type II Cas proteins), chimeric Type II Cas proteins comprising one or more domains of an EAJR Type II Cas protein and one or more domains of one or more different proteins (e.g., one or more different Type II Cas proteins), and chimeric Type II Cas proteins comprising one or more domains of a CFDE Type II Cas protein and one or more domains of one or more different proteins (e.g., one or more different Type II Cas proteins). [0115] The domain boundaries of wild-type Type II Cas proteins of the disclosure were determined by multiple alignment with the amino acid sequences of Type II Cas proteins for which the crystal structure is known and for which it is thus possible to define the boundaries of each functional domain. Domains identified in Type II Cas proteins include: the RuvC catalytic domain (discontinuous, represented by RuvC-I, RuvC-II, and RuvC-III domains), bridge helix (BH), recognition (REC) domain (which can be divided into REC1_1, REC2, and REC1_2 domains), HNH catalytic domain, and PAM-interacting domain (PID). The PID domain can be divided into a topoisomerase homology (Topo) domain and a C-terminal domain (CTD). [0116] Table 2 below report the amino acid positions corresponding to the boundaries between different functional domains in wild-type DGHJ (SEQ ID NO:2), CUAZ (SEQ ID NO:8), AHWY (SEQ ID NO:14), CBGI (SEQ ID NO:20), ASDR (SEQ ID NO:26), BCZZ (SEQ ID NO:32), DRCY (SEQ ID NO:38), DRPY (SEQ ID NO:44), EAJR (SEQ ID NO:50), and CFDE (SEQ ID NO:56) Type II Cas proteins. Table 2 Amino Acid Positions of DGHJ, CUAZ, AHWY CBGI, ASDR, BCZZ, DRCY, DRPY, EAJR, and CFDE Domains D^{GHJ CUAZ AHWY CBGI ASDR BCZZ DRCY DRPY EAJR CFDE} R_uvC-I^{1-49 1-50 1-51 1-51 1-52 1-54 1-52 1-51 1-55 1-52} B_H^{50-83 51-79 52-85 53-86 53-86 55-88 53-86 52-85 56-89 53-86} 84- 80- 86-178 87- 87- 89- 87- 86- 90- 87- REC1_1 176 153 178 167 180 178 171 179 167 177- 154- 179- 179- 168- 181- 179- 172- 180- 168- REC2 303 285 322 309 303 307 316 315 313 303 304- 286- 323- 310- 304- 308- 317- 316- 314- 304- REC1_2 738 699 769 765 765 706 765 741 727 757 739- 700- 770- 766- 766- 707- 766- 742- 728- 758- RuvC-II 794 764 825 821 822 762 822 798 784 814 795- 765- 826- 822- 823- 763- 823- 799- 785- 815- HNH 934 924 966 956 964 902 959 945 924 956 935- 925- 967- 957- 965- 903- 960- 946- 925- 957- RuvC-III 1075 1102 1130 1121 1149 1043 1120 1082 1058 1141 1076- 1103- 1131- 1122- 1150- 1044- 1121- 1083- 1059- 1142- Topo 1176 1205 1234 1221 1250 1143 1226 1191 1151 1246 1177- 1206- 1235- 1222- 1251- 1144- 1227- 1192- 1152- 1247- CTD 1344 1408 1392 1374 1404 1311 1388 1377 1324 1396 [0117] A chimeric Type II Cas protein can comprise one of more of the following domains (e.g., one or more, two or more, three or more, four or more, five or more, six or more, seven or more) from a CUAZ Type II Cas protein, AHWY Type II Cas protein, DGHJ Type II Cas protein, CBGI Type II Cas protein, ASDR Type II Cas protein, BCZZ Type II Cas protein, DRCY Type II Cas protein, DRPY Type II Cas protein, EAJR Type II Cas protein, and/or CFDE Type II Cas protein, and one or more domains from one or more other proteins, for example SaCas9, SpCas9 or a Type II Cas protein described in US 2020/0332273, US 2019/0169648, or 2015/0247150 (the contents of each of which are incorporated herein by reference in their entirety): RuvC-I, BH, REC (or a subpart thereof, for example one, two, or three of REC1_1, REC2, and REC1_2), RuvC-II, HNH, RuvC-III, PID (or a subpart thereof, for example one or both of the Topo domain and CTD). For example, the PID domain can be swapped between different Type II Cas proteins to change the PAM specificity of the resulting chimeric protein (which is given by the donor PID domain). Swapping of other domains or portions of them is also within the scope of the disclosure (e.g., through protein shuffling). [0118] In some embodiments, a Type II Cas protein of the disclosure comprises one, two, three, four, five, six, seven, eight, nine or ten of a RuvC-I domain, a BH domain, a REC1_1 domain, a REC2 domain, a REC1_2 domain, a RuvC-II domain, a HNH domain, a RuvC-III domain, a Topo domain and a CTD arranged in the N-terminal to C-terminal direction. In some embodiments, all domains are from a DGHJ Type II Cas protein (e.g., a DGHJ Type II Cas protein whose amino acid sequence comprises SEQ ID NO:1, 2, or 3). In some embodiments, all domains are from a CUAZ Type II Cas protein (e.g., a CUAZ Type II Cas protein whose amino acid sequence comprises SEQ ID NO:7, 8, or 9). In some embodiments, all domains are from an AHWY Type II Cas protein (e.g., an AHWY Type II Cas protein whose amino acid sequence comprises SEQ ID NO:13, 14, or 15). In some embodiments, all domains are from a CBGI Type II Cas protein (e.g., a CBGI Type II Cas protein whose amino acid sequence comprises SEQ ID NO:19, 20, or 21). In some embodiments, all domains are from an ASDR Type II Cas protein (e.g., an ASDR Type II Cas protein whose amino acid sequence comprises SEQ ID NO:25, 26, or 27). In some embodiments, all domains are from a BCZZ Type II Cas protein (e.g., a BCZZ Type II Cas protein whose amino acid sequence comprises SEQ ID NO:31, 32, or 33). In some embodiments, all domains are from a DRCY Type II Cas protein (e.g., a DRCY Type II Cas protein whose amino acid sequence comprises SEQ ID NO:37, 38, or 39). In some embodiments, all domains are from a DRPY Type II Cas protein (e.g., a DRPY Type II Cas protein whose amino acid sequence comprises SEQ ID NO:43, 44, or 45). In some embodiments, all domains are from an EAJR Type II Cas protein (e.g., an EAJR Type II Cas protein whose amino acid sequence comprises SEQ ID NO:49, 50, or 51). In some embodiments, all domains are from a CFDE Type II Cas protein (e.g., a CFDE Type II Cas protein whose amino acid sequence comprises SEQ ID NO:55, 56, or 57). In other embodiments, one or more domains (e.g., one domain), e.g., a PID domain, is from another Type II Cas protein. [0119] In addition, one or more amino acid substitutions can be introduced in one or more domains to modify the properties of the resulting nuclease in terms of editing activity, targeting specificity or PAM recognition specificity. For example, one or more amino acid substitutions can be introduced to provide nickase activity. Exemplary amino acid substitutions in SpCas9 providing nickase activity are the D10A substitution in the RuvC domain and the H840A substitution in the HNH domain. Combining both the D10A and H840A substitutions in SpCas9 provides a catalytically inactive nuclease. Corresponding substitutions can be introduced into the Type II Cas nucleases of the disclosure to provide nickases and catalytically inactive Cas proteins. For example, a DGHJ Type II Cas protein can include a D12A substitution (corresponding to D10A in SpCas9) or a H868A substitution (corresponding to H840A in SpCas9) to provide a nickase, or D12A and H868A substitutions to provide a catalytically inactive Cas protein, where the positions of the D12A and H868A substitutions are defined with respect to amino acid numbering of SEQ ID NO:2. Positions corresponding to D10 and H840 of SpCas9 for Type II Cas proteins of the disclosure as shown in Table 3. Nickases and catalytically inactive Type II Cas proteins of the disclosure can be used, for example, in base editors comprising a cytosine or adenosine deaminase fusion partner. Catalytically inactive Type II Cas proteins can also be used, for example, as fusion partners for transcriptional activators or repressors. Table 3 Position Type II corresponding Position Reference SEQ ID NO Cas to D10 of corresponding to defining amino acid Protein SpCas9 H840 of SpCas9 numbering DGHJ 12 868 2 Table 3 Position Type II corresponding Position Reference SEQ ID NO Cas to D10 of corresponding to defining amino acid Protein SpCas9 H840 of SpCas9 numbering CUAZ 18 845 8 AHWY 14 900 14 CBGI 15 890 20 ASDR 15 898 26 BCZZ 17 836 32 DRCY 15 893 38 DRPY 14 878 44 EAJR 18 857 50 CFDE 15 890 56 6.3. Guide RNAs [0120] The disclosure provides gRNA molecules that can be used with Type II Cas proteins of the disclosure to edit genomic DNA, for example mammalian DNA, e.g., human DNA. gRNAs of the GLVFORVXUH^W\SLFDOO\^FRPSULVH^D^VSDFHU^RI^^^^WR^^^^QXFOHRWLGHV^LQ^OHQJWK^^7KH^VSDFHU^FDQ^EH^SRVLWLRQHG^^¶^ of a crRNA scaffold to form a full crRNA. The crRNA can be used with a tracrRNA to effect cleavage of a target genomic sequence. [0121] An exemplary crRNA scaffold sequence that can be used for DGHJ Type II Cas gRNAs comprises GUUUUAGAAUUAUGUUAUUUUAGAUAGAAAUAAUAA (SEQ ID NO:61) and an exemplary tracrRNA sequence that can be used for DGHJ Type II Cas gRNAs comprises UAUUAAUUCUAUCUAAAAUAGCAUAAUGAGUUAAAAUAAGGUUUAACCUUUAAUGCCGAAUUGCU UCGGUUUCACUUAGGUGAAAAUUGCUCUGUUCAUCAGAGC (SEQ ID NO:62). [0122] An exemplary crRNA scaffold sequence that can be used for CUAZ Type II Cas gRNAs comprises GAUUUUAAGUAUGGUUCUUUUUACUAAGUUACUGAC (SEQ ID NO:63) and an exemplary tracrRNA sequence that can be used for CUAZ Type II Cas gRNAs comprises GUAACUUAGUAAAAGGAUACCAUAACGAAAAAUUAUGAAUAUUCUACUACUUUUAGAGGGCAUAA UUGGUUGCCCUCA (SEQ ID NO:64). [0123] An exemplary crRNA scaffold sequence that can be used for AHWY Type II Cas gRNAs comprises GUUUUGGAGCAGUGGCAUUCUGACUGGUAAUCAAAC (SEQ ID NO:65) and an exemplary tracrRNA sequence that can be used for AHWY Type II Cas gRNAs comprises UUGAUACCAGUCAAGAUGUCACUGCGAGUCAAAAUAAAGACUUAGUCUUAAAUGUACCUGUAAUA GGGACGCCGCGUAGGCGGCACUUUUGAAUCGCGAAAGAGCCUUCGGGCUCUUU (SEQ ID NO:66). [0124] An exemplary crRNA scaffold sequence that can be used for CBGI Type II Cas gRNAs comprises GUUUUGGAGCAGUGUCGUUCUGCUGGUAAUCCAAC (SEQ ID NO:67) and an exemplary tracrRNA sequence that can be used for CBGI Type II Cas gRNAs comprises UGAUUACCAGUCAGGACGACACUGCGAGUCAAAAUACGGCUUUGCCAAAAAUGCCUUCGGCGCC GCGUAGGCGGCAG (SEQ ID NO:68). [0125] An exemplary crRNA scaffold sequence that can be used for ASDR Type II Cas gRNAs comprises GUUUUGGAGCAGUGUCGAUCUAACUGGUAAUCAAAC (SEQ ID NO:69) and an exemplary tracrRNA sequence that can be used for ASDR Type II Cas gRNAs comprises GAUUACCAGUUACAUCGACACUGUGAGUCAAAAUACGGCUAAGCCAAAAAUGCUGCUCUUUCGG GAGCAUCACCCCGUAGGGGGAA (SEQ ID NO:70). [0126] An exemplary crRNA scaffold sequence that can be used for BCZZ Type II Cas gRNAs comprises GUUUUAGUAUUGUGUUAUUUUAGAUAGUAAUAAAAC (SEQ ID NO:71) and an exemplary tracrRNA sequence that can be used for BCZZ Type II Cas gRNAs comprises UAUUAAAUUACUAUCUAAAAUAGCACAAUGAGUUAAAAUAAGGUUUAAACCUUAAAUGCCAACUUG AUGUUGGUUUCACUUAGGUGAA (SEQ ID NO:72). [0127] An exemplary crRNA scaffold sequence that can be used for DRCY Type II Cas gRNAs comprises GUUUUGGAGCAGUGUCAAUCUGACUGGUAAUCAAAC (SEQ ID NO:73) and an exemplary tracrRNA sequence that can be used for DRCY Type II Cas gRNAs comprises GAUUACCAGUCGGAUCGACACUGCGAGUCAAAAUACGGCUUUGCCAAAAAUGCCUGCUUGCAGG CGCCACGUAGGUGGCAA (SEQ ID NO:74). [0128] An exemplary crRNA scaffold sequence that can be used for DRPY Type II Cas gRNAs comprises GUUUGAGAGCAGUGUUAAUCCAUAGGGGAUUGAAAC (SEQ ID NO:75) and an exemplary tracrRNA sequence that can be used for DRPY Type II Cas gRNAs comprises GUGUCAAUCCCCUAUGGAUUAACACUGCGAGUUCAAAUAACGUUAUCGCAACGAAAUCGCCGUAU AUCGGUUGCACAGUGUGUGCA (SEQ ID NO:76). [0129] An exemplary crRNA scaffold sequence that can be used for EAJR Type II Cas gRNAs comprises GUUUGAGAGUAGUGUAAUCCCAAGUGGUAGUAGUGU (SEQ ID NO:77) and an exemplary tracrRNA sequence that can be used for EAJR Type II Cas gRNAs comprises CUACUACCACUUGGGAUUACACUACAAGUUCAAAUAAGCGUUCUGCGCCCUUACCGCAUUAAGUU GCGGAUUCACAGUGUGUGA (SEQ ID NO:78). [0130] An exemplary crRNA scaffold sequence that can be used for CFDE Type II Cas gRNAs comprises GUUUUGGAGCAGUGUCGAUCUAACUGGUAAUCAAAC (SEQ ID NO:79) and an exemplary tracrRNA sequence that can be used for CFDE Type II Cas gRNAs comprises GAUUACCAGUUACAUCGACACUGCGAGUCAAAAUACGGCUUUGCCAAAAAUGCCUUCCUCGGAAG GCGUCCCGUAGGGGACAAA (SEQ ID NO:80). [0131] gRNAs of the disclosure are in some embodiments single guide RNAs (sgRNAs), which typically FRPSULVH^WKH^VSDFHU^DW^WKH^^¶^HQG^RI^WKH^PROHFXOH^DQG^D^^¶^VJ51$^VFDIIROd. Alternatively, gRNAs can comprise separate crRNA and tracrRNA molecules. [0132] Further features of exemplary gRNA spacer sequences are described in Section 6.3.1 and IXUWKHU^IHDWXUHV^RI^H[HPSODU\^^¶^VJ51$^VFDIIROGV^DUH^GHVFULEHG^LQ^6HFWLRQ^6.3.2. 6.3.1. Spacers [0133] The spacer sequence is partially or fully complementary to a target sequence found in a genomic DNA sequence, for example a human genomic DNA sequence. For example, a spacer sequence can be partially or fully complementary to a nucleotide sequence in a gene having a disease causing mutation. A spacer that is partially complementary to a target sequence can have, for example, one, two, or three mismatches with the target sequence. [0134] gRNAs of the disclosure can comprise a spacer that is 15 to 30 nucleotides in length (e.g., 15 to 25, 16 to 24, 17 to 23, 18 to 22, 19 to 21, 18 to 30, 20 to 28, 22 to 26, or 23 to 25 nucleotides in length). In some embodiments, a spacer is 15 nucleotides in length. In other embodiments, a spacer is 16 nucleotides in length. In other embodiments, a spacer is 17 nucleotides in length. In other embodiments, a spacer is 18 nucleotides in length. In other embodiments, a spacer is 19 nucleotides in length. In other embodiments, a spacer is 20 nucleotides in length. In other embodiments, a spacer is 21 nucleotides in length. In other embodiments, a spacer is 22 nucleotides in length. In other embodiments, a spacer is 23 nucleotides in length. In other embodiments, a spacer is 24 nucleotides in length. In other embodiments, a spacer is 25 nucleotides in length. In other embodiments, a spacer is 26 nucleotides in length. In other embodiments, a spacer is 27 nucleotides in length. In other embodiments, a spacer is 28 nucleotides in length. In other embodiments, a spacer is 29 nucleotides in length. In other embodiments, a spacer is 30 nucleotides in length. [0135] Type II Cas endonucleases require a specific sequence, called a protospacer adjacent motif (PAM) that is downstream (e.g., directly downstream) of the target sequence on the non-target strand. Thus, spacer sequences for targeting a gene of interest can be identified by scanning the gene for PAM sequences recognized by the Type II Cas protein. Exemplary PAM sequences for Type II Cas proteins are shown in Table 4. Table 4 Type II Cas protein PAM(s) Most preferred PAM(s) DGHJ Type II Cas NGG NGG CUAZ Type II Cas NNGNC NNGNCNY AHWY Type II Cas NNRC NNAC, NNACNT CBGI Type II Cas NNRTA NNRTA ASDR Type II Cas NNNNCC NNANCC, NNACCC BCZZ Type II Cas NGG NGG DRCY Type II Cas In silico predicted : NNAT, In vitro determined: NRATA NNATM, NNATA, NVAT, NVATM In vitro determined: NVRTA, NRRTA, NVATA DRPY Type II Cas NNNAC NRNAC, NRHAC, NRNACD, NRHACD, NRNACA, NRHACA EAJR Type II Cas NNNRNY NRNRNY,NRNRNC, NANANC, NNNRDC CFDE Type II Cas NNART NRART N = A, T, C, or G; M = A or C; V = G, C, or A; R = A or G; Y = C or T; H = A, C or T; D = A, G or T Table 4 Type II Cas protein PAM(s) Most preferred PAM(s) All PAMs are in vitro determined PAMs, except where noted (see Example 2). [0136] Example 3 describes exemplary sequences that can be used to target TRAC, B2M, and PD1 genomic sequences. In some embodiments, a gRNA of the disclosure comprises a spacer sequence targeting TRAC. In some embodiments, a gRNA of the disclosure comprises a spacer sequence targeting B2M. In some embodiments, a gRNA of the disclosure comprises a spacer sequence targeting PD1. [0137] Additional exemplary spacer sequences that can be used in gRNAs of the disclosure are set forth in Table 5A, Table 5B, and Table 5C. Table 5A Exemplary Spacer Sequences Targeting B2M Type II Cas Name Spacer Sequence SEQ ID NO: AHWY sgRNA-AHWY-B2M-g1 CAGAAAGAGAGAGUAGCGCGAGC 138 AHWY sgRNA-AHWY-B2M-g2 CCGUGGCCUUAGCUGUGCUCGCG 139 AHWY sgRNA-AHWY-B2M-g3 UCAGGUACUCCAAAGAUUCAGGU 140 AHWY sgRNA-AHWY-B2M-g4 GUACUCCAAAGAUUCAGGUUUAC 141 AHWY sgRNA-AHWY-B2M-g5 GACACAUAGCAAUUCAGGAAAUU 142 ASDR sgRNA-ASDR-B2M-g1 ACUCUCUCUUUCUGGCCUGGAGG 143 ASDR sgRNA-ASDR-B2M-g2 CCAAAGAUUCAGGUUUACUCACG 144 ASDR sgRNA-ASDR-B2M-g3 CUGAAUUGCUAUGUGUCUGGGUU 145 ASDR sgRNA-ADSR-B2M-g4 AUUGCUAUGUGUCUGGGUUUCAU 146 ASDR sgRNA-ASDR-B2M-g5 GUCAACUUCAAUGUCGGAUGGAUG 147 ASDR sgRNA-ASDR-B2M-g5_20nt ACUUCAAUGUCGGAUGGAUG 148 CBGI sgRNA-CBGI-B2M-g1 GAUAGCCUCCAGGCCAGAAAGAGA 149 CBGI sgRNA-CBGI-B2M-g2 GGAGGGUAGGAGAGACUCACGCU 150 CBGI sgRNA-CBGI-B2M-g3 GACGUGAGUAAACCUGAAUCUUUG 151 CBGI sgRNA-CBGI-B2M-g4 UUCCAUUCUCUGCUGGAUGACGU 152 CBGI sgRNA-CBGI-B2M-g5 GUCGGAUGGAUGAAACCCAGAC 153 CFDE sgRNA-CFDE-B2M-g1 GAUAGCCUCCAGGCCAGAAAGAG 154 CFDE sgRNA-CFDE-B2M-g2 UCUGGCCUGGAGGCUAUCCAGCG 155 CFDE sgRNA-CFDE-B2M-g3 GACGUGAGUAAACCUGAAUCUUU 156 CFDE sgRNA-CFDE-B2M-g4 GUUUACUCACGUCAUCCAGCAG 157 CFDE sgRNA-CFDE-B2M-g5 GAGAAUGGAAAGUCAAAUUUCC 158 DRPY sgRNA-DRPY-B2M-g1 UUCAGUGGGGGUGAAUUCAGUGU 159 DRPY sgRNA-DRPY-B2M-g2 GCCAGAAAGAGAGAGUAGCGCG 160 DRPY sgRNA-DRPY-B2M-g3 CAAUGUCGGAUGGAUGAAACCCA 161 DRPY sgRNA-DRPY-B2M-g4 GCAAGGACUGGUCUUUCUAUCUCU 162 Table 5B Exemplary Spacer Sequences Targeting TRAC Type II Cas Name Spacer Sequence SEQ ID NO: AHWY sgRNA-AHWY-TRAC-g1 AUCAAAAUCGGUGAAUAGGCAGAC 163 AHWY sgRNA-AHWY-TRAC-g2 ACACAUCAGAAUCCUUACUUUGUG 164 AHWY sgRNA-AHWY-TRAC-g3 GCACUGUUGCUCUUGAAGUCCAU 165 AHWY sgRNA-AHWY-TRAC-g4 AACUGUGCUAGACAUGAGGUCUAU 166 AHWY sgRNA-AHWY-TRAC-g5 GUAUAUCACAGACAAAACUGUGCU 167 ASDR sgRNA-ASDR-TRAC-g1 UCUUGUCCCACAGAUAUCCA 168 ASDR sgRNA-ASDR-TRAC-g2 CCCACAGAUAUCCAGAACCC 169 ASDR sgRNA-ASDR-TRAC-g3 GUGUACCAGCUGAGAGACUCU 170 ASDR sgRNA-ASDR-TRAC-g4 UGUCUGUGAUAUACACAUCA 171 ASDR sgRNA-ASDR-TRAC-g5 GCCACAGCACUGUUGCUCUUG 172 CBGI sgRNA-CBGI-TRAC-g1 GAUAUCCAGAACCCUGACCCUGCC 173 CBGI sgRNA-CBGI-TRAC-g2 CACUGGAUUUAGAGUCUCUCAGC 174 CBGI sgRNA-CBGI-TRAC-g3 UUUGUUUGAGAAUCAAAAUCGGU 175 CBGI sgRNA-CBGI-TRAC-g4 GAUUCUCAAACAAAUGUGUCACA 176 CFDE sgRNA-CFDE-TRAC-g5 GUCACAAAGUAAGGAUUCUGAU 177 CFDE sgRNA-CFDE-TRAC-g1 GCCGUGUACCAGCUGAGAGACUC 178 CFDE sgRNA-CFDE-TRAC-g2 GCUGAGAGACUCUAAAUCCAGUGA 179 CFDE sgRNA-CFDE-TRAC-g3 GCAGACAGACUUGUCACUGGAUUU 180 CFDE sgRNA-CFDE-TRAC-g4 AUUCACCGAUUUUGAUUCUCAAA 181 CFDE sgRNA-CFDE-TRAC-g5 AUUUGUUUGAGAAUCAAAAUCGG 182 CFDE sgRNA-CFDE-TRAC-g6 UGAUUCUCAAACAAAUGUGUCAC 183 CFDE sgRNA-CFDE-TRAC-g7 UUACUUUGUGACACAUUUGUUUG 184 CFDE sgRNA-CFDE-TRAC-g8 GUUUUGUCUGUGAUAUACACAUC 185 CFDE sgRNA-CFDE-TRAC-g9 CAGGCCACAGCACUGUUGCUCUU 186 CFDE sgRNA-CFDE-TRAC-g10 gAACAGUGCUGUGGCCUGGAGCAA 187 DRPY sgRNA-DRPY-TRAC-g1 ACUGGAUUUAGAGUCUCUCAGCU 188 DRPY sgRNA-DRPY-TRAC-g2 GCCUAUUCACCGAUUUUGAUUCU 189 DRPY sgRNA-DRPY-TRAC-g3 ACACAUCAGAAUCCUUACUUUGU 190 DRPY sgRNA-DRPY-TRAC-g4 CUAGCACAGUUUUGUCUGUGAUA 191 DRPY sgRNA-DRPY-TRAC-g5 UGAAGUCCAUAGACCUCAUGUCU 192 DRPY sgRNA-DRPY-TRAC-g6 GUCUAGCACAGUUUUGUCUGUGA 193 DRPY sgRNA-DRPY-TRAC-g7 AUAAUGCUGUUGUUGAAGGCGUU 194 Table 5C Exemplary Spacer Sequences Targeting PD1 Type II Cas Name Spacer Sequence SEQ ID NO: AHWY sgRNA-AHWY-PD1-g1 AUCCUGGCCGCCAGCCCAGUUGUA 195 AHWY sgRNA-AHWY-PD1-g2 GGGGCUGCUCCAGGCAUGCAGAUC 196 AHWY sgRNA-AHWY-PD1-g3 GCCCAGUUGUAGCACCGCCCAGA 197 AHWY sgRNA-AHWY-PD1-g4 GGCCAGUCGUCUGGGCGGUGCUA 198 Table 5C Exemplary Spacer Sequences Targeting PD1 Type II Cas Name Spacer Sequence SEQ ID NO: ASDR sgRNA-ASDR-PD1-g1 GUGGGGCUGCUCCAGGCAUGC 199 ASDR sgRNA-ASDR-PD1-g2 CGACCCCACCUACCUAAGAA 200 ASDR sgRNA-ASDR-PD1-g3 CAGGCAUGCAGAUCCCACAG 201 ASDR sgRNA-ASDR-PD1-g4 GCCGCCAGCCCAGUUGUAGCA 202 ASDR sgRNA-ASDR-PD1-g5 CUAAGAACCAUCCUGGCCGC 203 ASDR sgRNA-ASDR-PD1-g6 UCAGACUCCCCAGACAGGCCCUG 204 ASDR sgRNA-ASDR-PD1-g7 CGGGGCCUCCGAGGCCGCACCUG 205 CBGI sgRNA-CBGI-PD1-g1 GAACCAUCCUGGCCGCCAGCCCAG 206 CBGI sgRNA-CBGI-PD1-g2 GGCUGGCGGCCAGGAUGGUUCUU 207 CBGI sgRNA-CBGI-PD1-g3 CAUCGGAGAGCUUCGUGCUAAAC 208 CBGI sgRNA-CBGI-PD1-g4 CCGUCUGGUUGCUGGGGCUCAUG 209 CBGI sgRNA-CBGI-PD1-g5 GGGCCAGGGAGAUGGCCCCACAG 210 CFDE sgRNA-CFDE-PD1-g1 GGGGGGUUCCAGGGCCUGUCUGG 211 CFDE sgRNA-CFDE-PD1-g2 CGGGCCCUGACCACGCUCAUGUG 212 DRPY sgRNA-DRPY-PD1-g1 GCCACCUUCACCUGCAGCUUCUCC 213 DRPY sgRNA-DRPY-PD1-g2 GGCCAGGACUGCCGCUUCCGUGU 214 DRPY sgRNA-DRPY-PD1-g3 UGGGGCUCAUGCGGUACCAGUUU 215 DRPY sgRNA-DRPY-PD1-g4 CAACACAUCGGAGAGCUUCGUGC 216 DRPY sgRNA-DRPY-PD1-g5 GUCACGCCCGUUGGGCAGUUGUGU 217 [0138] In some embodiments, a gRNA of the disclosure has a spacer whose nucleotide sequence comprises 15 or more consecutive nucleotides from a sequence shown in Table 5A. In some embodiments, a gRNA of the disclosure has a spacer whose nucleotide sequence comprises 16 or more consecutive nucleotides from a sequence shown in Table 5A. In some embodiments, a gRNA of the disclosure has a spacer whose nucleotide sequence comprises 17 or more consecutive nucleotides from a sequence shown in Table 5A. In some embodiments, a gRNA of the disclosure has a spacer whose nucleotide sequence comprises 18 or more consecutive nucleotides from a sequence shown in Table 5A. In some embodiments, a gRNA of the disclosure has a spacer whose nucleotide sequence comprises 19 or more consecutive nucleotides from a sequence shown in Table 5A. In some embodiments, a gRNA of the disclosure has a spacer whose nucleotide sequence comprises 20 or more consecutive nucleotides from a sequence shown in Table 5A. In some embodiments, a gRNA of the disclosure has a spacer whose nucleotide sequence comprises 21 or more consecutive nucleotides from a sequence shown in Table 5A. In some embodiments, a gRNA of the disclosure has a spacer whose nucleotide sequence comprises 22 or more consecutive nucleotides from a sequence shown in Table 5A. In some embodiments, a gRNA of the disclosure has a spacer whose nucleotide sequence comprises 23 or more consecutive nucleotides from a sequence shown in Table 5A. In some embodiments, a gRNA of the disclosure has a spacer whose nucleotide sequence comprises 24 consecutive nucleotides from a sequence shown in Table 5A. [0139] In some embodiments, a gRNA of the disclosure has a spacer whose nucleotide sequence comprises 15 or more consecutive nucleotides from a sequence shown in Table 5B. In some embodiments, a gRNA of the disclosure has a spacer whose nucleotide sequence comprises 16 or more consecutive nucleotides from a sequence shown in Table 5B. In some embodiments, a gRNA of the disclosure has a spacer whose nucleotide sequence comprises 17 or more consecutive nucleotides from a sequence shown in Table 5B. In some embodiments, a gRNA of the disclosure has a spacer whose nucleotide sequence comprises 18 or more consecutive nucleotides from a sequence shown in Table 5B. In some embodiments, a gRNA of the disclosure has a spacer whose nucleotide sequence comprises 19 or more consecutive nucleotides from a sequence shown in Table 5B. In some embodiments, a gRNA of the disclosure has a spacer whose nucleotide sequence comprises 20 or more consecutive nucleotides from a sequence shown in Table 5B. In some embodiments, a gRNA of the disclosure has a spacer whose nucleotide sequence comprises 21 or more consecutive nucleotides from a sequence shown in Table 5B. In some embodiments, a gRNA of the disclosure has a spacer whose nucleotide sequence comprises 22 or more consecutive nucleotides from a sequence shown in Table 5B. In some embodiments, a gRNA of the disclosure has a spacer whose nucleotide sequence comprises 23 or more consecutive nucleotides from a sequence shown in Table 5B. In some embodiments, a gRNA of the disclosure has a spacer whose nucleotide sequence comprises 24 consecutive nucleotides from a sequence shown in Table 5B. [0140] In some embodiments, a gRNA of the disclosure has a spacer whose nucleotide sequence comprises 15 or more consecutive nucleotides from a sequence shown in Table 5C. In some embodiments, a gRNA of the disclosure has a spacer whose nucleotide sequence comprises 16 or more consecutive nucleotides from a sequence shown in Table 5C. In some embodiments, a gRNA of the disclosure has a spacer whose nucleotide sequence comprises 17 or more consecutive nucleotides from a sequence shown in Table 5C. In some embodiments, a gRNA of the disclosure has a spacer whose nucleotide sequence comprises 18 or more consecutive nucleotides from a sequence shown in Table 5C. In some embodiments, a gRNA of the disclosure has a spacer whose nucleotide sequence comprises 19 or more consecutive nucleotides from a sequence shown in Table 5C. In some embodiments, a gRNA of the disclosure has a spacer whose nucleotide sequence comprises 20 or more consecutive nucleotides from a sequence shown in Table 5C. In some embodiments, a gRNA of the disclosure has a spacer whose nucleotide sequence comprises 21 or more consecutive nucleotides from a sequence shown in Table 5C. In some embodiments, a gRNA of the disclosure has a spacer whose nucleotide sequence comprises 22 or more consecutive nucleotides from a sequence shown in Table 5C. In some embodiments, a gRNA of the disclosure has a spacer whose nucleotide sequence comprises 23 or more consecutive nucleotides from a sequence shown in Table 5C. In some embodiments, a gRNA of the disclosure has a spacer whose nucleotide sequence comprises 24 consecutive nucleotides from a sequence shown in Table 5C. 6.3.2. sgRNA Molecules [0141] gRNAs of the disclosure can be single-guide RNA (sgRNA) molecules. A sgRNA can comprise, in the 5' to 3' direction, an optional spacer extension sequence, a spacer sequence, a minimum CRISPR repeat sequence, a single-PROHFXOH^JXLGH^OLQNHU^^D^PLQLPXP^WUDFU51$^VHTXHQFH^^D^^¶^WUDFU51$^ sequence and an optional tracrRNA extension sequence. The optional tracrRNA extension can comprise elements that contribute additional functionality (e.g., stability) to the guide RNA. The single-molecule guide linker can link the minimum CRISPR repeat and the minimum tracrRNA sequence to form a hairpin structure. The optional tracrRNA extension can comprise one or more hairpins. [0142] The sgRNA can comprise a variable length spacer sequence (e.g.^^^^^WR^^^^QXFOHRWLGHV^^DW^WKH^^¶^ HQG^RI^WKH^VJ51$^VHTXHQFH^DQG^D^^¶^VJ51$^VHJPHQW^^ [0143] Type II Cas gRNAs typically comprise a repeat-antirepeat duplex and/or one or more stem-loops JHQHUDWHG^E\^WKH^J51$¶V^VHFRQdary structure. The length of the repeat-antirepeat duplex and/or one or more stem-loops can be modified in order to modulate (e.g., increase) the editing efficacy of a Type II Cas nuclease, and/or to reduce the size of a guide RNA for easier vectorization in situations in which the cargo size of the vector is limiting (e.g., AAV vectors). [0144] For example, the repeat-antirepeat duplex (which in a sgRNA is fused through a synthetic linker to become an additional stem loop in the structure) can be trimmed at different lengths without generally having detrimental effects on nuclease function and in some cases even producing increased enzymatic activity. If bulges are present within this duplex they generally should be retained in the final guide RNA sequence. [0145] Further optimization of the structure can be obtained by introducing targeted base changes into the stems of the gRNA to increase their stability and folding. Such base changes will preferably correspond to the introduction of G:C couples, which are known to generate the strongest Watson-Crick pairing. For the sake of clarity, these substitutions can consist in the introduction of a G or a C in a specific position of a stem together with a complementary substitution in another position of the gRNA sequence which is predicted to base pair with the former, for example according to available bioinformatic tools for RNA folding such as UNAfold or RNAfold. [0146] Stem-loop trimming can also be exploited to stabilize desired secondary structures by removing portions of the guide RNA producing unwanted secondary structures through annealing with other regions of the RNA molecule. [0147] ([HPSODU\^^¶^VJ51$^VFDIIROG^VHTXHQFHV^IRU^Type II Cas sgRNAs are shown in Table 6. Table 6 Sequences of sgRNA Scaffolds for Type IIA Cas Proteins Name Sequence SEQ ID NO: DGHJ Type II GUUUUAGAAUUAUGUUAUUUUAGAUAGAAAUAAUAAGAAAUAUUAAU 81 Cas sgRNA UCUAUCUAAAAUAGCAUAAUGAGUUAAAAUAAGGUUUAACCUUUAAU scaffold GCCGAAUUGCUUCGGUUUCACUUAGGUGAAAAUUGCUCUGUUCAUC AGAGC DGHJ Type II GUUUUAGAAUUAUGGAAACAUAAUGAGUUAAAAUAAGGUUUAACCUU 82 Cas sgRNA UAAUGCCGAAUUGCUUCGGUUUCACUUAGGUGAAAAUUGCUCUGUU scaffold CAUCAGAGC (trimmed) (v1) DGHJ Type II GUUUUAGAAUUAUGGAAACAUAAUGAGUUAAAAUAAGGUUUAACCUU 83 Cas sgRNA UAAUGCCGAAUUGCUUCGGUUUCACUUAGGUGAAAA scaffold (trimmed) (alternate) (v2) Table 6 Sequences of sgRNA Scaffolds for Type IIA Cas Proteins Name Sequence SEQ ID NO: CUAZ Type II GAUUUUAAGUAUGGUUCUUUUUACUAAGUUACUGACGAAAGUAACU 84 Cas sgRNA UAGUAAAAGGAUACCAUAACGAAAAAUUAUGAAUAUUCUACUACUUU scaffold UAGAGGGCAUAAUUGGUUGCCCUCA CUAZ Type II GAUUUUAAGUAUGGGAAACCAUAACGAAAAAUUAUGAAUAUUCUACU 85 Cas sgRNA ACUUUUAGAGGGCAUAAUUGGUUGCCCUCA scaffold (trimmed) AHWY Type II GUUUUGGAGCAGUGGCAUUCUGACUGGUAAUCAAACGAAAUUGAUA 86 Cas sgRNA CCAGUCAAGAUGUCACUGCGAGUCAAAAUAAAGACUUAGUCUUAAAU scaffold GUACCUGUAAUAGGGACGCCGCGUAGGCGGCACUUUUGAAUCGCGA AAGAGCCUUCGGGCUC AHWY Type II GUUUUGGAGCAGUGGAAACACUGCGAGUCAAAAUAAAGACUUAGUC 87 Cas sgRNA UUAAAUGUACCUGUAAUAGGGACGCCGCGUAGGCGGCACUUUUGAA scaffold UCGCGAAAGAGCCUUCGGGCUC (trimmed) (v1) AHWY Type II GUUUUGGAGCAGUGGAAACACUGCGAGUCAAAAUAAAGACUUAGUC 88 Cas sgRNA UUAAAUGUACCUGUAAUAGGGACGCCGCGUAGGCGGCAC scaffold (trimmed) (alternate) (v2) CBGI Type II GUUUUGGAGCAGUGUCGUUCUGCUGGUAAUCCAACGAAAUGAUUAC 89 Cas sgRNA CAGUCAGGACGACACUGCGAGUCAAAAUACGGCUUUGCCAAAAAUG scaffold CCUUCGGCGCCGCGUAGGCGGCAG CBGI Type II GUUUUGGAGCAGUGGAAACACUGCGAGUCAAAAUACGGCUUUGCCA 90 Cas sgRNA AAAAUGCCUUCGGCGCCGCGUAGGCGGCAG scaffold (trimmed) ASDR Type II GUUUUGGAGCAGUGUCGAUCUAACUGGUAAUCAAACGAAAGAUUAC 91 Cas sgRNA CAGUUACAUCGACACUGUGAGUCAAAAUACGGCUAAGCCAAAAAUGC scaffold UGCUCUUUCGGGAGCAUCACCCCGUAGGGGGAA ASDR Type II GUUUUGGAGCAGUGGAAACACUGUGAGUCAAAAUACGGCUAAGCCA 92 Cas sgRNA AAAAUGCUGCUCUUUCGGGAGCAUCACCCCGUAGGGGGAA scaffold (trimmed) BCZZ Type II GUUUUAGUAUUGUGUUAUUUUAGAUAGUAAUAAAACGAAAUAUUAAA 93 Cas sgRNA UUACUAUCUAAAAUAGCACAAUGAGUUAAAAUAAGGUUUAAACCUUA scaffold AAUGCCAACUUGAUGUUGGUUUCACUUAGGUGAA BCZZ Type II GUUUUAGUAUUGAAAAAUGAGUUAAAAUAAGGUUUAAACCUUAAAUG 94 Cas sgRNA CCAACUUGAUGUUGGUUUCACUUAGGUGAA scaffold (trimmed) DRCY Type II GUUUUGGAGCAGUGUCAAUCUGACUGGUAAUCAAACGAAAGAUUAC 95 Cas sgRNA CAGUCGGAUCGACACUGCGAGUCAAAAUACGGCUUUGCCAAAAAUG scaffold CCUGCUUGCAGGCGCCACGUAGGUGGCAA DRCY Type II GUUUUGGAGCGAAAGCGAGUCAAAAUACGGCUUUGCCAAAAAUGCC 96 Cas sgRNA UGCUUGCAGGCGCCACGUAGGUGGCAA scaffold (trimmed) DRPY Type II GUUUGAGAGCAGUGUUAAUCCAUAGGGGAUUGAAACGAAAGUGUCA 97 Cas sgRNA AUCCCCUAUGGAUUAACACUGCGAGUUCAAAUAACGUUAUCGCAACG scaffold AAAUCGCCGUAUAUCGGUUGCACAGUGUGUGCA DRPY Type II GUUUGAGAGCGAAAGCGAGUUCAAAUAACGUUAUCGCAACGAAAUC 98 Cas sgRNA GCCGUAUAUCGGUUGCACAGUGUGUGCA Table 6 Sequences of sgRNA Scaffolds for Type IIA Cas Proteins Name Sequence SEQ ID NO: scaffold (trimmed) EAJR Type II GUUUGAGAGUAGUGUAAUCCCAAGUGGUAGUAGUGUGAAACUACUA 99 Cas sgRNA CCACUUGGGAUUACACUACAAGUUCAAAUAAGCGUUCUGCGCCCUU scaffold ACCGCAUUAAGUUGCGGAUUCACAGUGUGUGA EAJR Type II GUUUGAGAGUGAAAACAAGUUCAAAUAAGCGUUCUGCGCCCUUACC 100 Cas sgRNA GCAUUAAGUUGCGGAUUCACAGUGUGUGA scaffold (trimmed) CFDE Type II GUUUUGGAGCAGUGUCGAUCUAACUGGUAAUCAAACGAAAGAUUAC 101 Cas sgRNA CAGUUACAUCGACACUGCGAGUCAAAAUACGGCUUUGCCAAAAAUGC scaffold CUUCCUCGGAAGGCGUCCCGUAGGGGACAAA CFDE Type II GUUUUGGAGCGAAAGCGAGUCAAAAUACGGCUUUGCCAAAAAUGCC 102 Cas sgRNA UUCCUCGGAAGGCGUCCCGUAGGGGACAAA scaffold (trimmed) [0148] The sgRNA (e.g., for use with CUAZ Type II Cas protein, AHWY Type II Cas protein, DGHJ Type II Cas protein, CBGI Type II Cas protein, ASDR Type II Cas protein, BCZZ Type II Cas protein, DRCY Type II Cas protein, DRPY Type II Cas protein, EAJR Type II Cas protein, and/or CFDE Type II Cas protein^^FDQ^FRPSULVH^QR^XUDFLO^EDVH^DW^WKH^^¶^HQG^RI^WKH^VJ51$^VHTXHQFH^^7\SLFDOO\^^KRZHYHU^^WKH^ VJ51$^FRPSULVHV^RQH^RU^PRUH^XUDFLO^EDVHV^DW^WKH^^¶^HQG^RI^WKH^VJ51$^VHTXHQFH^^IRU^H[DPple to SURPRWH^FRUUHFW^VJ51$^IROGLQJ^^)RU^H[DPSOH^^WKH^VJ51$^FDQ^FRPSULVH^^^XUDFLO^^8^^DW^WKH^^¶^HQG^RI^WKH^ VJ51$^VHTXHQFH^^7KH^VJ51$^FDQ^FRPSULVH^^^XUDFLO^^88^^DW^WKH^^¶^HQG^RI^WKH^VJ51$^VHTXHQFH^^7KH^ VJ51$^FDQ^FRPSULVH^^^XUDFLO^^888^^DW^WKH^^¶^HQG^RI^WKH sgRNA sequence. The sgRNA can comprise 4 XUDFLO^^8888^^DW^WKH^^¶^HQG^RI^WKH^VJ51$^VHTXHQFH^^7KH^VJ51$^FDQ^FRPSULVH^^^XUDFLO^^88888^^DW^WKH^ ^¶^HQG^RI^WKH^VJ51$^VHTXHQFH^^7KH^VJ51$^FDQ^FRPSULVH^^^XUDFLO^^888888^^DW^WKH^^¶^HQG^RI^WKH^VJ51$^ sequence. The sgRN$^FDQ^FRPSULVH^^^XUDFLO^^8888888^^DW^WKH^^¶^HQG^RI^WKH^VJ51$^VHTXHQFH^^7KH^ VJ51$^FDQ^FRPSULVH^^^XUDFLO^^88888888^^DW^WKH^^¶^HQG^RI^WKH^VJ51$^VHTXHQFH^^'LIIHUHQW^OHQJWK^ VWUHWFKHV^RI^XUDFLO^FDQ^EH^DSSHQGHG^DW^WKH^^¶HQG^RI^D^VJ51$^DV^WHUPLQDWRUV^^7KXV^^IRU^H[DPSOH^^WKH^^¶^ sgRNA sequences set forth in Table 6 can be modified by adding (or removing) one or more uracils at the end of the sequence. [0149] In some embodiments, a sgRNA scaffold for use with a DGHJ Type II Cas protein comprises the sequence GUUUUAGAAUUAUGGAAACAUAAUGAGUUAAAAUAAGGUUUAACCUUUAAUGCCGAAUUGCUUCG GUUUCACUUAGGUGAAAAUUGCUCUGUUCAUCAGAGCUUUUUU (SEQ ID NO:103). In some embodiments, a sgRNA scaffold for use with a DGHJ Type II Cas protein comprises the sequence GUUUUAGAAUUAUGGAAACAUAAUGAGUUAAAAUAAGGUUUAACCUUUAAUGCCGAAUUGCUUCG GUUUCACUUAGGUGAAAAUUUUUU (SEQ ID NO:104). [0150] In some embodiments, a sgRNA scaffold for use with a CUAZ Type II Cas protein comprises the sequence GAUUUUAAGUAUGGGAAACCAUAACGAAAAAUUAUGAAUAUUCUACUACUUUUAGAGGGCAUAAU UGGUUGCCCUCAUUUUUU (SEQ ID NO:105). [0151] In some embodiments, a sgRNA scaffold for use with an AHWY Type II Cas protein comprises the sequence GUUUUGGAGCAGUGGCAUUCUGACUGGUAAUCAAACGAAAUUGAUACCAGUCAAGAUGUCACUG CGAGUCAAAAUAAAGACUUAGUCUUAAAUGUACCUGUAAUAGGGACGCCGCGUAGGCGGCACUU UUGAAUCGCGAAAGAGCCUUCGGGCUCUUU (SEQ ID NO:106). In some embodiments, a sgRNA scaffold for use with an AHWY Type II Cas protein comprises the sequence GUUUUGGAGCAGUGGAAACACUGCGAGUCAAAAUAAAGACUUAGUCUUAAAUGUACCUGUAAUAG GGACGCCGCGUAGGCGGCACUUUUGAAUCGCGAAAGAGCCUUCGGGCUCUUUUUU (SEQ ID NO:107). In some embodiments, a sgRNA scaffold for use with an AHWY Type II Cas protein comprises the sequence GUUUUGGAGCAGUGGAAACACUGCGAGUCAAAAUAAAGACUUAGUCUUAAAUGUACCUGUAAUAG GGACGCCGCGUAGGCGGCACUUUUUU (SEQ ID NO:108). [0152] In some embodiments, a sgRNA scaffold for use with a CBGI Type II Cas protein comprises the sequence GUUUUGGAGCAGUGGAAACACUGCGAGUCAAAAUACGGCUUUGCCAAAAAUGCCUUCGGCGCCG CGUAGGCGGCAGUUUUUU (SEQ ID NO:109). [0153] In some embodiments, a sgRNA scaffold for use with an ASDR Type II Cas protein comprises the sequence GUUUUGGAGCAGUGGAAACACUGUGAGUCAAAAUACGGCUAAGCCAAAAAUGCUGCUCUUUCGG GAGCAUCACCCCGUAGGGGGAAUUUUUU (SEQ ID NO:110). [0154] In some embodiments, a sgRNA scaffold for use with a BCZZ Type II Cas protein comprises the sequence GUUUUAGUAUUGAAAAAUGAGUUAAAAUAAGGUUUAAACCUUAAAUGCCAACUUGAUGUUGGUUU CACUUAGGUGAAUUUUUU (SEQ ID NO:111). [0155] In some embodiments, a sgRNA scaffold for use with a DRCY Type II Cas protein comprises the sequence GUUUUGGAGCGAAAGCGAGUCAAAAUACGGCUUUGCCAAAAAUGCCUGCUUGCAGGCGCCACGU AGGUGGCAAUUUUUU (SEQ ID NO:112). [0156] In some embodiments, a sgRNA scaffold for use with a DRPY Type II Cas protein comprises the sequence GUUUGAGAGCGAAAGCGAGUUCAAAUAACGUUAUCGCAACGAAAUCGCCGUAUAUCGGUUGCAC AGUGUGUGCAUUUUUU (SEQ ID NO:113). [0157] In some embodiments, a sgRNA scaffold for use with an EAJR Type II Cas protein comprises the sequence GUUUGAGAGUGAAAACAAGUUCAAAUAAGCGUUCUGCGCCCUUACCGCAUUAAGUUGCGGAUUC ACAGUGUGUGAUUUUUU (SEQ ID NO:114). [0158] In some embodiments, a sgRNA scaffold for use with a CFDE Type II Cas protein comprises the sequence GUUUUGGAGCGAAAGCGAGUCAAAAUACGGCUUUGCCAAAAAUGCCUUCCUCGGAAGGCGUCCC GUAGGGGACAAAUUUUUU (SEQ ID NO:115). 6.3.3. Modified gRNA Molecules [0159] Guide RNAs can be readily synthesized by chemical means, enabling a number of modifications to be readily incorporated, as described in the art. The disclosed gRNA (e.g., sgRNA) molecules can be unmodified or can contain any one or more of an array of chemical modifications. [0160] While chemical synthetic procedures are continually expanding, purifications of such RNAs by procedures such as high-performance liquid chromatography (HPLC, which avoids the use of gels such as PAGE) tends to become more challenging as polynucleotide lengths increase significantly beyond a hundred or so nucleotides. One approach that can be used for generating chemically modified RNAs of greater length is to produce two or more molecules that are ligated together. Much longer RNAs, such as those encoding a Type II Cas endonuclease, are more readily generated enzymatically. While fewer types of modifications are available for use in enzymatically produced RNAs, there are still modifications that can be used to, for instance, enhance stability, reduce the likelihood or degree of innate immune response, and/or enhance other attributes, as described herein and in the art. [0161] By way of illustration of various types of modifications, especially those used frequently with smaller chemically synthesized RNAs, modifications can comprise one or more nucleotides modified at the 2' position of the sugar, for instance a 2'-O-alkyl, 2'-O-alkyl-O-alkyl, or 2'-fluoro-modified nucleotide. In some examples, RNA modifications can comprise 2'-fluoro, 2'-amino or 2'-O-methyl modifications on the ribose of pyrimidines, abasic residues, or an inverted base at the 3' end of the RNA. Such modifications can be routinely incorporated into oligonucleotides and these oligonucleotides have been shown to have a higher Tm (thus, higher target binding affinity) than 2'-deoxyoligonucleotides against a given target. [0162] A number of nucleotide and nucleoside modifications have been shown to make the oligonucleotide into which they are incorporated more resistant to nuclease digestion than the native oligonucleotide; these modified oligos survive intact for a longer time than unmodified oligonucleotides. Specific examples of modified oligonucleotides include those comprising modified backbones, for example, phosphorothioates, phosphotriesters, methyl phosphonates, short chain alkyl or cycloalkyl intersugar linkages or short chain heteroatomic or heterocyclic intersugar linkages. Some oligonucleotides are oligonucleotides with phosphorothioate backbones and those with heteroatom backbones, particularly CH2-NH-O-CH2, CH,~N(CH3)-O-CH2 (known as a methylene(methylimino) or MMI backbone), CH2-O-N (CH3)-CH2, CH2 -N (CH3)-N (CH3)-CH2 and O-N (CH3)- CH2 -CH2 backbones, wherein the native phosphodiester backbone is represented as O- P- O- CH,); amide backbones (see De Mesmaeker et al.1995, Ace. Chem. Res., 28:366-374); morpholino backbone structures (see U.S. Patent No.5,034,506); peptide nucleic acid (PNA) backbone (wherein the phosphodiester backbone of the oligonucleotide is replaced with a polyamide backbone, the nucleotides being bound directly or indirectly to the aza nitrogen atoms of the polyamide backbone, see Nielsen et al., 1991, Science 254:1497). Phosphorus-containing linkages include, but are not limited to, phosphorothioates, chiral phosphorothioates, phosphorodithioates, phosphotriesters, aminoalkylphosphotriesters, methyl and other alkyl phosphonates comprising 3'alkylene phosphonates and chiral phosphonates, phosphinates, phosphoramidates comprising 3'-amino phosphoramidate and aminoalkylphosphoramidates, thionophosphoramidates, thionoalkylphosphonates, thionoalkylphosphotriesters, and boranophosphates having normal 3'-5' linkages, 2'-5' linked analogs of these, and those having inverted polarity wherein the adjacent pairs of nucleoside units are linked 3'-5' to 5'-3' or 2'-5' to 5'-2'; see U.S. Patent Nos.3,687,808; 4,469,863; 4,476,301; 5,023,243; 5,177,196; 5,188,897; 5,264,423; 5,276,019; 5,278,302; 5,286,717; 5,321,131; 5,399,676; 5,405,939; 5,453,496; 5,455,233; 5,466,677; 5,476,925; 5,519,126; 5,536,821; 5,541,306; 5,550,111; 5,563,253; 5,571,799; 5,587,361; and 5,625,050. [0163] Morpholino-based oligomeric compounds are described in Braasch and David Corey, 2002, Biochemistry, 41(14):4503-4510; Genesis, Volume 30, Issue 3, (2001); Heasman, 2002, Dev. Biol., 243: 209-214; Nasevicius et al., 2000, Nat. Genet., 26:216-220; Lacerra et al., 2000, Proc. Natl. Acad. Sci., 97: 9591-9596; and U.S. Patent No.5,034,506. [0164] Cyclohexenyl nucleic acid oligonucleotide mimetics are described in Wang et al., 2000, J. Am. Chem. Soc., 122: 8595-8602. [0165] Modified oligonucleotide backbones that do not include a phosphorus atom therein have backbones that are formed by short chain alkyl or cycloalkyl internucleoside linkages, mixed heteroatom and alkyl or cycloalkyl internucleoside linkages, or one or more short chain heteroatomic or heterocyclic intemucleoside linkages. These comprise those having morpholino linkages (formed in part from the sugar portion of a nucleoside); siloxane backbones; sulfide, sulfoxide and sulfone backbones; formacetyl and thioformacetyl backbones; methylene formacetyl and thioformacetyl backbones; alkene containing backbones; sulfamate backbones; methyleneimino and methylenehydrazino backbones; sulfonate and sulfonamide backbones; amide backbones; and others having mixed N, O, S, and CH2 component parts; see U.S. Patent Nos.5,034,506; 5,166,315; 5,185,444; 5,214,134; 5,216,141; 5,235,033; 5,264,562; 5,264,564; 5,405,938; 5,434,257; 5,466,677; 5,470,967; 5,489,677; 5,541,307; 5,561,225; 5,596,086; 5,602,240; 5,610,289; 5,602,240; 5,608,046; 5,610,289; 5,618,704; 5,623,070; 5,663,312; 5,633,360; 5,677,437; and 5,677,439. [0166] One or more substituted sugar moieties can also be included, e.g., one of the following at the 2' position: OH, SH, SCH₃, F, OCN, OCH₃, OCH₃ O(CH₂)n CH₃, O(CH₂)n NH₂, or O(CH₂)n CH₃, where n is from 1 to about 10; C1 to C10 lower alkyl, alkoxyalkoxy, substituted lower alkyl, alkaryl or aralkyl; Cl; Br; CN; CF3; OCF3; O-, S-, or bi- alkyl; O-, S-, or N-alkenyl; SOCH3; SO2 CH3; ONO2; NO2; N3; NH2; heterocycloalkyl; heterocycloalkaryl; aminoalkylamino; polyalkylamino; substituted silyl; an RNA cleaving group; a reporter group; an intercalator; a group for improving the pharmacokinetic properties of an oligonucleotide; or a group for improving the pharmacodynamic properties of an oligonucleotide and other substituents having similar properties. In some aspects, a modification includes 2'-methoxyethoxy (2'-O-CH₂CH₂OCH₃, also known as 2'-O-(2-methoxyethyl)) (Martin et al., 1995, Helv. Chim. Acta, 78, 486). Other modifications include 2'-methoxy (2'-O-CH₃), 2'-propoxy (2'- OCH₂ CH₂CH₃) and 2'-fluoro (2'- F). Similar modifications can also be made at other positions on the oligonucleotide, particularly the 3' position of the sugar on the 3' terminal nucleotide and the 5' position of 5' terminal nucleotide. Oligonucleotides can also have sugar mimetics, such as cyclobutyls in place of the pentofuranosyl group. [0167] In some examples, both a sugar and an internucleoside linkage (in the backbone) of the nucleotide units can be replaced with novel groups. The base units can be maintained for hybridization with an appropriate nucleic acid target compound. One such oligomeric compound, an oligonucleotide mimetic that has been shown to have excellent hybridization properties, is referred to as a peptide nucleic acid (PNA). In PNA compounds, the sugar- backbone of an oligonucleotide can be replaced with an amide containing backbone, for example, an aminoethylglycine backbone. The nucleobases can be retained and bound directly or indirectly to aza nitrogen atoms of the amide portion of the backbone. Representative U.S. patents that teach the preparation of PNA compounds include, but are not limited to, U.S. Patent Nos.5,539,082; 5,714,331; and 5,719,262. Further teaching of PNA compounds can be found in Nielsen et al., 1991, Science, 254: 1497-1500. [0168] RNAs such as guide RNAs can also include, additionally or alternatively, nucleobase (often referred to in the art simply as "base") modifications or substitutions. As used herein, "unmodified" or "natural" nucleobases include adenine (A), guanine (G), thymine (T), cytosine (C), and uracil (U). Modified nucleobases include nucleobases found only infrequently or transiently in natural nucleic acids, e.g., hypoxanthine, 6-methyladenine, 5-Me pyrimidines, particularly 5- methylcytosine (also referred to as 5-methyl-2' deoxy cytosine and often referred to in the art as 5-Me-C), 5-hydroxymethylcytosine (HMC), glycosyl HMC and gentobiosyl HMC, as well as synthetic nucleobases, e.g., 2-aminoadenine, 2- (methylamino) adenine, 2- (imidazolylalkyl)adenine, 2-(aminoalklyamino) adenine or other heterosub stituted alkyladenines, 2-thiouracil, 2-thiothymine, 5-bromouracil, 5-hydroxymethyluracil, 8-azaguanine, 7- deazaguanine, N6 (6-aminohexyl) adenine, and 2,6-diaminopurine. Komberg, A., DNA Replication, W. H. Freeman & Co., San Francisco, pp.75-77 (1980); Gebeyehu et al., Nucl. Acids Res.15:4513 (1997). A "universal" base known in the art, e.g., inosine, can also be included.5-Me-C substitutions have been shown to increase nucleic acid duplex stability by about 0.6-1.2 °C. (Sanghvi, Y. S., in Crooke, S. T. and Lebleu, B., eds., Antisense Research and Applications, CRC Press, Boca Raton, 1993, pp.276-278) and are aspects of base substitutions. [0169] Modified nucleobases can comprise other synthetic and natural nucleobases, such as 5- methylcytosine (5-me-C), 5-hydroxymethyl cytosine, xanthine, hypoxanthine, 2-aminoadenine, 6-methyl and other alkyl derivatives of adenine and guanine, 2-propyl and other alkyl derivatives of adenine and guanine, 2-thiouracil, 2-thiothymine and 2-thiocytosine, 5-halouracil and cytosine, 5-propynyl uracil and cytosine, 6-azo uracil, cytosine and thymine, 5-uracil (pseudo-uracil), 4-thiouracil, 8-halo, 8-amino, 8- thiol, 8-thioalkyl, 8-hydroxyl and other 8-substituted adenines and guanines, 5-halo particularly 5-bromo, 5-trifluoromethyl and other 5-substituted uracils and cytosines, 7-methylquanine and 7-methyladenine, 8- azaguanine and 8-azaadenine, 7-deazaguanine and 7-deazaadenine, and 3-deazaguanine and 3- deazaadenine. [0170] Further, nucleobases can comprise those disclosed in U.S. Patent No.3,687,808, those disclosed in 'The Concise Encyclopedia of Polymer Science and Engineering', 858-859, Kroschwitz, J.I., ed. John Wiley & Sons, 1990, those disclosed by Englisch et al., Angewandle Chemie, International Edition', 1991, 30, p.613, and those disclosed by Sanghvi, Y. S., Chapter 15, Antisense Research and Applications', 289-302, Crooke, S.T. and Lebleu, B. ea., CRC Press, 1993. Certain of these nucleobases can be useful for increasing the binding affinity of the oligomeric compounds of the invention. These include 5-substituted pyrimidines, 6-azapyrimidines and N-2, N-6 and O-6 substituted purines, comprising 2-aminopropyladenine, 5-propynyluracil and 5-propynylcytosine.5-methylcytosine substitutions have been shown to increase nucleic acid duplex stability by about 0.6-1.2°C (Sanghvi, Y.S., Crooke, S.T. and Lebleu, B., eds, 'Antisense Research and Applications', CRC Press, Boca Raton, 1993, 276-278) and are aspects of base substitutions, even more particularly when combined with 2'-O-methoxyethyl sugar modifications. Modified nucleobases are described in U.S. Patent No.3,687,808, as well as 4,845,205; 5,130,302; 5,134,066; 5,175,273; 5,367,066; 5,432,272; 5,457,187; 5,459,255; 5,484,908; 5,502,177; 5,525,711; 5,552,540; 5,587,469; 5,596,091; 5,614,617; 5,681,941; 5,750,692; 5,763,588; 5,830,653; 6,005,096; and U.S. Patent Application Publication 2003/0158403. [0171] Thus, a modified gRNA can include, for example, one or more non-natural sugars, internucleotide linkages and/or bases. It is not necessary for all positions in a given gRNA to be uniformly modified, and in fact more than one of the aforementioned modifications can be incorporated in a single oligonucleotide, or even in a single nucleoside within an oligonucleotide. [0172] The guide RNAs and/or mRNA (or DNA) encoding an endonuclease can be chemically linked to one or more moieties or conjugates that enhance the activity, cellular distribution, or cellular uptake of the oligonucleotide. Such moieties comprise, but are not limited to, lipid moieties such as a cholesterol moiety (Letsinger et al.1989, Proc. Natl. Acad. Sci. USA, 86: 6553-6556); cholic acid (Manoharan et al, 1994, Bioorg. Med. Chem. Let., 4: 1053- 1060); a thioether, e.g., hexyl-S- tritylthiol (Manoharan et al, 1992, Ann. N. Y. Acad. Sci., 660: 306-309; Manoharan et al., 1993, Bioorg. Med. Chem. Let., 3: 2765- 2770); a thiocholesterol (Oberhauser et al., 1992, Nucl. Acids Res., 20: 533-538); an aliphatic chain, e.g., dodecandiol or undecyl residues (Kabanov et al, 1990, FEBS Lett., 259: 327-330; Svinarchuk et al, 1993, Biochimie, 75: 49- 54); a phospholipid, e.g., di-hexadecyl-rac-glycerol or triethylammonium 1,2-di-O- hexadecyl-rac-glycero-3-H-phosphonate (Manoharan et al., 1995, Tetrahedron Lett., 36: 3651-3654; and Shea et al, 1990, Nucl. Acids Res., 18: 3777-3783); a polyamine or a polyethylene glycol chain (Mancharan et al, 1995, Nucleosides & Nucleotides, 14: 969-973); adamantane acetic acid (Manoharan et al, 1995, Tetrahedron Lett., 36: 3651-3654); a palmityl moiety (Mishra et al., 1995, Biochim. Biophys. Acta, 1264: 229- 237); or an octadecylamine or hexylamino-carbonyl-t oxycholesterol moiety (Crooke et al, 1996, J. Pharmacol. Exp. Ther., 277: 923-937). See also U.S. Patent Nos.4,828,979; 4,948,882; 5,218,105; 5,525,465; 5,541,313; 5,545,730; 5,552,538; 5,578,717; 5,580,731; 5,580,731; 5,591,584; 5,109,124; 5,118,802; 5,138,045; 5,414,077; 5,486,603; 5,512,439; 5,578,718; 5,608,046; 4,587,044; 4,605,735; 4,667,025; 4,762,779; 4,789,737; 4,824,941; 4,835,263; 4,876,335; 4,904,582; 4,958,013; 5,082,830; 5,112,963; 5,214,136; 5,082,830; 5,112,963; 5,214,136; 5,245,022; 5,254,469; 5,258,506; 5,262,536; 5,272,250; 5,292,873; 5,317,098; 5,371,241; 5,391,723; 5,416,203; 5,451,463; 5,510,475; 5,512,667; 5,514,785; 5,565,552; 5,567,810; 5,574,142; 5,585,481; 5,587,371; 5,595,726; 5,597,696; 5,599,923; 5,599, 928 and 5,688,941. [0173] Sugars and other moieties can be used to target proteins and complexes comprising nucleotides, such as cationic polysomes and liposomes, to particular sites. For example, hepatic cell directed transfer can be mediated via asialoglycoprotein receptors (ASGPRs); see, e.g., Hu, et al., 2014, Protein Pept Lett.21(10):1025-30. Other systems known in the art and regularly developed can be used to target biomolecules of use in the present case and/or complexes thereof to particular target cells of interest. [0174] Targeting moieties or conjugates can include conjugate groups covalently bound to functional groups, such as primary or secondary hydroxyl groups. Conjugate groups of the present disclosure include intercalators, reporter molecules, polyamines, polyamides, polyethylene glycols, polyethers, groups that enhance the pharmacodynamic properties of oligomers, and groups that enhance the pharmacokinetic properties of oligomers. Typical conjugate groups include cholesterols, lipids, phospholipids, biotin, phenazine, folate, phenanthridine, anthraquinone, acridine, fluoresceins, rhodamines, coumarins, and dyes. Groups that enhance the pharmacodynamic properties, in the context of this present disclosure, include groups that improve uptake, enhance resistance to degradation, and/or strengthen sequence-specific hybridization with the target nucleic acid. Groups that enhance the pharmacokinetic properties, in the context of this disclosure, include groups that improve uptake, distribution, metabolism or excretion of the compounds of the present disclosure. Representative conjugate groups are disclosed in International Patent Application Publication WO1993007883, and U.S. Patent No.6,287,860. Conjugate moieties include, but are not limited to, lipid moieties such as a cholesterol moiety, cholic acid, a thioether, e.g., hexyl-5 -trityl thiol, a thiocholesterol, an aliphatic chain, e.g., dodecandiol or undecyl residues, a phospholipid, e.g., di-hexadecyl-rac- glycerol or triethylammonium 1,2-di-O-hexadecyl-rac-glycero-3-H-phosphonate, a polyamine or a polyethylene glycol chain, or adamantane acetic acid, a palmityl moiety, or an octadecylamine or hexylamino-carbonyl- oxy cholesterol moiety. See, e.g., U.S. Pat. Nos.4,828,979; 4,948,882; 5,218,105; 5,525,465; 5,541,313; 5,545,730; 5,552,538; 5,578,717, 5,580,731; 5,580,731; 5,591,584; 5,109,124; 5,118,802; 5,138,045; 5,414,077; 5,486,603; 5,512,439; 5,578,718; 5,608,046; 4,587,044; 4,605,735; 4,667,025; 4,762,779; 4,789,737; 4,824,941; 4,835,263; 4,876,335; 4,904,582; 4,958,013; 5,082,830; 5,112,963; 5,214,136; 5,082,830; 5,112,963; 5,214,136; 5,245,022; 5,254,469; 5,258,506; 5,262,536; 5,272,250; 5,292,873; 5,317,098; 5,371,241; 5,391,723; 5,416,203, 5,451,463; 5,510,475; 5,512,667; 5,514,785; 5,565,552; 5,567,810; 5,574,142; 5,585,481; 5,587,371; 5,595,726; 5,597,696; 5,599,923; 5,599,928 and 5,688,941. [0175] A large variety of modifications have been developed and applied to enhance RNA stability, reduce innate immune responses, and/or achieve other benefits that can be useful in connection with the introduction of polynucleotides into human cells, as described herein; see, e.g., the reviews by Whitehead KA et al., 2011, Annual Review of Chemical and Biomolecular Engineering, 2: 77-96; Gaglione and Messere, 2010, Mini Rev Med Chem, 10(7):578-95; Chernolovskaya et al, 2010, Curr Opin Mol Ther., 12(2): 158-67; Deleavey et al., 2009, Curr Protoc Nucleic Acid Chem Chapter 16:Unit 16.3; Behlke, 2008, Oligonucleotides 18(4):305-19; Fucini et al, 2012, Nucleic Acid Ther 22(3): 205-210; Bremsen et al, 2012, Front Genet 3: 154. 6.4. Systems [0176] The disclosure provides systems comprising a Type II Cas protein of the disclosure (e.g., as described in Section 6.2) and a means for targeting the Type II Cas protein to a target genomic sequence. The means for targeting the Type II Cas protein to a target genomic sequence can be a guide RNA (gRNA) (e.g., as described in Section 6.3). [0177] The disclosure also provides systems comprising a Type II Cas protein of the disclosure (e.g., as described in Section 6.2) and a gRNA (e.g., as described in Section 6.3). The systems can comprise a ribonucleoprotein particle (RNP) in which a Type II Cas protein is complexed with a gRNA, for example a sgRNA or separate crRNA and tracrRNA. Systems of the disclosure can in some embodiments further comprise genomic DNA complexed with the Type II Cas protein and the gRNA. Accordingly, the disclosure provides systems comprising a Type II Cas protein, a genomic DNA, and gRNA, all complexed with one another. [0178] The systems of the disclosure can exist within a cell (whether the cell is in vivo, ex vivo, or in vitro) or outside a cell (e.g., in a particle our outside of a particle). 6.5. Nucleic Acids [0179] The disclosure provides nucleic acids (e.g., DNA or RNA) encoding Type II Cas proteins (e.g., CUAZ Type II Cas proteins, AHWY Type II Cas proteins, DGHJ Type II Cas proteins, CBGI Type II Cas proteins, ASDR Type II Cas proteins, BCZZ Type II Cas proteins, DRCY Type II Cas proteins, DRPY Type II Cas proteins, EAJR Type II Cas proteins, and CFDE Type II Cas proteins), nucleic acids encoding gRNAs of the disclosure, nucleic acids encoding both Type II Cas proteins and gRNAs, and pluralities of nucleic acids, for example comprising a nucleic acid encoding a Type II Cas protein and a gRNA. [0180] A nucleic acid encoding a Type II Cas protein and/or gRNA can be, for example, a plasmid or a viral genome (e.g., a lentivirus, retrovirus, adenovirus, or adeno-associated virus genome). Plasmids can be, for example, plasmids for producing virus particles, e.g., lentivirus particles, or plasmids for propagating the Type II Cas and gRNA coding sequences in bacterial (e.g., E. coli) or eukaryotic (e.g., yeast) cells. [0181] A nucleic acid encoding a Type II Cas protein can, in some embodiments, further encode a gRNA. Alternatively, a gRNA can be encoded by a separate nucleic acid (e.g., DNA or mRNA). [0182] Nucleic acids encoding a Type II Cas protein can be codon optimized, e.g., where at least one non-common codon or less-common codon has been replaced by a codon that is common in a host cell. For example, a codon optimized nucleic acid can direct the synthesis of an optimized messenger mRNA, e.g., optimized for expression in a mammalian expression system. As an example, if the intended target nucleic acid is within a human cell, a human codon-optimized polynucleotide encoding Type II Cas can be used for producing a Type II Cas polypeptide. Exemplary codon-optimized sequences are shown in Tables 1A-1K. [0183] Nucleic acids of the disclosure, e.g., plasmids and viral vectors, can comprise one or more regulatory elements such as promoters, enhancers, and other expression control elements (e.g., transcription termination signals, such as polyadenylation signals and poly-U sequences). Such regulatory elements are described, for example, in Goeddel, 1990, GENE EXPRESSION TECHNOLOGY: METHODS IN ENZYMOLOGY 185, Academic Press, San Diego, Calif. Regulatory elements include those that direct constitutive expression of a nucleotide sequence in many types of host cell and those that direct expression of the nucleotide sequence only in certain host cells (e.g., tissue- specific regulatory sequences). A tissue-specific promoter may direct expression primarily in a desired tissue of interest or in particular cell types. Regulatory elements may also direct expression in a temporal- dependent manner, such as in a cell-cycle dependent or developmental stage-dependent manner, which may or may not also be tissue or cell-type specific. In some embodiments, a nucleic acid of the disclosure comprises one or more pol III promoter (e.g., 1, 2, 3, 4, 5, or more pol III promoters), one or more pol II promoters (e.g., 1, 2, 3, 4, 5, or more pol II promoters), one or more pol I promoters (e.g., 1, 2, 3, 4, 5, or more pol I promoters), or combinations thereof, e.g., to express a Type II Cas protein and a gRNA separately. Examples of pol III promoters include, but are not limited to, U6 and H1 promoters. Examples of pol II promoters include, but are not limited to, the retroviral Rous Sarcoma virus (RSV) LTR promoter (optionally with the RSV enhancer), the cytomegalovirus (CMV) promoter (optionally with the CMV enhancer) (see, e.g., Boshart et al, 1985, Cell 41:521-530), the SV40 promoter, the dihydrofolate reducWDVH^SURPRWHU^^WKH^ȕ-DFWLQ^SURPRWHU^^WKH^SKRVSKRJO\FHURO^NLQDVH^^3*.^^SURPRWHU^^DQG^()^Į^ SURPRWHUV^^IRU^H[DPSOH^^IXOO^OHQJWK^()^Į^SURPRWHU^DQG^WKH^()6^SURPRWHU^^ZKLFK^LV^D^VKRUW^^LQWURQ-less IRUP^RI^WKH^IXOO^()^Į^SURPRWHU^^^([HPSODU\^HQKDQFHU^HOHPHQWV^LQFOude WPRE; CMV enhancers; the R- 8^ƍ^VHJPHQW^LQ^/75^RI^+7/9-I; SV40 enhancer; and the intron sequence between exons 2 and 3 of UDEELW^ȕ-globin. It will be appreciated by those skilled in the art that the design of an expression vector can depend on such factors as the choice of the host cell, the level of expression desired, etc. [0184] The term "vector" refers to a polynucleotide molecule capable of transporting another nucleic acid to which it has been linked. One type of polynucleotide vector includes a "plasmid", which refers to a circular double-stranded DNA loop into which additional nucleic acid segments are or can be ligated. Another type of polynucleotide vector is a viral vector; wherein additional nucleic acid segments can be ligated into the viral genome. Certain vectors are capable of autonomous replication in a host cell into which they are introduced (e.g., bacterial vectors having a bacterial origin of replication and episomal mammalian vectors). Other vectors (e.g., non-episomal mammalian vectors) are integrated into the genome of a host cell upon introduction into the host cell, and thereby are replicated along with the host genome. [0185] In some examples, vectors can be capable of directing the expression of nucleic acids to which they are operably linked. Such vectors can be referred to herein as "recombinant expression vectors", or more simply "expression vectors", which serve equivalent functions. [0186] The term "operably linked" means that the nucleotide sequence of interest is linked to regulatory sequence(s) in a manner that allows for expression of the nucleotide sequence. The term "regulatory sequence" is intended to include, for example, promoters, enhancers and other expression control elements (e.g., polyadenylation signals). Such regulatory sequences are well known in the art and are described, for example, in Goeddel; Gene Expression Technology: Methods in Enzymology 185, Academic Press, San Diego, CA (1990). Regulatory sequences include those that direct constitutive expression of a nucleotide sequence in many types of host cells, and those that direct expression of the nucleotide sequence only in certain host cells (e.g., tissue-specific regulatory sequences). It will be appreciated by those skilled in the art that the design of the expression vector can depend on such factors as the choice of the target cell, the level of expression desired, and the like. [0187] Vectors can include, but are not limited to, viral vectors based on vaccinia virus, poliovirus, adenovirus, adeno-associated virus (e.g., AAV2, AAV5, AAV7m8, AAV8, AAV9, AAVrh8r, AAVrh10), SV40, herpes simplex virus, human immunodeficiency virus, retrovirus (e.g., Murine Leukemia Virus, spleen necrosis virus, and vectors derived from retroviruses such as Rous Sarcoma Virus, Harvey Sarcoma Virus, avian leukosis virus, a lentivirus, human immunodeficiency virus, myeloproliferative sarcoma virus, and mammary tumor virus) and other recombinant vectors. Other vectors contemplated for eukaryotic target cells include, but are not limited to, the vectors pXTl, pSG5, pSVK3, pBPV, pMSG, and pSVLSV40 (Pharmacia). Additional vectors contemplated for eukaryotic target cells include, but are not limited to, the vectors pCTx-l, pCTx-2, and pCTx-3. Other vectors can be used so long as they are compatible with the host cell. [0188] In some examples, a vector can comprise one or more transcription and/or translation control elements. Depending on the host/vector system utilized, any of a number of suitable transcription and translation control elements, including constitutive and inducible promoters, transcription enhancer elements, transcription terminators, etc. can be used in the expression vector. The vector can be a self- inactivating vector that either inactivates the viral sequences or the components of the CRISPR machinery or other elements. [0189] Non-limiting examples of suitable eukaryotic promoters (promoters functional in a eukaryotic cell) include those from cytomegalovirus (CMV) immediate early, herpes simplex virus (HSV) thymidine kinase, early and late SV40, long terminal repeats (LTRs) from retrovirus, human elongation factor-l SURPRWHUV^^IRU^H[DPSOH^^WKH^IXOO^()^Į^SURPRWHU^DQG^WKH^()6^SURPRWHU^^^D^K\EULG^FRQVWUXFW^FRPSULVLQJ^WKH^ cytomegalovirus (CMV) enhancer fused to the chicken beta-actin promoter (CAG), murine stem cell virus promoter (MSCV), phosphoglycerate kinase-1 locus promoter (PGK), and mouse metallothionein-I. [0190] An expression vector can also contain a ribosome binding site for translation initiation and a transcription terminator. The expression vector can also comprise appropriate sequences for amplifying expression. The expression vector can also include nucleotide sequences encoding non-native tags (e.g., histidine tag, hemagglutinin tag, green fluorescent protein, etc.) that are fused to the site-directed polypeptide, thus resulting in a fusion protein. [0191] A promoter can be an inducible promoter (e.g., a heat shock promoter, tetracycline- regulated promoter, steroid-regulated promoter, metal-regulated promoter, estrogen receptor-regulated promoter, etc.). The promoter can be a constitutive promoter (e.g., CMV promoter, UBC promoter). In some cases, the promoter can be a spatially restricted and/or temporally restricted promoter (e.g., a tissue specific promoter, for example a human RHO promoter or human rhodopsin kinase promoter (hGRK), a cell type specific promoter, etc.). 6.6. Particles and Cells [0192] The disclosure further provides particles comprising a Type II Cas protein of the disclosure (e.g., a DGHJ Type II Cas protein, a CUAZ Type II Cas protein, an AHWY Type II Cas protein, a CBGI Type II Cas protein, an ASDR Type II Cas protein, a BCZZ Type II Cas protein, a DRCY Type II Cas protein, a DRPY Type II Cas protein, an EAJR Type II Cas protein, or a CFDE Type II Cas protein), particles comprising a gRNA of the disclosure, particles comprising a system of the disclosure, and particles comprising a nucleic acid or plurality of nucleic acids of the disclosure. The particles can in some embodiments comprise or further comprise a gRNA, or a nucleic acid encoding the gRNA (e.g., DNA or mRNA). For example, the particles can comprise a RNP of the disclosure. Exemplary particles include lipid nanoparticles, vesicles, viral-like particles (VLPs) and gold nanoparticles. See, e.g., WO 2020/012335, the contents of which are incorporated herein by reference in their entireties, which describes vesicles that can be used to deliver gRNA molecules and Type II Cas proteins to cells (e.g., complexed together as a RNP). [0193] The disclosure provides particles (e.g., virus particles) comprising a nucleic acid encoding a Type II Cas protein of the disclosure. The particles can further comprise a nucleic acid encoding a gRNA. Alternatively, a nucleic acid encoding a Type II Cas protein can further encode a gRNA. [0194] The disclosure further provides pluralities of particles (e.g., pluralities of virus particles). Such pluralities can include a particle encoding a Type II Cas protein and a different particle encoding a gRNA. For example, a plurality of particles can comprise a virus particle (e.g., an AAV2, AAV5, AAV7m8, AAV8, AAV9, AAVrh8r, or AAVrh10 virus particle) encoding a Type II Cas protein and a second virus particle (e.g., an AAV2, AAV5, AAV7m8, AAV8, AAV9, AAVrh8r, or AAVrh10 virus particle) encoding a gRNA. Alternatively, a plurality of particles can comprise a plurality of virus particles where each particle encodes a Type II Cas protein and a gRNA. [0195] The disclosure further provides cells and populations of cells (e.g., ex vivo cells and populations of cells) that can comprise a Type II Cas protein (e.g., introduced to the cell as a RNP) or a nucleic acid encoding the Type II Cas protein (e.g., DNA or mRNA) (optionally also encoding a gRNA). The disclosure further provides cells and populations of cells comprising a gRNA of the disclosure (optionally complexed with a Type II Cas protein) or a nucleic acid encoding the gRNA (e.g., DNA or mRNA) (optionally also encoding a Type II Cas protein). The cells and populations of cells can be, for example, human cells such as a stem cell, e.g., a hematopoietic stem cell (HSC), a pluripotent stem cell, an induced pluripotent stem cell (iPS), or an embryonic stem cell. Methods for introducing proteins and nucleic acids to cells are known in the art. For example, a RNP can be produced by mixing a Type II Cas protein and one or more guide RNAs in an appropriate buffer. An RNP can be introduced to a cell, for example, via electroporation and other methods known in the art. [0196] The cell populations of the disclosure can be cells in which gene editing by the systems of the disclosure has taken place, or cells in which the components of a system of the disclosure have been introduced or expressed but gene editing has not taken place, or a combination thereof. A cell population can comprise, for example, a population in which at least 1%, at least 5%, at least 10%, at least 15%, at least 20%, at least 30%, at least 40%, at least 50%, at least 60%, or at least 70% of the cells have undergone gene editing by a system of the disclosure. 6.7. Pharmaceutical Compositions [0197] Also disclosed herein are pharmaceutical formulations and medicaments comprising a Type II Cas protein, gRNA, nucleic acid or plurality of nucleic acids, system, particle, or plurality of particles of the disclosure together with a pharmaceutically acceptable excipient. [0198] Suitable excipients include, but are not limited to, salts, diluents, (e.g., Tris-HCl, acetate, phosphate), preservatives (e.g., Thimerosal, benzyl alcohol, parabens), binders, fillers, solubilizers, disintegrants, sorbents, solvents, pH modifying agents, antioxidants, antinfective agents, suspending agents, wetting agents, viscosity modifiers, tonicity agents, stabilizing agents, and other components and combinations thereof. Suitable pharmaceutically acceptable excipients can be selected from materials which are generally recognized as safe (GRAS), and may be administered to an individual without causing undesirable biological side effects or unwanted interactions. Suitable excipients and their formulations are described in Remington's Pharmaceutical Sciences, 16th ed.1980, Mack Publishing Co. In addition, such compositions can be complexed with polyethylene glycol (PEG), metal ions, or incorporated into polymeric compounds such as polyacetic acid, polyglycolic acid, hydrogels, etc., or incorporated into liposomes, microemulsions, micelles, unilamellar or multilamellar vesicles, erythrocyte ghosts or spheroblasts. Suitable dosage forms for administration, e.g., parenteral administration, include solutions, suspensions, and emulsions. [0199] The components of the pharmaceutical formulation can be dissolved or suspended in a suitable solvent such as, for example, water, Ringer's solution, phosphate buffered saline (PBS), or isotonic sodium chloride. The formulation may also be a sterile solution, suspension, or emulsion in a nontoxic, parenterally acceptable diluent or solvent such as 1,3-butanediol. [0200] In some cases, formulations can include one or more tonicity agents to adjust the isotonic range of the formulation. Suitable tonicity agents are well known in the art and include glycerin, mannitol, sorbitol, sodium chloride, and other electrolytes. In some cases, the formulations can be buffered with an effective amount of buffer necessary to maintain a pH suitable for parenteral administration. Suitable buffers are well known by those skilled in the art and some examples of useful buffers are acetate, borate, carbonate, citrate, and phosphate buffers. [0201] In some embodiments, the formulation can be distributed or packaged in a liquid form, or alternatively, as a solid, obtained, for example by lyophilization of a suitable liquid formulation, which can be reconstituted with an appropriate carrier or diluent prior to administration. In some embodiments, the formulations can comprise a guide RNA and a Type II Cas protein in a pharmaceutically effective amount sufficient to edit a gene in a cell. The pharmaceutical compositions can be formulated for medical and/or veterinary use. 6.8. Methods of Altering a Cell [0202] The disclosure further provides methods of using the Type II Cas proteins, gRNAs, nucleic acids (including pluralities of nucleic acids), systems, and particles (including pluralities of particles) of the disclosure for altering cells. [0203] In one aspect, a method of altering a cell comprises contacting a eukaryotic cell (e.g., a human cell) with a nucleic acid, particle, system or pharmaceutical composition described herein. [0204] Contacting a cell with a disclosed nucleic acid, particle, system or pharmaceutical composition can be achieved by any method known in the art and can be performed in vivo, ex vivo, or in vitro. In some embodiments, the methods can include obtaining one or more cells from a subject prior to contacting the cell(s) with a herein disclosed nucleic acid, particle, system or pharmaceutical composition. In some embodiments, the methods can further comprise returning or implanting the contacted cell or a progeny thereof to the subject. [0205] Type II Cas and gRNA, as well as nucleic acids encoding Type II Cas and gRNAs can be delivered to a cell by any means known in the art, for example, by viral or non-viral delivery vehicles, electroporation or lipid nanoparticles. [0206] A polynucleotide encoding Type II Cas and a gRNA, can be delivered to a cell (ex vivo or in vivo) by a lipid nanoparticle (LNP). LNPs can have, for example, a diameter of less than 1000 nm, 500 nm, 250 nm, 200 nm, 150 nm, 100 nm, 75 nm, 50 nm, or 25 nm. Alternatively, a nanoparticle can range in size from 1-1000 nm, 1-500 nm, 1-250 nm, 25-200 nm, 25-100 nm, 35-75 nm, or 25-60 nm. LNPs can be made from cationic, anionic, neutral lipids, and combinations thereof. Neutral lipids, such as the fusogenic phospholipid DOPE or the membrane component cholesterol, can be included in LNPs as 'helper lipids' to enhance transfection activity and nanoparticle stability. [0207] LNPs can also be comprised of hydrophobic lipids, hydrophilic lipids, or both hydrophobic and hydrophilic lipids. Lipids and combinations of lipids that are known in the art can be used to produce a LNP. Examples of lipids used to produce LNPs are: DOTMA, DOSPA, DOTAP, DMRIE, DC- cholesterol, DOTAP-cholesterol, GAP-DMORIE-DPyPE, and GL67A-DOPE-DMPE- polyethylene glycol (PEG). Examples of cationic lipids are: 98N12-5, C12-200, DLin-KC2- DMA (KC2), DLin-MC3-DMA (MC3), XTC, MD1, and 7C1. Examples of neutral lipids are: DPSC, DPPC, POPC, DOPE, and SM. Examples of PEG- modified lipids are: PEG-DMG, PEG- CerCl4, and PEG-CerC20. Lipids can be combined in any number of molar ratios to produce a LNP. In addition, the polynucleotide(s) can be combined with lipid(s) in a wide range of molar ratios to produce a LNP. [0208] Type II Cas and/or gRNAs can be delivered to a cell via an adeno-associated viral vector (e.g., of an AAV2, AAV5, AAV7m8, AAV8, AAV9, AAVrh8r, or AAVrh10 serotype), or by another viral vector. Other viral vectors include, but are not limited to lentivirus, adenovirus, alphavirus, enterovirus, pestivirus, baculovirus, herpesvirus, Epstein Barr virus, papovavirus, poxvirus, vaccinia virus, and herpes simplex virus. In some embodiments, a Type II Cas mRNA is formulated in a lipid nanoparticle, while a sgRNA is delivered to a cell in an AAV or other viral vector. In some embodiments, one or more AAV vectors (e.g., one or more AAV2, AAV5, AAV7m8, AAV8, AAV9, AAVrh8r, or AAVrh10 serotype) are used to deliver both a sgRNA and a Type II Cas. In some embodiments, a Type II Cas and a sgRNA are delivered using separate vectors. In other embodiments, a Type II Cas and a sgRNA are delivered using a single vector. [0209] Compositions and methods for delivering Type II Cas and gRNAs to a cell and/or subject are further described in PCT Patent Application Publications WO 2019/102381, WO 2020/012335, and WO 2020/053224, each of which is incorporated by reference herein in its entirety. [0210] DNA cleavage can result in a single-strand break (SSB) or double-strand break (DSB) at particular locations within the DNA molecule. Such breaks can be and regularly are repaired by natural, endogenous cellular processes, such as homology-dependent repair (HDR) and non-homologous end- joining (NHEJ). These repair processes can edit the targeted polynucleotide by introducing a mutation, WKHUHE\^UHVXOWLQJ^LQ^D^SRO\QXFOHRWLGH^KDYLQJ^D^VHTXHQFH^ZKLFK^GLIIHUV^IURP^WKH^SRO\QXFOHRWLGH¶V^VHTXHQFH^ prior to cleavage by a Type II Cas. [0211] NHEJ and HDR DNA repair processes consist of a family of alternative pathways. Non- homologous end-joining (NHEJ) refers to the natural, cellular process in which a double-stranded DNA- break is repaired by the direct joining of two non-homologous DNA segments. See, e.g. Cahill et al., 2006, Front. Biosci.11:1958-1976. DNA repair by non-homologous end-joining is error-prone and frequently results in the untemplated addition or deletion of DNA sequences at the site of repair. Thus, NHEJ repair mechanisms can introduce mutations into the coding sequence which can disrupt gene function. NHEJ directly joins the DNA ends resulting from a double-strand break, sometimes with a modification of the polynucleotide sequence such as a loss of or addition of nucleotides in the polynucleotide sequence. The modification of the polynucleotide sequence can disrupt (or perhaps enhance) gene expression. [0212] Homology-dependent repair (HDR) utilizes a homologous sequence, or donor sequence, as a template for inserting a defined DNA sequence at the break point. The homologous sequence can be in the endogenous genome, such as a sister chromatid. Alternatively, the donor can be an exogenous nucleic acid, such as a plasmid, a single-strand oligonucleotide, a double- stranded oligonucleotide, a duplex oligonucleotide or a virus, that has regions of high homology with the nuclease-cleaved locus, but which can also contain additional sequence or sequence changes including deletions that can be incorporated into the cleaved target locus. [0213] A third repair mechanism includes microhomology-mediated end joining (MMEJ), also referred to DV^³$OWHUQDWLYH^1+(-^^$1+(-^´^^LQ^ZKLFK^WKH^JHQHWLF^RXWFRPH^LV^VLPLODU^WR^1+(-^LQ^WKDW^VPDOO^GHOHWLRQV^ and insertions can occur at the cleavage site. MMEJ can make use of homologous sequences of a few base pairs flanking the DNA break site to drive a more favored DNA end joining repair outcome. In some instances, it may be possible to predict likely repair outcomes based on analysis of potential microhomologies at the site of the DNA break. [0214] Modifications of a cleaved polynucleotide by HDR, NHEJ, and/or ANHEJ can result in, for example, mutations, deletions, alterations, integrations, gene correction, gene replacement, gene tagging, transgene insertion, nucleotide deletion, gene disruption, translocations and/or gene mutation. The aforementioned process outcomes are examples of editing a polynucleotide. [0215] Advantages of ex vivo cell therapy approaches include the ability to conduct a comprehensive analysis of the therapeutic prior to administration. Nuclease-based therapeutics can have some level of off-target effects. Performing gene correction ex vivo allows a method user to characterize the corrected cell population prior to implantation, including identifying any undesirable off-target effects. Where undesirable effects are observed, a method user may opt not to implant the cells or cell progeny, may further edit the cells, or may select new cells for editing and analysis. Other advantages include ease of genetic correction in iPSCs compared to other primary cell sources. iPSCs are prolific, making it easy to obtain the large number of cells that will be required for a cell-based therapy. Furthermore, iPSCs are an ideal cell type for performing clonal isolations. This allows screening for the correct genomic correction, without risking a decrease in viability. [0216] Although certain cells present an attractive target for ex vivo treatment and therapy, increased efficacy in delivery may permit direct in vivo delivery to such cells. Ideally the targeting and editing is directed to the relevant cells. Cleavage in other cells can also be prevented by the use of promoters only active in certain cell types and/or developmental stages. [0217] Additional promoters are inducible, and therefore can be temporally controlled if the nuclease is delivered as a plasmid. The amount of time that delivered protein and RNA remain in the cell can also be adjusted using treatments or domains added to change the half-life. In vivo treatment would eliminate a number of treatment steps, but a lower rate of delivery can require higher rates of editing. In vivo treatment can eliminate problems and losses from ex vivo treatment and engraftment. [0218] An advantage of in vivo gene therapy can be the ease of therapeutic production and administration. The same therapeutic approach and therapy has the potential to be used to treat more than one patient, for example a number of patients who share the same or similar genotype or allele. In contrast, ex vivo cell therapy typically requires using D^VXEMHFW¶V^RZQ^FHOOV^^ZKLFK^DUH^LVRODWHG^^ manipulated and returned to the same patient. [0219] Progenitor cells (also referred to as stem cells herein) are capable of both proliferation and giving rise to more progenitor cells, which in turn have the ability to generate a large number of cells that can in turn give rise to differentiated or differentiable daughter cells. The daughter cells themselves can be induced to proliferate and produce progeny that subsequently differentiate into one or more mature cell types, while also retaining one or more cells with parental developmental potential. The term "stem cell" refers then to a cell with the capacity or potential, under particular circumstances, to differentiate to a more specialized or differentiated phenotype, and which retains the capacity, under certain circumstances, to proliferate without substantially differentiating. In one aspect, the term progenitor or stem cell refers to a generalized mother cell whose descendants (progeny) specialize, often in different directions, by differentiation, e.g., by acquiring completely individual characters, as occurs in progressive diversification of embryonic cells and tissues. Cellular differentiation is a complex process typically occurring through many cell divisions. A differentiated cell can derive from a multipotent cell that itself is derived from a multipotent cell, and so on. While each of these multipotent cells can be considered stem cells, the range of cell types that each can give rise to can vary considerably. Some differentiated cells also have the capacity to give rise to cells of greater developmental potential. Such capacity can be natural or can be induced artificially upon treatment with various factors. In many biological instances, stem cells can also be "multipotent" because they can produce progeny of more than one distinct cell type, but this is not required. [0220] Human cells described herein can be induced pluripotent stem cells (iPSCs). An advantage of using iPSCs in the methods of the disclosure is that the cells can be derived from the same subject to which the progenitor cells are to be administered. That is, a somatic cell can be obtained from a subject, reprogrammed to an induced pluripotent stem cell, and then differentiated into a progenitor cell to be administered to the subject (e.g., an autologous cell). Because progenitors are essentially derived from an autologous source, the risk of engraftment rejection or allergic response can be reduced compared to the use of cells from another subject or group of subjects. In addition, the use of iPSCs negates the need for cells obtained from an embryonic source. Thus, in one aspect, the stem cells used in the disclosed methods are not embryonic stem cells. [0221] Methods are known in the art that can be used to generate pluripotent stem cells from somatic cells. Pluripotent stem cells generated by such methods can be used in the method of the disclosure. [0222] Reprogramming methodologies for generating pluripotent cells using defined combinations of transcription factors have been described. Mouse somatic cells can be converted to ES cell-like cells with expanded developmental potential by the direct transduction of Oct4, Sox2, Klf4, and c-Myc; see, e.g., Takahashi and Yamanaka, 2006, Cell 126(4): 663-76. iPSCs resemble ES cells, as they restore the pluripotency-associated transcriptional circuitry and much of the epigenetic landscape. In addition, mouse iPSCs satisfy all the standard assays for pluripotency: specifically, in vitro differentiation into cell types of the three germ layers, teratoma formation, contribution to chimeras, germline transmission (see, e.g., Maherali and Hochedlinger, 2008, Cell Stem Cell.3(6):595-605), and tetraploid complementation. [0223] Human iPSCs can be obtained using similar transduction methods, and the transcription factor trio, OCT4, SOX2, and NANOG, has been established as the core set of transcription factors that govern pluripotency; see, e.g., 2014, Budniatzky and Gepstein, Stem Cells Transl Med.3(4):448-57; Barrett et al, 2014, Stem Cells Trans Med 3: 1-6 sctm.2014-0121; Focosi et al, 2014, Blood Cancer Journal 4: e211. The production of iPSCs can be achieved by the introduction of nucleic acid sequences encoding stem cell-associated genes into an adult, somatic cell, historically using viral vectors. [0224] iPSCs can be generated or derived from terminally differentiated somatic cells, as well as from adult stem cells, or somatic stem cells. That is, a non-pluripotent progenitor cell can be rendered pluripotent or multipotent by reprogramming. In such instances, it may not be necessary to include as many reprogramming factors as required to reprogram a terminally differentiated cell. Further, reprogramming can be induced by the non-viral introduction of reprogramming factors, e.g., by introducing the proteins themselves, or by introducing nucleic acids that encode the reprogramming factors, or by introducing messenger RNAs that upon translation produce the reprogramming factors (see e.g., Warren et al., 2010, Cell Stem Cell, 7(5):6l8- 30. Reprogramming can be achieved by introducing a combination of nucleic acids encoding stem cell-associated genes, including, for example, Oct-4 (also known as Oct-3/4 or Pouf5l), Soxl, Sox2, Sox3, Sox 15, Sox 18, NANOG, Klfl, Klf2, Klf4, Klf5, NR5A2, c- Myc, 1- Myc, n-Myc, Rem2, Tert, and LIN28. Reprogramming using the methods and compositions described herein can further comprise introducing one or more of Oct-3/4, a member of the Sox family, a member of the Klf family, and a member of the Myc family to a somatic cell. The methods and compositions described herein can further comprise introducing one or more of each of Oct-4, Sox2, Nanog, c-MYC and Klf4 for reprogramming. As noted above, the exact method used for reprogramming is not necessarily critical to the methods and compositions described herein. However, where cells differentiated from the reprogrammed cells are to be used in, e.g., human therapy, in one aspect the reprogramming is not affected by a method that alters the genome. Thus, in such examples, reprogramming can be achieved, e.g., without the use of viral or plasmid vectors. [0225] Efficiency of reprogramming (the number of reprogrammed cells) derived from a population of starting cells can be enhanced by the addition of various agents, e.g., small molecules, as shown by Shi et al., 2008, Cell-Stem Cell 2:525-528; Huangfu et al., 2008, Nature Biotechnology 26(7):795-797; and Marson et al., 2008, Cell-Stem Cell 3: 132-135. Thus, an agent or combination of agents that enhance the efficiency or rate of induced pluripotent stem cell production can be used in the production of patient- specific or disease-specific iPSCs. Some non-limiting examples of agents that enhance reprogramming efficiency include soluble Wnt, Wnt conditioned media, BIX-01294 (a G9a histone methyltransferase), PD0325901 (a MEK inhibitor), DNA methyltransferase inhibitors, histone deacetylase (HD AC) inhibitors, valproic acid, 5'-azacytidine, dexamethasone, suberoylanilide, hydroxamic acid (SAHA), vitamin C, and trichostatin (TSA), among others. Other non-limiting examples of reprogramming enhancing agents include: Suberoylanilide Hydroxamic Acid (SAHA ( e.g ., MK0683, vorinostat) and other hydroxamic acids), BML-210, Depudecin (e.g., (-)-Depudecin), HC Toxin, Nullscript (4-(l,3-Dioxo-lH,3H- benzo[de]isoquinolin-2-yl)-N-hydroxybutanamide), Phenylbutyrate (e.g., sodium phenylbutyrate) and Valproic Acid ((VP A) and other short chain fatty acids), Scriptaid, Suramin Sodium, Trichostatin A (TSA), APHA Compound 8, Apicidin, Sodium Butyrate, pi valoyloxy methyl butyrate (Pivanex, AN-9), Trapoxin B, Chlamydocin, Depsipeptide (also known as FR901228 or FK228), benzamides (e.g., CI-994 (e.g., N- acetyl dinaline) and MS-27- 275), MGCD0103, NVP-LAQ-824, CBHA (m-carboxycinnaminic acid bishydroxamic acid), JNJ16241199, Tubacin, A-161906, proxamide, oxamflatin, 3-C1-UCHA (e.g., 6-(3- chlorophenylureido)caproic hydroxamic acid), AOE (2-amino-8-oxo-9, 10-epoxy decanoic acid), CHAP31 and CHAP 50. Other reprogramming enhancing agents include, for example, dominant negative forms of the HDACs (e.g, catalytically inactive forms), siRNA inhibitors of the HDACs, and antibodies that specifically bind to the HDACs. Such inhibitors are available, e.g., from BIOMOL International, Fukasawa, Merck Biosciences, Novartis, Gloucester Pharmaceuticals, Titan Pharmaceuticals, MethylGene, and Sigma Aldrich. [0226] To confirm the induction of pluripotent stem cells, isolated clones can be tested for the expression of a stem cell marker. Such expression in a cell derived from a somatic cell identifies the cells as induced pluripotent stem cells. Stem cell markers can be selected from the non-limiting group including SSEA3, SSEA4, CD9, Nanog, Fbxl5, Ecatl, Esgl, Eras, Gdfi, Fgf4, Cripto, Daxl, Zpf296, Slc2a3, Rexl, Utfl, and Natl. In one case, for example, a cell that expresses Oct4 or Nanog is identified as pluripotent. Methods for detecting the expression of such markers can include, for example, RT-PCR and immunological methods that detect the presence of the encoded polypeptides, such as Western blots or flow cytometric analyses. Detection can involve not only RT-PCR, but also detection of protein markers. Intracellular markers can be best identified via RT-PCR, or protein detection methods such as immunocytochemistry, while cell surface markers are readily identified, e.g., by immunocytochemistry. [0227] Pluripotency of isolated cells can be confirmed by tests evaluating the ability of the iPSCs to differentiate into cells of each of the three germ layers. As one example, teratoma formation in nude mice can be used to evaluate the pluripotent character of the isolated clones. The cells can be introduced into nude mice and histology and/or immunohistochemistry can be performed on a tumor arising from the cells. The growth of a tumor comprising cells from all three germ layers, for example, further indicates that the cells are pluripotent stem cells. [0228] Patient-specific iPS cells or cell line can be created. There are many established methods in the art for creating patient specific iPS cells, e.g., as described in Takahashi and Yamanaka 2006; Takahashi, Tanabe et al.2007. For example, the creating step can comprise: a) isolating a somatic cell, such as a skin cell or fibroblast, from the patient; and b) introducing a set of pluripotency-associated genes into the somatic cell in order to induce the cell to become a pluripotent stem cell. The set of pluripotency-associated genes can be one or more of the genes selected from the group consisting of OCT4, SOX1, SOX2, SOX3, SOX15, SOX18, NANOG, KLF1, KLF2, KLF4, KLF5, c-MYC, n-MYC, REM2, TERT and LIN28. [0229] In some aspects, a biopsy or aspirate of a subjHFW¶V^ERQH^PDUURZ^FDQ^EH^SHUIRUPHG^^$^ELRSV\^RU^ aspirate is a sample of tissue or fluid taken from the body. There are many different kinds of biopsies or aspirates. Nearly all of them involve using a sharp tool to remove a small amount of tissue. If the biopsy will be on the skin or other sensitive area, numbing medicine can be applied first. A biopsy or aspirate can be performed according to any of the known methods in the art. For example, in a bone marrow aspirate, a large needle is used to enter the pelvis bone to collect bone marrow. [0230] In some aspects, a mesenchymal stem cell can be isolated from a subject. Mesenchymal stem FHOOV^FDQ^EH^LVRODWHG^DFFRUGLQJ^WR^DQ\^PHWKRG^NQRZQ^LQ^WKH^DUW^^VXFK^DV^IURP^D^VXEMHFW¶V^ERQH^PDUURZ^RU^ peripheral blood. For example, marrow aspirate can be collected into a syringe with heparin. Cells can be ZDVKHG^DQG^FHQWULIXJHG^RQ^D^3HUFROO^^GHQVLW\^JUDGLHQW^^&HOOV^^VXFK^DV^EORRG^FHOOV^^OLYHU^FHOOV^^LQWHUVWLWLDO^ cells, macrophages, mast cells, and thymocytes, can be separated using density gradient centrifugation PHGLD^^3HUFROO^^^7KH^FHOOV^FDQ^WKHQ^EH^FXOWXUHG^LQ^'XOEHFFR^V^PRGLILHG^(DJOH^V^PHGLXP^^'0(0^^^ORZ^ glucose) containing 10% fetal bovine serum (FBS) (Pittinger et. al., 1999, Science 284: 143-147). 6.8.1. Exemplary Genomic Targets [0231] The Type II Cas proteins and gRNAs of the disclosure can be used to alter various genomic targets. In some aspects, the methods of altering a cell are methods for altering a CCR5, EMX1, Fas, FANCF, HBB, ZSCAN2, Chr6, ADAMTSL1, B2M, CXCR4, PD1, DNMT1, Match8, TRAC, TRBC, VEGFAsite2, VEGFAsite3, CACNA, HEKsite3, HEKsite4, Chr8, BCR, ATM, HBG1, HPRT, IL2RG, NF1, USH2A, RHO, BcLenh, or CTFR genomic sequence. In some aspects, the methods of altering a cell are methods of altering a TRAC, B2M, PD1, or LAG3 genomic sequence. Reference sequences of RHO, TRAC, B2M, PD1, and LAG3 are available in public databases, for example those maintained by NCBI. For example, RHO has the NCBI gene ID: 6010; TRAC has the NCBI gene ID:28755; B2M has the NCBI gene ID: 567; PD1 has the NCBI gene ID:5133; and LAG3 has the NCBI gene ID: 3902. [0232] In some embodiments, the methods of altering a cell are methods for altering a hemoglobin VXEXQLW^EHWD^^+%%^^JHQH^^+%%^PXWDWLRQV^DUH^DVVRFLDWHG^ZLWK^ȕ-thalassemia and SCD. Dever et al., 2016 Nature 539(7629):384-389. [0233] In some embodiments, the methods of altering a cell are methods for altering a CCR5 gene. CCR5 has demonstrated involvement in several different disease states including, but not limited to, human immunodeficiency virus (HIV) and acquired immune deficiency syndrome (AIDS). WO 2018/119359 describes CCR5 editing by CRISPR-Cas to make loss of function CCR5 in order to provide protection against HIV infection, decrease one or more symptoms of HIV infection, halt or delay progression of HIV to AIDS, and/or decrease one or more symptoms of AIDS. [0234] In some embodiments, the methods of altering a cell are methods for altering a PD1, B2M gene, TRAC gene, or a combination thereof. CAR-T cells having PD1, B2M and TRAC genes disrupted by CRISPR-Type II Cas have demonstrated enhanced activity in preclinical glioma models. Choi et al., 2019, Journal for ImmunoTherapy of Cancer 7:309. [0235] Targeting of (one or more of) human TRAC, human B2M, human PD1, and human LAG3 genes can be used, for example, in the engineering of chimeric antigen receptor (CAR) T cells. For example, CRISPR/Cas technology has been used to deliver CAR-encoding DNA sequences to loci such as TRAC and PD1 (see, e.g., Eyquem et al., 2017, Nature 543(7643):113-117; Hu et al., 2023, eClinicalMedicine 60:102010), while TRAC, B2M, PD1, and LAG3 knockout CAR T-cells have been reported (see, e.g., Dimitri et al., 2022, Molecular Cancer 21:78; Liu et al., 2016, Cell Research 27:154-157; Ren et al., 2017, Clin Cancer Res.23(9):2255-2266; Zhang et al., 2017, Front Med.11(4):554-562). Thus, the Type II Cas proteins and TRAC, B2M, PD1, and LAG3 guides of the disclosure can be used for targeted knock-in of an exogenous DNA sequence to a desired genomic site in a human cell and/or knock-out of TRAC, B2M, PD1, or LAG3 in a human cell, for example a human T cell. In some embodiments, T cells are edited ex vivo to produce CAR-T cells and subsequently administered to a subject in need of CAR-T cell therapy. [0236] In some embodiments, the methods of altering a cell are methods for altering an USH2A gene. Mutations in the USH2A gene can cause Usher syndrome type 2A, which is characterized by progressive hearing and vision loss. [0237] In some embodiments, the methods of altering a cell are methods for altering a RHO gene. Mutations in the RHO gene can cause retinitis pigmentosa (RP). [0238] In some embodiments, the methods of altering a cell are methods for altering a DNMT1 gene. Mutations in the DNMT1 gene can cause DNMT1-related disorder, which is a degenerative disorder of the central and peripheral nervous systems. DNMT1-related disorder is characterized by sensory impairment, loss of sweating, dementia, and hearing loss. 7. EXAMPLES 7.1. Example 1: Identification and Characterization of Type II Cas Proteins [0239] This Example describes studies performed to identify and characterize DGHJ, CUAZ, AHWY, CBGI, ASDR, BCZZ, DRCY, DRPY, EAJR, and CFDE Type II Cas proteins. 7.1.1. Materials and Methods 7.1.1.1. Identification of Type II Cas Proteins From Metagenomic Data [0240] 154,723 bacterial and archaeal metagenome-assembled genomes (MAGs) reconstructed from the human microbiome (Pasolli, et al., 2019, Cell 176(3):649-662.e20) were screened in order to find new Type II Cas proteins. cas1, cas2 and cas9 genes were identified from the protein annotation, performed with Prokka version 1.12 (Seemann, 2014, Bioinformatics 30(14):2068±2069). CRISPR arrays were identified using MinCED version 0.4.2 (with default parameters) (Bland, et al., 2007, BMC bioinformatics 8:209). Only loci having a CRISPR array and cas1-2-9 genes at a maximum distance of 10 kbp from each other were considered. Loci containing Type II Cas proteins shorter than 950 aa were discarded. Type II Cas proteins from the same species, having similar length but slightly different sequence, were compared by multiple sequence alignment. Proteins presenting deletions in nucleasic domains were discarded. The remaining proteins were compared for sequencing coverage and the ortholog with the highest coverage was selected for each species. 7.1.1.2. tracrRNA Identification [0241] Identification of tracrRNAs for CRISPR-Type II Cas loci of interest was performed with a method based on a work by Chyou and Brown (Chyou and Brown, 2019, RNA biology 16(4):423±434). Starting from unique direct repeats in the CRISPR array, BLAST® (National Library of Medicine) version 2.2.31 (with parameters -task blastn-short -gapopen 2 -gapextend 1 -penalty -1 -reward 1 -evalue 1 -word_size 8) (Altschul, et al., 1990, Journal of Molecular Biology 215(3):403±410) was used to identify anti-repeats within a 3000 bp window flanking the CRISPR-Type II Cas locus. A custom version of RNIE (Gardner, et al., 2011, Nucleic Acids Research 39(14):5845±5852) was used to predict Rho-independent transcription terminators (RITs) near anti-repeats. Putative tracrRNA sequences, starting with an anti-repeat and ending with either a RIT (when found) or a poly-T, were combined with directed repeats to form sgRNA scaffolds. The secondary structure of sgRNA scaffolds was predicted using RNAsubopt version 2.4.14 (with parameters --noLP -e 5) (Lorenz, et al., 2011, Algorithms for Molecular Biology 6(1):26). sgRNAs lacking the functional modules identified by (Briner, et al., 2014 Molecular Cell 56(2):333±339), namely the repeat:anti-UHSHDW^GXSOH[^^QH[XV^DQG^^¶^KDLUSLQ-like folds, were discarded. 7.1.2. Results [0242] DGHJ, CUAZ, AHWY, CBGI, ASDR, BCZZ, DRCY, DRPY, EAJR, and CFDE Type II Cas proteins were identified: ^ DGHJ Type II Cas, originating from an unclassified Firmicutes, 1343 amino acids (aa) long; ^ CUAZ Type II Cas, originating from an unclassified bacterium from the Prochlorococcus genus, 1408 aa long; ^ AHWY Type II Cas, originating from an unclassified bacterium from the Eggerthellaceae family, 1392 aa long; ^ CBGI Type II Cas, originating from an unclassified bacterium from the Clostridia class, 1374 aa long; ^ ASDR Type II Cas, originating from an unclassified bacterium from the Eggerthellaceae family, 1404 aa long; ^ BCZZ Type II Cas, originating from an unclassified Clostridium, 1311 aa long; ^ DRCY Type II Cas, originating from Ellagibacter isourolithinifaciens, 1388 aa long; ^ DRPY Type II Cas, originating from an unclassified Lachnospiraceae bacterium, 1377 aa long; ^ EAJR Type II Cas, originating from an unclassified bacterium from the Ruminococcaceae family, 1324 aa long; and ^ CFDE Type II Cas, originating from an Unclassified bacterium from the Acholeplasmatales order, 1396 aa long. [0243] Amino acid sequences of DGHJ, CUAZ, AHWY, CBGI, ASDR, BCZZ, DRCY, DRPY, EAJR, and CFDE Type II Cas proteins and nucleotide sequences encoding exemplary DGHJ, CUAZ, AHWY, CBGI, ASDR, BCZZ, DRCY, DRPY, EAJR, and CFDE Type II Cas proteins are shown in Tables 1A-1K. crRNA and tracrRNA for the nucleases are described in Section 6.3. sgRNAs were designed for each Type II Cas protein by combining the identified tracrRNAs with the corresponding crRNAs extracted from the CRISPR arrays of each of the two nucleases. Exemplary sgRNA scaffolds are shown in Table 6. The predicted hairpin structure of the full length sgRNA molecules was used to establish how to trim the sgRNA scaffolds at the level of the repeat:antirepeat loop to generate the more compact molecules represented in FIG.1-10, which have been used throughout the studies. 7.2. Example 2: Additional Characterization of Type II Cas Proteins [0244] This Example describes studies performed to further characterize DGHJ, CUAZ, AHWY, CBGI, ASDR, BCZZ, DRCY, DRPY, EAJR, and CFDE Type II Cas proteins. 7.2.1. Materials and Methods 7.2.1.1. In vitro Cas PAM identification assay [0245] The in vitro PAM evaluation of the novel Type II Cas proteins was performed according to the protocol from Karvelis, et al., 2019, Methods in Enzymology, 616, pp.219±240. In brief: the human codon optimized version of the Type II Cas protein gene obtained as a synthetic construct (Twist Bioscience) was cloned into an expression vector for in vitro transcription and translation (IVT) (pT7-N-His-GST, Thermo Fisher Scientific). The sgRNAs to perform the assay were obtained by in vitro transcription of the guide using the HighYield T7 RNA Synthesis Kit (Jena Bioscience) starting from a PCR template generated by amplification from each sgRNA expression construct, as commonly done in the field. The primers used to generate the IVT templates are reported in Table 7. In vitro transcribed gRNAs were subsequently purified using the MEGAClear Transcription Clean-up kit (Thermo Fisher Scientific). The in vitro transcription and translation reaction for Cas expression was performed according to the PDQXIDFWXUHU¶V^SURWRFRO (1-Step Human High-Yield Mini IVT Kit, Thermo Fisher Scientific). The nuclease- JXLGH^51$^513^FRPSOH[^ZDV^DVVHPEOHG^E\^FRPELQLQJ^^^^^/^RI^WKH^VXSHUQDWDQW^FRQWDLQLQJ^WKH^VROXEOH^ 7\SH^,,^&DV^SURWHLQ^ZLWK^^^/^RI^5LER/RFN^51DVH^,QKLELWRU^^7KHUPR^)LVKHU^6FLHQWLILF^^DQG^^^J^RI^JXLGH^ RNA (previously transcribed in vitro). The RNP complex was used to digest 1ug of a PAM plasmid DNA OLEUDU\^^FRQWDLQLQJ^D^GHILQHG^WDUJHW^VHTXHQFH^IODQNHG^DW^WKH^^¶-end by a randomized 8 nucleotide PAM sequence) for 1 hour at 37°C. Table 7 Sequences of the primers used for PCR amplification of gRNAs used as templates for in vitro transcription Primer name 6HTXHQFH^^^¶^ĺ^^¶^ SEQ ID NO: IVT-T7-DGHJ- cctcTAATACGACTCACTATAGGtcGTCGCCCTCGA gRNA-F_{ACTTCACCTGTTTTAGAATTATGGAAACATAATGA}²¹⁸ IVT-DGHJ-gRNAv1- R^{AAAAAAGCTCTGATGAACAGAG 219} IVT-DGHJ-gRNAv2- R^{AAAAAATTTTCACCTAAGTGAAAC 220} IVT-T7-CUAZ- cctcTAATACGACTCACTATAGGtcGTCGCCCTCGA gRNA-F_{ACTTCACCTGATTTTAAGTATGGGAAACCATAAC}²²¹ IVT-CUAZ-gRNA-R AAAAAATGAGGGCAACCAATTAT 222 IVT-T7-AHWY- cctcTAATACGACTCACTATAGGtcGTCGCCCTCGA gRNA-F_{ACTTCACCTGTTTTGGAGCAGTGGAAACAC}²²³ IVT-AHWY- g_RNAv1-R^{AAAAAAGAGCCCGAAGGCTC 224} IVT-AHWY- g_RNAv2-R^{AAAAAAGTGCCGCCTACGC 225} IVT-T7- CBGI/ASDR-gRNA- cctcTAATACGACTCACTATAGGtcGTCGCCCTCGA A_CTT²²³ F_{CACCTGTTTTGGAGCAGTGGAAACAC} IVT-CBGI-gRNA-R AAAAAACTGCCGCCTACGC 226 IVT-ASDR-gRNA-R AAAAAATTCCCCCTACGGGG 227 IVT-T7-BCZZ- cctcTAATACGACTCACTATAGGtcGTCGCCCTCGA gRNA-F_{ACTTCACCTGTTTTAGTATTGAAAAATGAGTTAAA}^{228IVT-BCZZ-gRNA-R}_{AAAAAATTCACCTAAGTGAAACC 229} IVT-T7-DRCY- cctcTAATACGACTCACTATAGGtcGTCGCCCTCGA gRNA-F_{ACTTCACCTGTTTTGGAGCGAAAGCGAG}²³⁰ IVT-DRCY-gRNA-R AAAAAATTGCCACCTACGTGGCG 231 IVT-T7-DRPY- cctcTAATACGACTCACTATAGGtcGTCGCCCTCGA gRNA-F_{ACTTCACCTGTTTGAGAGCGAAAGCGAGT}²³² IVT-DRPY-gRNA-R AAAAAATGCACACACTGTGCAA 233 IVT-T7-EAJR- cctcTAATACGACTCACTATAGGtcGTCGCCCTCGA gRNA-F_{ACTTCACCTGTTTGAGAGTGAAAACAAGTTCAAA}²³⁴ IVT-EAJR-gRNA-R AAAAAATCACACACTGTGAATCCG 235 IVT-T7-CFDE- cctcTAATACGACTCACTATAGGtcGTCGCCCTCGA gRNA-F_{ACTTCACCTGTTTTGGAGCGAAAGCGAGT}²³⁶ IVT-CFDE-gRNA-R AAAAAATTTGTCCCCTACGGGA 237 [0246] A double stranded DNA adapter (Table 8) was ligated to the DNA ends generated by the targeted Cas cleavage and the final ligation product was purified using a GeneJet PCR Purification Kit (Thermo Fisher Scientific). Table 8 Sequences of the two oligonucleotides used to prepare the dsDNA adapter for the in vitro PAM assay 6HTXHQFH^^^¶^ĺ^^¶^ SEQ ID NO: Oligo UP CGGCATTCCTGCTGAACCGCTCTTCCGATCT 238 Oligo BOTTOM GATCGGAAGAGCGGTTCAGCAGGAATGCCG 239 [0247] One round of a two-step PCR (Phusion® HF DNA polymerase, Thermo Fisher Scientific) was performed to enrich the sequences that were cut using a set of forward primers annealing on the adapter and a reverse primer designed on the plasmid backbone downstream of the PAM (Table 9). A second round of PCR was performed to attach the Illumina indexes and adapters. PCR products were purified using the GeneJet PCR Purification Kit (Thermo Fisher Scientific). Table 9 Sequences of the primers used for NGS library preparation in the in vitro PAM assay Primer 6HTXHQFH^^^¶^ĺ^^¶^ SEQ ID NO: name_F4a^{TCGTCGGCAGCGTCAGATGTGTATAAGAGACAGCTGCTGAACCGC} 240 T_CTTCCGATCF4b^{TCGTCGGCAGCGTCAGATGTGTATAAGAGACAGTAAGACTGCTGAA} 241 C_{CGCTCTTCCGATCF4c}^{TCGTCGGCAGCGTCAGATGTGTATAAGAGACAGGCTAGACCTAATG} 242 T_{GATCTGCTGAACCGCTCTTCCGATCR3}^{GTCTCGTGGGCTCGGAGATGTGTATAAGAGACAGTCTGCGTTCTGA} 243 T_{TTAATCTGTATCAGGC} [0248] The library was analyzed with a 71-bp single read sequencing, using a flow cell v2 micro, on an Illumina MiSeq® sequencer. [0249] PAM sequences were extracted from Illumina MiSeq reads and used to generate PAM sequence logos, using Logomaker version 0.8. PAM heatmaps were used to display PAM enrichment, computed dividing the frequency of PAM sequences in the cleaved library by the frequency of the same sequences in a control uncleaved library. 7.2.2. Results [0250] Having determined the sgRNA requirements for the selected Type II Cas proteins, it was possible to proceed with the discovery of the PAM sites recognized by each nuclease. To this aim, a previously described in vitro PAM assay was used as described in Section 7.2.1.1. Briefly, the assay uses in vitro translated Type II Cas proteins coupled with an in vitro synthesized sgRNA to generate a functional ribonucleoprotein complex to cleave a plasmid library characterized by a defined target sequence followed by a randomized 8-nt stretch corresponding to the putative PAMs. Cleaved PAMs can then be recovered after library preparation by next generation sequencing. Table 10 contains the PAM preferences as determined based on the assay outcome. The PAM logos and the PAM heatmaps reporting the nucleotide preferences for specific positions along the PAMs are reported in FIGS.11-20. For DGHJ and AHWY Type II Cas proteins, two different types of sgRNA scaffolds were used, a longer (v1, FIGS.1A and 3A) and a shorter (v2, FIGS.1B and 3B) version, both of which yielded the same results in terms of PAM preferences (FIGS.11 and 13). Surprisingly, the DRPY Type II Cas protein can efficiently use guide RNAs related to DRCY Type II Cas protein, demonstrating unvaried PAM recognition sites (FIG.18). Table 10 PAM preferences as determined by in vitro assay Type II Cas protein In vitro-determined PAM Most preferred PAMs CUAZ Type II Cas NNGNC NNGNCNY AHWY Type II Cas NNRC NNAC, NNACNT DGHJ Type II Cas NGG NGG CBGI Type II Cas NNRTA NNRTA ASDR Type II Cas NNNNCC NNANCC, NNACCC BCZZ Type II Cas NGG NGG DRPY Type II Cas NNNAC NRNAC, NRHAC, NRNACD, NRHACD, NRNACA, NRHACA CFDE Type II Cas NNART NRART EAJR Type II Cas NNNRNY NRNRNY,NRNRNC, NANANC, NNNRDC DRCY Type II Cas NVRTA, NRRTA, NVATA NRATA 7.3. Example 3: Mammalian Cell Editing with Type II Cas Proteins [0251] This Example describes studies performed to evaluate editing using DGHJ, CUAZ, AHWY, CBGI, ASDR, BCZZ, DRCY, DRPY, EAJR, and CFDE Type II Cas proteins in mammalian cells. 7.3.1. Materials and Methods 7.3.1.1. Plasmids [0252] EF1alpha-driven expression plasmids were used to express each Type II Cas protein in mammalian cells. Briefly, the human codon-optimized coding sequences of the different Type II Cas were cloned into the aforementioned expression plasmid. An sgRNA scaffold for each Type II Cas (reported in Table 6^^ZLWK^DSSHQGHG^^¶^XUDFLOV^DV^VKRZQ^LQ^6(4^,'^126^^^^-115) was cloned into an expression plasmid containing a human U6 promoter to drive guide RNA expression in mammalian cells. Each Type II Cas coding sequence, modified by the addition of an SV5 tag at the N-terminus and two bipartite nuclear localization signals (1 at the N-term and 1 at the C-term) and human codon-optimized, as well as the sgRNA expression cassettes (U6 promoter + sgRNA scaffolds), were obtained as synthetic constructs from Twist Bioscience. Spacer sequences were cloned into the sgRNA expression plasmids as annealed DNA oligonucleotides using a double BsaI site present in the plasmid. The list of spacer sequences and relative cloning oligonucleotides used in the present example are reported in Tables 11- 14. Table 11 Oligonucleotides used for cloning sgRNA spacers targeting EGFP Spacer sequences Oligo used to clone the spacer in the sgRNA plasmid (*) SEQ ID SEQ ID SEQ Name Protospacer Oligo 1 (5'>3') Oligo 2 (5'>3') NO NO ID NO sgRNA- 244 245 246 AHWY- GCCCATCCTGGTCG caccGCCCATCCTGG aaacCCGTCCAGCTCGACCAGGA GFP-g1 AGCTGGACGG TCGAGCTGGACGG TGGGC sgRNA- 247 248 249 AHWY- GGCAGCTTGCCGGT caccGGCAGCTTGCC aaacATCTGCACCACCGGCAAGC GFP-g2 GGTGCAGAT GGTGGTGCAGAT TGCC sgRNA- 250 251 252 AHWY- GCAGTGCTTCAGCC caccGCAGTGCTTCA aaacTCGGGGTAGCGGCTGAAG GFP-g3 GCTACCCCGA GCCGCTACCCCGA CACTGC sgRNA- 253 254 255 ASDR- GTCCAGCTCGACCA caccGTCCAGCTCGA aaacTGCCCATCCTGGTCGAGCT GFP-g1 GGATGGGCA CCAGGATGGGCA GGAC sgRNA- 256 257 258 ASDR- GGGCGATGCCACCT caccGGGCGATGCCA aaacGCTTGCCGTAGGTGGCATC GFP-g2 ACGGCAAGC CCTACGGCAAGC GCCC sgRNA- 259 260 261 ASDR- CGTGCCCTGGCCCA caccGCGTGCCCTGG aaacTCACGAGGGTGGGCCAGG GFP-g3 CCCTCGTGA CCCACCCTCGTGA GCACGC sgRNA- 262 263 264 CBGI- CATCCTGGTCGAGC caccGCATCCTGGTC aaacCGCCGTCCAGCTCGACCAG GFP-g1 TGGACGGCG GAGCTGGACGGCG GATGC sgRNA- 265 266 267 CBGI- GAGATGAACTTCAG caccGAGATGAACTT aaacCAAGCTGACCCTGAAGTTC GFP-g2 GGTCAGCTTG CAGGGTCAGCTTG ATCTC sgRNA- 268 269 270 CBGI- GGTAGCGGCTGAAG caccGGTAGCGGCTG aaacCGTGCAGTGCTTCAGCCGC GFP-g3 CACTGCACG AAGCACTGCACG TACC sgRNA- 271 272 273 CUAZ- CGCTGAACTTGTGG caccGCGCTGAACTT aatcCGTAAACGGCCACAAGTTC GFP-g1 CCGTTTACG GTGGCCGTTTACG AGCGC sgRNA- 274 275 276 CUAZ- GTGCAGATGAACTT caccGTGCAGATGAA aatcGCTGACCCTGAAGTTCATCT GFP-g2 CAGGGTCAGC CTTCAGGGTCAGC GCAC sgRNA- 277 278 279 CUAZ- GAAGCAGCACGACT caccGAAGCAGCACG aatcACTTGAAGAAGTCGTGCTG GFP-g3 TCTTCAAGT ACTTCTTCAAGT CTTC sgRNA- 280 281 282 DGHJ- GTCGCCGTCCAGCT caccGTCGCCGTCCA aaacATCCTGGTCGAGCTGGACG GFP-g1 CGACCAGGAT GCTCGACCAGGAT GCGAC sgRNA- 283 284 285 DGHJ- GCCGGTGGTGCAGA caccGCCGGTGGTGC aaacTGAAGTTCATCTGCACCAC GFP-g2 TGAACTTCA AGATGAACTTCA CGGC sgRNA- 286 287 288 DGHJ- GAAGCACTGCACGC caccGAAGCACTGCA aaacTGACCTACGGCGTGCAGTG GFP-g3 CGTAGGTCA CGCCGTAGGTCA CTTC sgRNA- 289 290 291 DRCY- TGGCCGACAAGCAG caccGTGGCCGACAA aaacGCCGTTCTTCTGCTTGTCG GFP-g1 AAGAACGGC GCAGAAGAACGGC GCCAC sgRNA- 292 293 294 DRCY- GCAACATCCTGGGG caccGCAACATCCTG aaacCAGCTTGTGCCCCAGGATG GFP-g2 CACAAGCTG GGGCACAAGCTG TTGC sgRNA- 295 296 297 BCZZ- GGTGGTGCCCATCC caccGGTGGTGCCCA aaacGCTCGACCAGGATGGGCA GFP-g1 TGGTCGAGC TCCTGGTCGAGC CCACC sgRNA- 298 299 300 BCZZ- CCACAAGTTCAGCG caccGCCACAAGTTC aaacCGCCGGACACGCTGAACTT GFP-g2 TGTCCGGCG AGCGTGTCCGGCG GTGGC sgRNA- 283 284 285 BCZZ- GCCGGTGGTGCAGA caccGCCGGTGGTGC aaacTGAAGTTCATCTGCACCAC GFP-g3 TGAACTTCA AGATGAACTTCA CGGC sgRNA- 301 302 303 CFDE- CTGGACGGCGACGT caccGCTGGACGGCG aaacTGGCCGTTTACGTCGCCGT GFP-g1 AAACGGCCA ACGTAAACGGCCA CCAGC sgRNA- 304 305 306 CFDE- GCCACCTACGGCAA caccGCCACCTACGG aaacAGGGTCAGCTTGCCGTAGG GFP-g2 GCTGACCCT CAAGCTGACCCT TGGC sgRNA- 307 308 309 CFDE- CCTTCGGGCATGGC caccGCCTTCGGGCA aaacTTCAAGTCCGCCATGCCCG GFP-g3 GGACTTGAA TGGCGGACTTGAA AAGGC sgRNA- 310 311 312 DRPY- GACCAGGATGGGCA caccGACCAGGATGG aaacCCGGGGTGGTGCCCATCCT GFP-g1 CCACCCCGG GCACCACCCCGG GGTC sgRNA- 313 314 315 DRPY- GGTGGCATCGCCCT caccGGTGGCATCGC aaacGCGAGGGCGAGGGCGATG GFP-g2 CGCCCTCGC CCTCGCCCTCGC CCACC sgRNA- 316 317 318 DRPY- CAAGGAGGACGGCA caccGCAAGGAGGAC aaacCCAGGATGTTGCCGTCCTC GFP-g3 ACATCCTGG GGCAACATCCTGG CTTGC sgRNA- 319 320 321 EAJR- GGCACCACCCCGGT caccGGCACCACCCC aaacGAGCTGTTCACCGGGGTG GFP-g1 GAACAGCTC GGTGAACAGCTC GTGCC sgRNA- 322 323 324 EAJR- ACTTGTGGCCGTTTA caccGACTTGTGGCC aaacCGGCGACGTAAACGGCCA GFP-g2 CGTCGCCG GTTTACGTCGCCG CAAGTC sgRNA- 325 326 327 EAJR- GGGCGAGGGCGAT caccGGGCGAGGGC aaacCGTAGGTGGCATCGCCCTC GFP-g3 GCCACCTACG GATGCCACCTACG GCCC ^^^^7KH^FORQLQJ^RYHUKDQJ^LV^UHSRUWHG^LQ^ORZHUFDVH^^1XFOHRWLGHV^KLJKOLJKWHG^LQ^EROG^UHSUHVHQW^^¶-G appended to favor transcription from canonical U6 Pol III promoter Table 12 Oligonucleotides used for cloning sgRNA spacers targeting the B2M locus Spacer sequences Oligo used to clone the spacer in the sgRNA plasmid (*) SEQ ID SEQ SEQ Name Protospacer Oligo 1 (5'>3') Oligo 2 (5'>3') NO ID NO ID NO sgRNA- 328 329 330 AHWY- CAGAAAGAGAGAG caccGCAGAAAGAGAGAGTA aaacGCTCGCGCTACTCTC B2M-g1 TAGCGCGAGC GCGCGAGC TCTTTCTGC sgRNA- 331 332 333 AHWY- CCGTGGCCTTAGCT caccGCCGTGGCCTTAGCTG aaacCGCGAGCACAGCTAA B2M-g2 GTGCTCGCG TGCTCGCG GGCCACGGC sgRNA- 334 335 336 AHWY- TCAGGTACTCCAAA caccGTCAGGTACTCCAAAG aaacACCTGAATCTTTGGAG B2M-g3 GATTCAGGT ATTCAGGT TACCTGAC sgRNA- 337 338 339 AHWY- GTACTCCAAAGATT caccGTACTCCAAAGATTCAG aaacGTAAACCTGAATCTTT B2M-g4 CAGGTTTAC GTTTAC GGAGTAC sgRNA- 340 341 342 AHWY- GACACATAGCAATT caccGACACATAGCAATTCA aaacAATTTCCTGAATTGCT B2M-g5 CAGGAAATT GGAAATT ATGTGTC sgRNA- 343 344 345 ASDR- ACTCTCTCTTTCTG caccGACTCTCTCTTTCTGGC aaacCCTCCAGGCCAGAAA B2M-g1 GCCTGGAGG CTGGAGG GAGAGAGTC sgRNA- 346 347 348 ASDR- CCAAAGATTCAGGT caccGCCAAAGATTCAGGTTT aaacCGTGAGTAAACCTGA B2M-g2 TTACTCACG ACTCACG ATCTTTGGC sgRNA- 349 350 351 ASDR- CTGAATTGCTATGT caccGCTGAATTGCTATGTGT aaacAACCCAGACACATAG B2M-g3 GTCTGGGTT CTGGGTT CAATTCAGC sgRNA- 352 353 354 ADSR- ATTGCTATGTGTCT caccGATTGCTATGTGTCTGG aaacATGAAACCCAGACAC B2M-g4 GGGTTTCAT GTTTCAT ATAGCAATC sgRNA- 355 356 357 ASDR- GTCAACTTCAATGT caccGTCAACTTCAATGTCG aaacCATCCATCCGACATTG B2M-g5 CGGATGGATG GATGGATG AAGTTGAC sgRNA- 358 359 360 ASDR- B2M-g5 ACTTCAATGTCGGA caccGACTTCAATGTCGGAT aaacCATCCATCCGACATTG _20nt TGGATG GGATG AAGTC sgRNA- 361 362 363 CBGI- GATAGCCTCCAGG caccGATAGCCTCCAGGCCA aaacTCTCTTTCTGGCCTGG B2M-g1 CCAGAAAGAGA GAAAGAGA AGGCTATC sgRNA- 364 365 366 CBGI- GGAGGGTAGGAGA caccGGAGGGTAGGAGAGAC aaacAGCGTGAGTCTCTCC B2M-g2 GACTCACGCT TCACGCT TACCCTCC sgRNA- 367 368 369 CBGI- GACGTGAGTAAAC caccGACGTGAGTAAACCTG aaacCAAAGATTCAGGTTTA B2M-g3 CTGAATCTTTG AATCTTTG CTCACGTC sgRNA- 370 371 372 CBGI- TTCCATTCTCTGCT caccGTTCCATTCTCTGCTGG aaacACGTCATCCAGCAGA B2M-g4 GGATGACGT ATGACGT GAATGGAAC sgRNA- 373 374 375 CBGI- GTCGGATGGATGA caccGTCGGATGGATGAAAC aaacGTCTGGGTTTCATCCA B2M-g5 AACCCAGAC CCAGAC TCCGAC sgRNA- 376 377 378 CFDE- GATAGCCTCCAGG caccGATAGCCTCCAGGCCA aaacCTCTTTCTGGCCTGG B2M-g1 CCAGAAAGAG GAAAGAG AGGCTATC sgRNA- 379 380 381 CFDE- TCTGGCCTGGAGG caccGTCTGGCCTGGAGGCT aaacCGCTGGATAGCCTCC B2M-g2 CTATCCAGCG ATCCAGCG AGGCCAGAC sgRNA- 382 383 384 CFDE- GACGTGAGTAAAC caccGACGTGAGTAAACCTG aaacAAAGATTCAGGTTTAC B2M-g3 CTGAATCTTT AATCTTT TCACGTC sgRNA- 385 386 387 CFDE- GTTTACTCACGTCA caccGTTTACTCACGTCATCC aaacCTGCTGGATGACGTG B2M-g4 TCCAGCAG AGCAG AGTAAAC sgRNA- 388 389 390 CFDE- GAGAATGGAAAGT caccGAGAATGGAAAGTCAA aaacGGAAATTTGACTTTCC B2M-g5 CAAATTTCC ATTTCC ATTCTC sgRNA- 391 392 393 DRPY- TTCAGTGGGGGTG caccGTTCAGTGGGGGTGAA aaacACACTGAATTCACCCC B2M-g1 AATTCAGTGT TTCAGTGT CACTGAAC sgRNA- 394 395 396 DRPY- GCCAGAAAGAGAG caccGCCAGAAAGAGAGAGT aaacCGCGCTACTCTCTCTT B2M-g2 AGTAGCGCG AGCGCG TCTGGC sgRNA- 397 398 399 DRPY- CAATGTCGGATGG caccGCAATGTCGGATGGAT aaacTGGGTTTCATCCATCC B2M-g3 ATGAAACCCA GAAACCCA GACATTGC sgRNA- 400 401 402 DRPY- GCAAGGACTGGTC caccGCAAGGACTGGTCTTT aaacAGAGATAGAAAGACC B2M-g4 TTTCTATCTCT CTATCTCT AGTCCTTGC ^^^^7KH^FORQLQJ^RYHUKDQJ^LV^UHSRUWHG^LQ^ORZHUFDVH^^1XFOHRWLGHV^KLJKOLJKWHG^LQ^EROG^UHSUHVHQW^^¶-G appended to favor transcription from canonical U6 Pol III promoter Table 13 Oligonucleotides used for cloning sgRNA spacers targeting the TRAC locus Spacer sequences Oligo used to clone the spacer in the sgRNA plasmid (*) SEQ ID SEQ SEQ Name Protospacer Oligo 1 (5'>3') Oligo 2 (5'>3') NO ID NO ID NO sgRNA- 403 404 405 AHWY- ATCAAAATCGGTGA caccGATCAAAATCGGTGAAT aaacGTCTGCCTATTCACCG TRAC-g1 ATAGGCAGAC AGGCAGAC ATTTTGATC sgRNA- 406 407 408 AHWY- ACACATCAGAATCC caccGACACATCAGAATCCTT aaacCACAAAGTAAGGATTC TRAC-g2 TTACTTTGTG ACTTTGTG TGATGTGTC sgRNA- 409 410 411 AHWY- GCACTGTTGCTCTT caccGCACTGTTGCTCTTGAA aaacATGGACTTCAAGAGC TRAC-g3 GAAGTCCAT GTCCAT AACAGTGC sgRNA- 412 413 414 AHWY- AACTGTGCTAGACA caccGAACTGTGCTAGACAT aaacATAGACCTCATGTCTA TRAC-g4 TGAGGTCTAT GAGGTCTAT GCACAGTTC sgRNA- 415 416 417 AHWY- GTATATCACAGACA caccGTATATCACAGACAAAA aaacAGCACAGTTTTGTCTG TRAC-g5 AAACTGTGCT CTGTGCT TGATATAC sgRNA- 418 419 420 ASDR- TCTTGTCCCACAGA caccGTCTTGTCCCACAGATA aaacTGGATATCTGTGGGA TRAC-g1 TATCCA TCCA CAAGAC sgRNA- 421 422 423 ASDR- CCCACAGATATCCA caccGCCCACAGATATCCAG aaacGGGTTCTGGATATCT TRAC-g2 GAACCC AACCC GTGGGC sgRNA- 424 425 426 ASDR- GTGTACCAGCTGA caccGTGTACCAGCTGAGAG aaacAGAGTCTCTCAGCTG TRAC-g3 GAGACTCT ACTCT GTACAC sgRNA- 427 428 429 ASDR- TGTCTGTGATATAC caccGTGTCTGTGATATACAC aaacTGATGTGTATATCACA TRAC-g4 ACATCA ATCA GACAC sgRNA- 430 431 432 ASDR- GCCACAGCACTGTT caccGCCACAGCACTGTTGC aaacCAAGAGCAACAGTGC TRAC-g5 GCTCTTG TCTTG TGTGGC sgRNA- 433 434 435 CBGI- GATATCCAGAACCC caccGATATCCAGAACCCTG aaacGGCAGGGTCAGGGTT TRAC-g1 TGACCCTGCC ACCCTGCC CTGGATATC sgRNA- 436 437 438 CBGI- CACTGGATTTAGAG caccGCACTGGATTTAGAGT aaacGCTGAGAGACTCTAA TRAC-g2 TCTCTCAGC CTCTCAGC ATCCAGTGC sgRNA- 439 440 441 CBGI- TTTGTTTGAGAATC caccGTTTGTTTGAGAATCAA aaacACCGATTTTGATTCTC TRAC-g3 AAAATCGGT AATCGGT AAACAAAC sgRNA- 442 443 444 CBGI- GATTCTCAAACAAA caccGATTCTCAAACAAATGT aaacTGTGACACATTTGTTT TRAC-g4 TGTGTCACA GTCACA GAGAATC sgRNA- 445 446 447 CFDE- GTCACAAAGTAAGG caccGTCACAAAGTAAGGATT aaacATCAGAATCCTTACTT TRAC-g5 ATTCTGAT CTGAT TGTGAC sgRNA- 448 449 450 CFDE- GCCGTGTACCAGC caccGCCGTGTACCAGCTGA aaacGAGTCTCTCAGCTGG TRAC-g1 TGAGAGACTC GAGACTC TACACGGC sgRNA- 451 452 453 CFDE- GCTGAGAGACTCTA caccGCTGAGAGACTCTAAAT aaacTCACTGGATTTAGAGT TRAC-g2 AATCCAGTGA CCAGTGA CTCTCAGC sgRNA- 454 455 456 CFDE- GCAGACAGACTTGT caccGCAGACAGACTTGTCA aaacAAATCCAGTGACAAGT TRAC-g3 CACTGGATTT CTGGATTT CTGTCTGC sgRNA- 457 458 459 CFDE- ATTCACCGATTTTG caccGATTCACCGATTTTGAT aaacTTTGAGAATCAAAATC TRAC-g4 ATTCTCAAA TCTCAAA GGTGAATC sgRNA- 460 461 462 CFDE- ATTTGTTTGAGAAT caccGATTTGTTTGAGAATCA aaacCCGATTTTGATTCTCA TRAC-g5 CAAAATCGG AAATCGG AACAAATC sgRNA- 463 464 465 CFDE- TGATTCTCAAACAA caccGTGATTCTCAAACAAAT aaacGTGACACATTTGTTTG TRAC-g6 ATGTGTCAC GTGTCAC AGAATCAC sgRNA- 466 467 468 CFDE- TTACTTTGTGACAC caccGTTACTTTGTGACACAT aaacCAAACAAATGTGTCAC TRAC-g7 ATTTGTTTG TTGTTTG AAAGTAAC sgRNA- 469 470 471 CFDE- GTTTTGTCTGTGAT caccGTTTTGTCTGTGATATA aaacGATGTGTATATCACAG TRAC-g8 ATACACATC CACATC ACAAAAC sgRNA- 472 473 474 CFDE- CAGGCCACAGCAC caccGCAGGCCACAGCACTG aaacAAGAGCAACAGTGCT TRAC-g9 TGTTGCTCTT TTGCTCTT GTGGCCTGC sgRNA- 475 476 477 CFDE- TRAC- gAACAGTGCTGTGG caccGAACAGTGCTGTGGCC aaacTTGCTCCAGGCCACA g10 CCTGGAGCAA TGGAGCAA GCACTGTTC sgRNA- 478 479 480 DRPY- ACTGGATTTAGAGT caccGACTGGATTTAGAGTCT aaacAGCTGAGAGACTCTA TRAC-g1 CTCTCAGCT CTCAGCT AATCCAGTC sgRNA- 481 482 483 DRPY- GCCTATTCACCGAT caccGCCTATTCACCGATTTT aaacAGAATCAAAATCGGTG TRAC-g2 TTTGATTCT GATTCT AATAGGC sgRNA- 484 485 486 DRPY- ACACATCAGAATCC caccGACACATCAGAATCCTT aaacACAAAGTAAGGATTCT TRAC-g3 TTACTTTGT ACTTTGT GATGTGTC sgRNA- 487 488 489 DRPY- CTAGCACAGTTTTG caccGCTAGCACAGTTTTGTC aaacTATCACAGACAAAACT TRAC-g4 TCTGTGATA TGTGATA GTGCTAGC sgRNA- 490 491 492 DRPY- TGAAGTCCATAGAC caccGTGAAGTCCATAGACC aaacAGACATGAGGTCTAT TRAC-g5 CTCATGTCT TCATGTCT GGACTTCAC sgRNA- 493 494 495 DRPY- GTCTAGCACAGTTT caccGTCTAGCACAGTTTTGT aaacTCACAGACAAAACTGT TRAC-g6 TGTCTGTGA CTGTGA GCTAGAC sgRNA- 496 497 498 DRPY- ATAATGCTGTTGTT caccGATAATGCTGTTGTTGA aaacAACGCCTTCAACAACA TRAC-g7 GAAGGCGTT AGGCGTT GCATTATC (*) The cloning overhang is reported in lowercase. Nucleotides highlighted LQ^EROG^UHSUHVHQW^^¶-G appended to favor transcription from canonical U6 Pol III promoter Table 14 Oligonucleotides used for cloning sgRNA spacers targeting the PD1 locus Spacer sequences Oligo used to clone the spacer in the sgRNA plasmid (*) SEQ SEQ SEQ Name Protospacer Oligo 1 (5'>3') Oligo 2 (5'>3') ID NO ID NO ID NO sgRNA- 499 500 501 AHWY- ATCCTGGCCGCCA caccGATCCTGGCCGCCAGC aaacTACAACTGGGCTGGC PD1-g1 GCCCAGTTGTA CCAGTTGTA GGCCAGGATC sgRNA- 502 503 504 AHWY- GGGGCTGCTCCAG caccGGGGCTGCTCCAGGCA aaacGATCTGCATGCCTGG PD1-g2 GCATGCAGATC TGCAGATC AGCAGCCCC sgRNA- 505 506 507 AHWY- GCCCAGTTGTAGC caccGCCCAGTTGTAGCACC aaacTCTGGGCGGTGCTAC PD1-g3 ACCGCCCAGA GCCCAGA AACTGGGC sgRNA- 508 509 510 AHWY- GGCCAGTCGTCTG caccGGCCAGTCGTCTGGGC aaacTAGCACCGCCCAGAC PD1-g4 GGCGGTGCTA GGTGCTA GACTGGCC sgRNA- 511 512 513 ASDR- GTGGGGCTGCTCC caccGTGGGGCTGCTCCAGG aaacGCATGCCTGGAGCAG PD1-g1 AGGCATGC CATGC CCCCAC sgRNA- 514 515 516 ASDR- CGACCCCACCTAC caccGCGACCCCACCTACCT aaacTTCTTAGGTAGGTGG PD1-g2 CTAAGAA AAGAA GGTCGC sgRNA- 517 518 519 ASDR- CAGGCATGCAGAT caccGCAGGCATGCAGATCC aaacCTGTGGGATCTGCAT PD1-g3 CCCACAG CACAG GCCTGC sgRNA- 520 521 522 ASDR- GCCGCCAGCCCAG caccGCCGCCAGCCCAGTTG aaacTGCTACAACTGGGCT PD1-g4 TTGTAGCA TAGCA GGCGGC sgRNA- 523 524 525 ASDR- CTAAGAACCATCCT caccGCTAAGAACCATCCTGG aaacGCGGCCAGGATGGTT PD1-g5 GGCCGC CCGC CTTAGC sgRNA- 526 527 528 ASDR- TCAGACTCCCCAGA caccGTCAGACTCCCCAGAC aaacCAGGGCCTGTCTGGG PD1-g6 CAGGCCCTG AGGCCCTG GAGTCTGAC sgRNA- 529 530 531 ASDR- CGGGGCCTCCGAG caccGCGGGGCCTCCGAGGC aaacCAGGTGCGGCCTCGG PD1-g7 GCCGCACCTG CGCACCTG AGGCCCCGC sgRNA- 532 533 534 CBGI- GAACCATCCTGGC caccGAACCATCCTGGCCGC aaacCTGGGCTGGCGGCCA PD1-g1 CGCCAGCCCAG CAGCCCAG GGATGGTTC sgRNA- 535 536 537 CBGI- GGCTGGCGGCCAG caccGGCTGGCGGCCAGGAT aaacAAGAACCATCCTGGC PD1-g2 GATGGTTCTT GGTTCTT CGCCAGCC sgRNA- 538 539 540 CBGI- CATCGGAGAGCTT caccGCATCGGAGAGCTTCG aaacGTTTAGCACGAAGCTC PD1-g3 CGTGCTAAAC TGCTAAAC TCCGATGC sgRNA- 541 542 543 CBGI- CCGTCTGGTTGCTG caccGCCGTCTGGTTGCTGG aaacCATGAGCCCCAGCAA PD1-g4 GGGCTCATG GGCTCATG CCAGACGGC sgRNA- 544 545 546 CBGI- GGGCCAGGGAGAT caccGGGCCAGGGAGATGGC aaacCTGTGGGGCCATCTC PD1-g5 GGCCCCACAG CCCACAG CCTGGCCC sgRNA- 547 548 549 CFDE- GGGGGGTTCCAGG caccGGGGGGTTCCAGGGCC aaacCCAGACAGGCCCTGG PD1-g1 GCCTGTCTGG TGTCTGG AACCCCCC sgRNA- 550 551 552 CFDE- CGGGCCCTGACCA caccGCGGGCCCTGACCACG aaacCACATGAGCGTGGTC PD1-g2 CGCTCATGTG CTCATGTG AGGGCCCGC sgRNA- 553 554 555 DRPY- GCCACCTTCACCTG caccGCCACCTTCACCTGCA aaacGGAGAAGCTGCAGGT PD1-g1 CAGCTTCTCC GCTTCTCC GAAGGTGGC sgRNA- 556 557 558 DRPY- GGCCAGGACTGCC caccGGCCAGGACTGCCGCT aaacACACGGAAGCGGCAG PD1-g2 GCTTCCGTGT TCCGTGT TCCTGGCC sgRNA- 559 560 561 DRPY- TGGGGCTCATGCG caccGTGGGGCTCATGCGGT aaacAAACTGGTACCGCAT PD1-g3 GTACCAGTTT ACCAGTTT GAGCCCCAC sgRNA- 562 563 564 DRPY- CAACACATCGGAG caccGCAACACATCGGAGAG aaacGCACGAAGCTCTCCG PD1-g4 AGCTTCGTGC CTTCGTGC ATGTGTTGC sgRNA- 565 566 567 DRPY- GTCACGCCCGTTG caccGTCACGCCCGTTGGGC aaacACACAACTGCCCAAC PD1-g5 GGCAGTTGTGT AGTTGTGT GGGCGTGAC ^^^^7KH^FORQLQJ^RYHUKDQJ^LV^UHSRUWHG^LQ^ORZHUFDVH^^1XFOHRWLGHV^KLJKOLJKWHG^LQ^EROG^UHSUHVHQW^^¶-G appended to favor transcription from canonical U6 Pol III promote 7.3.1.2. Cells [0253] HEK293T cells (obtained from ATCC) and U2OS-EFGP cells, harboring a single integrated copy of an EGFP reporter gene, were cultured in DMEM (Life Technologies) supplemented with 10% FBS (Life Technologies), 2 mM L-Glutamine (Life Technologies) and penicillin/streptomycin (Life Technologies). T cells were cultured in RPMI 1640 medium (Gibco, Life Technologies) supplemented with 10% FBS (Life Technologies) and 100 U/ml Il-2 (Immunotools) in plates supporting suspension growth. T cells were DFWLYDWHG^^^KRXUV^DIWHU^WKDZLQJ^XVLQJ^,PPXQR&XOW^^+XPDQ^&'^^&'^^^&'^^7^&HOO^$FWLYDWRU^^^^^L/1*10⁶ cells) (Stem Cell) and IL-^^^^^^8^^/^^and kept at concentration of 2*10⁶ cells/mL. All cells were incubated at 37°C and 5% CO₂ in a humidified atmosphere. All cells tested mycoplasma negative (PlasmoTest, Invivogen). 7.3.1.3. Cell line transfections [0254] To perform editing studies on endogenous target loci 10⁵ HEK293T cells were seeded in a 24- well plate 24 hours before transfection. Cells were then transfected with 500 ng of nuclease-expressing plasmid together with 250 ng of sgRNA expression vector targeting the locus of interest using the TransIT^TM-LT1 rHDJHQW^^0LUXV^%LR^^DFFRUGLQJ^WR^WKH^PDQXIDFWXUHU¶V^SURWRFRO^ Cell pellets were collected 3 days from transfection for analysis. To perform EGFP editing studies, 2*10⁵ U2OS-EGFP cells were nucleofected with 500 ng of nuclease-expressing plasmid and 250 ng of sgRNA-expressing plasmid containing a guide designed to target EGFP using the 4D-1XFOHRIHFWRU^^6(^.LW^^/RQ]D^^^'1-100 SURJUDP^^DFFRUGLQJ^WR^WKH^PDQXIDFWXUHU¶V^SURWRFRO^^$IWHU^HOHFWURSRUDWLRQ^^FHOOV^ZHUH^SODWHG^LQ^D^^^-well plate. EGFP knock-out was analyzed 4 days after nucleofection using a BD FACSymphony^TM A1 (BD) flow cytometer. 7.3.1.4. Primary T cells transfections [0255] Commercial T cells were electroporated with mRNAs for the expression of selected candidates together with chemically synthesized sgRNAs (obtained from IDT). Briefly, plasmids encoding for Type II Cas proteins under the control of a T7 RNA polymerase promoter were linearized via enzymatic digestion and used as template for in vitro transcription (IVT) using the HighYield^TM T7 RNA Synthesis kit (Jena %LRVFLHQFH^^IROORZLQJ^WKH^PDQXIDFWXUHU¶V^SURWRFRO^ [0256] To perform single-plex editing studies, 1x10⁶ commercially purified CD3+ T cells were nucleofected with 3 μg of nuclease-expressing IVT mRNA and 100 pmol of synthetic sgRNAs targeting selected endogenous loci using the 4D-1XFOHRIHFWRU^^3^^.LW^^/RQ]D^^^(^-115 program, according to the PDQXIDFWXUHU¶V^SURWRFRO^^$IWHU^HOHFWURSRUDWLRQ^^FHOOV^ZHUH^SODWHG^LQ^D^^^-well plate and cultured in RPMI Media enriched with IL-2 (100 U/mL). [0257] For multi-plexing studies, 3 μg of nuclease-expressing IVT mRNA and 200 pmol of synthetic sgRNAs ± instead of 100 pmol - targeting each locus were used. 7.3.1.5. Evaluation of gene editing [0258] For evaluation of gene editing in cell lines, three days after transfection cells were collected and the genomic DNA was extracted using the QuickExtract^TM DNA Extraction Solution (Lucigen) according WR^WKH^PDQXIDFWXUHU¶V^LQVWUXFWLRQV^^7R^DPSOLI\^WKH^WDUJHW^ORFL^^3&5^UHDFWLRQV^ZHUH^SHUIRUPHG^XVLQJ^WKH^ HOT FIREPol® polymerase (Solis BioDyne) and the oligonucleotides listed in Table 15. The amplified products were purified, Sanger sequenced (EasyRun service, Microsynth) and analyzed with the TIDE web tool (shinyapps.datacurators.nl/tide/) to quantify indels. The same primers were used for Sanger sequencing reactions on amplicons, selecting the forward or reverse primer by taking into account the position of the cut site within each amplicon. For evaluation of gene editing in T cells, genomic DNA was extracted using the NucleoSspin^TM Tissue (Machery-1DJHO^^NLW^DFFRUGLQJ^WR^WKH^PDQXIDFWXUHU¶V^ instructions. The amplification of the target loci was obtained with the same procedure described for in cell lines. Table 15 Oligonucleotides used to amplify target regions to perform TIDE analysis Locus )RU^^^¶^ĺ^^¶^ SEQ ID NO 5HY^^^¶^ĺ^^¶^ SEQ ID NO TRAC CATCACGAGCAGCTGGTTTC 568 TGGCAATGGATAAGGCCGAG 569 B2M ex1 CTCTAACCTGGCACTGCGTC 570 CGAAGTCCACAGCTCTCCAG 571 B2M ex2 TGGCCAGAGTGGAAATGGAA 572 TGTATTTGTGCAAGTGCTGCT 573 PD-1 ex1 CACTGCCTCTGTCACTCTCG 574 GACTTGGGCCAGGGGAGGAG 575 PD-1 ex2 CCACCCACCCAGACCAGTTA 576 CTCTTTGATCTGCGCCTTGG 577 [0259] For the evaluation of productive knockouts at the B2M, TRAC and PD1 loci in T cells, cells were stained using the following antibodies: APC anti-human ß2-microglobulin (B2M) (BioLegend), Pacific %OXH^^DQWL-human TCR alpha/Beta (TRAC) (BioLegend), and PE anti-human CD279 (PD-1) (BioLegend); and analyzed by cytofluorimetry 4 days post-electroporation. 7.3.1.6. Off-target measurements [0260] Genome-wide off-targets were evaluated using the GUIDE-seq protocol (Tsai et al., 2015, Nat Biotechnol 33(2):187-197) with some modifications (Casini et al., 2018, Nat Biotechnol 36(3):265-271). Briefly, 1.8x10⁵ HEK293T cells were seeded in a 24-well plate the day before transfection. Cells were then transfected with 500 ng of each Type II Cas protein together with 250 ng of the corresponding sgRNA expression plasmids, 10 pmol of bait dsODN (sequence reported in the cited publications) using Lipofectamine 3^^^^^7KHUPR^6FLHQWLILF^^DFFRUGLQJ^WR^PDQXIDFWXUHU¶V^LQVWUXFWLRQV^^^^K^KRXUV^DIWHU^ transfection, genomic DNA was extracted using the NucleoSpin^TM Tissue kit (Macherey-Nagel) and libraries were prepared and sequenced according to previously published protocols (Tsai et al., 2015, Nat Biotechnol 33(2):187-197). A maximum of 5 different libraries were loaded on a single Illumina Miseq^TM Reagent kit v2 ± 300 cycles flow cell. Raw sequencing data were analyzed using an available dedicated computational pipeline (Tsai et al., 2016, Nat Biotechnol 34(5):483). 7.3.2. Results 7.3.2.1. Evaluation of the editing activity using an EGFP reporter system [0261] After the determination of the PAM sequences and sgRNAs for the selected Type II Cas proteins and after obtaining preliminary information on the ability of these nucleases to cut a desired target in vitro (plasmid target used during the PAM assay), their ability to cleave selected targets in mammalian cells was investigated. An EGFP reporter system was used as it allowed an easier readout on the editing activity, based on the loss of fluorescence of treated cells quantitatively measured by cytofluorimetry. sgRNAs targeting the EGFP coding sequence (2/3 for each evaluated Type II Cas protein) were thus designed for all the Type II Cas proteins and evaluated in U2OS cells stably expressing a single copy of an EGFP reporter by transient electroporation. Surprisingly, as reported in FIG.21, some of the evaluated guides in combination with their respective Type II Cas protein were able to significantly downregulate EGFP expression in target cells. In particular, AHWY, ASDR, CGBI, CFDE and DRPY Type II Cas proteins showed very high (>80% EGFP KO) with one or more of the evaluated guides; CUAZ, DRCY and EAJR Type II Cas proteins showed appreciable knock-out activity (>20% EGFP KO) with at least one of the evaluated sgRNAs; BCZZ and DGJH Type II Cas proteins showed only marginal activity (<10% EGFP KO) with the evaluated guide RNAs. These data clearly demonstrate that evaluated Type II Cas proteins were able to modify very efficiently genetic targets in mammalian cells and can thus be exploited to edit the mammalian genome. For AHWY and DGHJ Type II Cas protein two alternative sgRNA scaffolds were evaluated (v1 and v2, which differ for the extent of trimming, see also Table 6) showing similar editing capabilities. For further studies with AHWY Type II Cas the v2 version was exploited. 7.3.2.2. Type II Cas efficiently edit endogenous genomic loci in cell lines [0262] To extensively evaluate the cleavage activity of AHWY, ASDR, CBGI, CFDE and DRPY Type II Cas, a panel of endogenous loci (B2M, TRAC, PD1) which are commonly targeted to generate allogeneic CAR-T cells (Chimeric Antigen Receptor T cells) was selected for editing studies. For each target locus multiple sgRNAs were designed and evaluated in parallel by transient plasmid transfection in HEK293T cells. [0263] As shown in FIGS.22A-22C, for each of the evaluated loci most of the nucleases showed significant editing activity with at least one of the selected guide RNAs, demonstrating that these novel Type II Cas proteins have the ability to effectively modify genomic targets of interest, in accordance with previous data obtained in EGFP reporter knock-out studies. Interestingly, an alternative version of the ASDR Type II Cas g5 targeting the B2M locus characterized by a shorter spacer sequence (20nt) demonstrated improved editing activity towards the target site. 7.3.2.3. Type II Cas are active in primary T cells [0264] The highest performing hits emerging from validation studies in HEK293T cells (DRPY, CFDE and ASDR Type II Cas proteins) were further characterized in another system by mRNA transfection in primary T cells. The most active sgRNA for each nuclease associated with each target (TRAC and B2M) was chemically synthesized with standard end-modifications to improve stability. Specifically the following guide/nuclease combinations shown in Table 16 were evaluated: Table 16 Guide RNAs used in T cell studies Type II Cas B2M TRAC DRPY g3 g5 CFDE g1, g2 g1 ASDR g5_20nt g3 [0265] To target PD1 with DRPY Type II Cas, g5 was employed. [0266] As shown in FIG.23A, most of the evaluated nuclease-guide combinations were able to induce significant indels at their target sites (e.g. for the TRAC locus), with the exception of CFDE Type II Cas where two sgRNAs targeting B2M were evaluated without obtaining strong modification of the target loci. Notably, among the evaluated nucleases, DRPY Type II Cas showed robust modification of both TRAC and B2M, at levels which were comparable with the SpCas9 benchmark. Measured indel levels were paralleled by similar downregulation of the two protein targets, as determined by cytofluorimetric analysis (FIG.23B). [0267] Since DRPY Type II Cas was shown to be particularly active towards the B2M and TRAC loci, the analysis was extended to the PD1 target, where high levels of indel formation were demonstrated (FIG. 23C). [0268] Given the excellent editing profile demonstrated in single-gene targeting studies, the editing activity of DRPY Type II Cas was additionally evaluated in multiplex targeting studies where the three previously targeted genes (B2M, TRAC and PD1) are edited in T cells simultaneously after transient electroporation of the nuclease-encoding mRNA and the corresponding pool of synthetic sgRNAs. Editing outcomes were evaluated through protein downregulation measured by cytofluorimetric analysis, demonstrating high levels of knock-out across all three targets at a level which was comparable to the benchmark nuclease SpCas9 (FIG.24). [0269] Overall, reported data clearly demonstrate that these novel editors can be delivered efficiently as mRNAs to target T cells producing appreciable levels of editing at multiple endogenous loci. One of the evaluated editors, DRPY Type II Cas proved to be particularly active demonstrating robust editing both in singleplex and multiplexing studies. 7.3.2.4. Genome-wide specificity of DRPY Type II Cas [0270] The genome-wide specificity of DRPY Type II Cas was evaluated in combination with two of the previously evaluated sgRNAs targeting the B2M and PD1 loci using the GUIDE-seq methodology exploiting plasmid transfection in HEK293T cells. Parallel studies were performed using SpCas9 and two guides targeting the same loci reported in literature (Ren et al., 2017, Clin Res 23(9):2255-2266; Liu et al., 2017, Cell Research 27:154-157). As shown in FIGS.25A-2B, no off-target sites were identified for DRPY Type II Cas for all the evaluated guides, while multiple sites were flagged when SpCas9 and its corresponding guide RNAs were used (FIGS.25C-25D). [0271] These data demonstrated the high level of target specificity offered by DRPY Type II Cas which is a clear advantage for human therapeutic applications. 8. SPECIFIC EMBODIMENTS [0272] The present disclosure is exemplified by the specific embodiments below. 1. A Type II Cas protein comprising an amino acid sequence having at least 50% sequence identity to: (a) the amino acid sequence of a RuvC-I domain of a reference protein sequence; (b) the amino acid sequence of a RuvC-II domain of a reference protein sequence; (c) the amino acid sequence of a RuvC-III domain of a reference protein sequence; (d) the amino acid sequence of a BH domain of a reference protein sequence; (e) the amino acid sequence of a REC1_1 domain of a reference protein sequence; (f) the amino acid sequence of a REC2 domain of a reference protein sequence; (g) the amino acid sequence of a REC1_2 domain of a reference protein sequence; (h) the amino acid sequence of a HNH domain of a reference protein sequence; (i) the amino acid sequence of a Topo domain of a reference protein sequence; (j) the amino acid sequence of a CTD domain of a reference protein sequence; or (k) the amino acid sequence of the full length of a reference protein sequence; wherein the reference protein sequence is SEQ ID NO:1, SEQ ID NO:2, SEQ ID NO:7, SEQ ID NO:8, SEQ ID NO:13, SEQ ID NO:14, SEQ ID NO:19, SEQ ID NO:20, SEQ ID NO:25, SEQ ID NO:26, SEQ ID NO:31, SEQ ID NO:32, SEQ ID NO:37, SEQ ID NO:38, SEQ ID NO:43, SEQ ID NO:44, SEQ ID NO:49, SEQ ID NO:50, SEQ ID NO:55, or SEQ ID NO:56. 2. The Type II Cas protein of embodiment 1, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 50% identical to the amino acid sequence of the RuvC-I domain of the reference protein sequence. 3. The Type II Cas protein of embodiment 1, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 55% identical to the amino acid sequence of the RuvC-I domain of the reference protein sequence. 4. The Type II Cas protein of embodiment 1, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 60% identical to the amino acid sequence of the RuvC-I domain of the reference protein sequence. 5. The Type II Cas protein of embodiment 1, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 65% identical to the amino acid sequence of the RuvC-I domain of the reference protein sequence. 6. The Type II Cas protein of embodiment 1, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 70% identical to the amino acid sequence of the RuvC-I domain of the reference protein sequence. 7. The Type II Cas protein of embodiment 1, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 75% identical to the amino acid sequence of the RuvC-I domain of the reference protein sequence. 8. The Type II Cas protein of embodiment 1, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 80% identical to the amino acid sequence of the RuvC-I domain of the reference protein sequence. 9. The Type II Cas protein of embodiment 1, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 85% identical to the amino acid sequence of the RuvC-I domain of the reference protein sequence. 10. The Type II Cas protein of embodiment 1, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 90% identical to the amino acid sequence of the RuvC-I domain of the reference protein sequence. 11. The Type II Cas protein of embodiment 1, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 95% identical to the amino acid sequence of the RuvC-I domain of the reference protein sequence. 12. The Type II Cas protein of embodiment 1, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 96% identical to the amino acid sequence of the RuvC-I domain of the reference protein sequence. 13. The Type II Cas protein of embodiment 1, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 97% identical to the amino acid sequence of the RuvC-I domain of the reference protein sequence. 14. The Type II Cas protein of embodiment 1, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 98% identical to the amino acid sequence of the RuvC-I domain of the reference protein sequence. 15. The Type II Cas protein of embodiment 1, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 99% identical to the amino acid sequence of the RuvC-I domain of the reference protein sequence. 16. The Type II Cas protein of embodiment 1, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is identical to the amino acid sequence of the RuvC-I domain of the reference protein sequence. 17. The Type II Cas protein of any one of embodiments 1 to 16, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 50% identical to the amino acid sequence of the RuvC-II domain of the reference protein sequence. 18. The Type II Cas protein of any one of embodiments 1 to 16, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 55% identical to the amino acid sequence of the RuvC-II domain of the reference protein sequence. 19. The Type II Cas protein of any one of embodiments 1 to 16, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 60% identical to the amino acid sequence of the RuvC-II domain of the reference protein sequence. 20. The Type II Cas protein of any one of embodiments 1 to 16, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 65% identical to the amino acid sequence of the RuvC-II domain of the reference protein sequence. 21. The Type II Cas protein of any one of embodiments 1 to 16, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 70% identical to the amino acid sequence of the RuvC-II domain of the reference protein sequence. 22. The Type II Cas protein of any one of embodiments 1 to 16, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 75% identical to the amino acid sequence of the RuvC-II domain of the reference protein sequence. 23. The Type II Cas protein of any one of embodiments 1 to 16, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 80% identical to the amino acid sequence of the RuvC-II domain of the reference protein sequence. 24. The Type II Cas protein of any one of embodiments 1 to 16, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 85% identical to the amino acid sequence of the RuvC-II domain of the reference protein sequence. 25. The Type II Cas protein of any one of embodiments 1 to 16, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 90% identical to the amino acid sequence of the RuvC-II domain of the reference protein sequence. 26. The Type II Cas protein of any one of embodiments 1 to 16, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 95% identical to the amino acid sequence of the RuvC-II domain of the reference protein sequence. 27. The Type II Cas protein of any one of embodiments 1 to 16, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 96% identical to the amino acid sequence of the RuvC-II domain of the reference protein sequence. 28. The Type II Cas protein of any one of embodiments 1 to 16, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 97% identical to the amino acid sequence of the RuvC-II domain of the reference protein sequence. 29. The Type II Cas protein of any one of embodiments 1 to 16, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 98% identical to the amino acid sequence of the RuvC-II domain of the reference protein sequence. 30. The Type II Cas protein of any one of embodiments 1 to 16, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 99% identical to the amino acid sequence of the RuvC-II domain of the reference protein sequence. 31. The Type II Cas protein of any one of embodiments 1 to 16, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is identical to the amino acid sequence of the RuvC-II domain of the reference protein sequence. 32. The Type II Cas protein of any one of embodiments 1 to 31, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 50% identical to the amino acid sequence of the RuvC-III domain of the reference protein sequence. 33. The Type II Cas protein of any one of embodiments 1 to 31, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 55% identical to the amino acid sequence of the RuvC-III domain of the reference protein sequence. 34. The Type II Cas protein of any one of embodiments 1 to 31, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 60% identical to the amino acid sequence of the RuvC-III domain of the reference protein sequence. 35. The Type II Cas protein of any one of embodiments 1 to 31, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 65% identical to the amino acid sequence of the RuvC-III domain of the reference protein sequence. 36. The Type II Cas protein of any one of embodiments 1 to 31, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 70% identical to the amino acid sequence of the RuvC-III domain of the reference protein sequence. 37. The Type II Cas protein of any one of embodiments 1 to 31, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 75% identical to the amino acid sequence of the RuvC-III domain of the reference protein sequence. 38. The Type II Cas protein of any one of embodiments 1 to 31, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 80% identical to the amino acid sequence of the RuvC-III domain of the reference protein sequence. 39. The Type II Cas protein of any one of embodiments 1 to 31, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 85% identical to the amino acid sequence of the RuvC-III domain of the reference protein sequence. 40. The Type II Cas protein of any one of embodiments 1 to 31, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 90% identical to the amino acid sequence of the RuvC-III domain of the reference protein sequence. 41. The Type II Cas protein of any one of embodiments 1 to 31, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 95% identical to the amino acid sequence of the RuvC-III domain of the reference protein sequence. 42. The Type II Cas protein of any one of embodiments 1 to 31, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 96% identical to the amino acid sequence of the RuvC-III domain of the reference protein sequence. 43. The Type II Cas protein of any one of embodiments 1 to 31, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 97% identical to the amino acid sequence of the RuvC-III domain of the reference protein sequence. 44. The Type II Cas protein of any one of embodiments 1 to 31, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 98% identical to the amino acid sequence of the RuvC-III domain of the reference protein sequence. 45. The Type II Cas protein of any one of embodiments 1 to 31, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 99% identical to the amino acid sequence of the RuvC-III domain of the reference protein sequence. 46. The Type II Cas protein of any one of embodiments 1 to 31, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is identical to the amino acid sequence of the RuvC-III domain of the reference protein sequence. 47. The Type II Cas protein of any one of embodiments 1 to 46, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 50% identical to the amino acid sequence of the BH domain of the reference protein sequence. 48. The Type II Cas protein of any one of embodiments 1 to 46, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 55% identical to the amino acid sequence of the BH domain of the reference protein sequence. 49. The Type II Cas protein of any one of embodiments 1 to 46, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 60% identical to the amino acid sequence of the BH domain of the reference protein sequence. 50. The Type II Cas protein of any one of embodiments 1 to 46, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 65% identical to the amino acid sequence of the BH domain of the reference protein sequence. 51. The Type II Cas protein of any one of embodiments 1 to 46, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 70% identical to the amino acid sequence of the BH domain of the reference protein sequence. 52. The Type II Cas protein of any one of embodiments 1 to 46, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 75% identical to the amino acid sequence of the BH domain of the reference protein sequence. 53. The Type II Cas protein of any one of embodiments 1 to 46, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 80% identical to the amino acid sequence of the BH domain of the reference protein sequence. 54. The Type II Cas protein of any one of embodiments 1 to 46, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 85% identical to the amino acid sequence of the BH domain of the reference protein sequence. 55. The Type II Cas protein of any one of embodiments 1 to 46, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 90% identical to the amino acid sequence of the BH domain of the reference protein sequence. 56. The Type II Cas protein of any one of embodiments 1 to 46, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 95% identical to the amino acid sequence of the BH domain of the reference protein sequence. 57. The Type II Cas protein of any one of embodiments 1 to 46, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 96% identical to the amino acid sequence of the BH domain of the reference protein sequence. 58. The Type II Cas protein of any one of embodiments 1 to 46, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 97% identical to the amino acid sequence of the BH domain of the reference protein sequence. 59. The Type II Cas protein of any one of embodiments 1 to 46, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 98% identical to the amino acid sequence of the BH domain of the reference protein sequence. 60. The Type II Cas protein of any one of embodiments 1 to 46, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 99% identical to the amino acid sequence of the BH domain of the reference protein sequence. 61. The Type II Cas protein of any one of embodiments 1 to 46, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is identical to the amino acid sequence of the BH domain of the reference protein sequence. 62. The Type II Cas protein of any one of embodiments 1 to 61, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 50% identical to the amino acid sequence of the REC1_1 domain of the reference protein sequence. 63. The Type II Cas protein of any one of embodiments 1 to 61, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 55% identical to the amino acid sequence of the REC1_1 domain of the reference protein sequence. 64. The Type II Cas protein of any one of embodiments 1 to 61, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 60% identical to the amino acid sequence of the REC1_1 domain of the reference protein sequence. 65. The Type II Cas protein of any one of embodiments 1 to 61, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 65% identical to the amino acid sequence of the REC1_1 domain of the reference protein sequence. 66. The Type II Cas protein of any one of embodiments 1 to 61, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 70% identical to the amino acid sequence of the REC1_1 domain of the reference protein sequence. 67. The Type II Cas protein of any one of embodiments 1 to 61, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 75% identical to the amino acid sequence of the REC1_1 domain of the reference protein sequence. 68. The Type II Cas protein of any one of embodiments 1 to 61, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 80% identical to the amino acid sequence of the REC1_1 domain of the reference protein sequence. 69. The Type II Cas protein of any one of embodiments 1 to 61, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 85% identical to the amino acid sequence of the REC1_1 domain of the reference protein sequence. 70. The Type II Cas protein of any one of embodiments 1 to 61, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 90% identical to the amino acid sequence of the REC1_1 domain of the reference protein sequence. 71. The Type II Cas protein of any one of embodiments 1 to 61, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 95% identical to the amino acid sequence of the REC1_1 domain of the reference protein sequence. 72. The Type II Cas protein of any one of embodiments 1 to 61, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 96% identical to the amino acid sequence of the REC1_1 domain of the reference protein sequence. 73. The Type II Cas protein of any one of embodiments 1 to 61, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 97% identical to the amino acid sequence of the REC1_1 domain of the reference protein sequence. 74. The Type II Cas protein of any one of embodiments 1 to 61, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 98% identical to the amino acid sequence of the REC1_1 domain of the reference protein sequence. 75. The Type II Cas protein of any one of embodiments 1 to 61, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 99% identical to the amino acid sequence of the REC1_1 domain of the reference protein sequence. 76. The Type II Cas protein of any one of embodiments 1 to 61, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is identical to the amino acid sequence of the REC1_1 domain of the reference protein sequence. 77. The Type II Cas protein of any one of embodiments 1 to 76, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 50% identical to the amino acid sequence of the REC2 domain of the reference protein sequence. 78. The Type II Cas protein of any one of embodiments 1 to 76, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 55% identical to the amino acid sequence of the REC2 domain of the reference protein sequence. 79. The Type II Cas protein of any one of embodiments 1 to 76, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 60% identical to the amino acid sequence of the REC2 domain of the reference protein sequence. 80. The Type II Cas protein of any one of embodiments 1 to 76, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 65% identical to the amino acid sequence of the REC2 domain of the reference protein sequence. 81. The Type II Cas protein of any one of embodiments 1 to 76, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 70% identical to the amino acid sequence of the REC2 domain of the reference protein sequence. 82. The Type II Cas protein of any one of embodiments 1 to 76, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 75% identical to the amino acid sequence of the REC2 domain of the reference protein sequence. 83. The Type II Cas protein of any one of embodiments 1 to 76, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 80% identical to the amino acid sequence of the REC2 domain of the reference protein sequence. 84. The Type II Cas protein of any one of embodiments 1 to 76, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 85% identical to the amino acid sequence of the REC2 domain of the reference protein sequence. 85. The Type II Cas protein of any one of embodiments 1 to 76, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 90% identical to the amino acid sequence of the REC2 domain of the reference protein sequence. 86. The Type II Cas protein of any one of embodiments 1 to 76, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 95% identical to the amino acid sequence of the REC2 domain of the reference protein sequence. 87. The Type II Cas protein of any one of embodiments 1 to 76, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 96% identical to the amino acid sequence of the REC2 domain of the reference protein sequence. 88. The Type II Cas protein of any one of embodiments 1 to 76, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 97% identical to the amino acid sequence of the REC2 domain of the reference protein sequence. 89. The Type II Cas protein of any one of embodiments 1 to 76, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 98% identical to the amino acid sequence of the REC2 domain of the reference protein sequence. 90. The Type II Cas protein of any one of embodiments 1 to 76, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 99% identical to the amino acid sequence of the REC2 domain of the reference protein sequence. 91. The Type II Cas protein of any one of embodiments 1 to 76, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is identical to the amino acid sequence of the REC2 domain of the reference protein sequence. 92. The Type II Cas protein of any one of embodiments 1 to 91, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 50% identical to the amino acid sequence of the REC1_2 domain of the reference protein sequence. 93. The Type II Cas protein of any one of embodiments 1 to 91, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 55% identical to the amino acid sequence of the REC1_2 domain of the reference protein sequence. 94. The Type II Cas protein of any one of embodiments 1 to 91, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 60% identical to the amino acid sequence of the REC1_2 domain of the reference protein sequence. 95. The Type II Cas protein of any one of embodiments 1 to 91, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 65% identical to the amino acid sequence of the REC1_2 domain of the reference protein sequence. 96. The Type II Cas protein of any one of embodiments 1 to 91, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 70% identical to the amino acid sequence of the REC1_2 domain of the reference protein sequence. 97. The Type II Cas protein of any one of embodiments 1 to 91, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 75% identical to the amino acid sequence of the REC1_2 domain of the reference protein sequence. 98. The Type II Cas protein of any one of embodiments 1 to 91, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 80% identical to the amino acid sequence of the REC1_2 domain of the reference protein sequence. 99. The Type II Cas protein of any one of embodiments 1 to 91, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 85% identical to the amino acid sequence of the REC1_2 domain of the reference protein sequence. 100. The Type II Cas protein of any one of embodiments 1 to 91, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 90% identical to the amino acid sequence of the REC1_2 domain of the reference protein sequence. 101. The Type II Cas protein of any one of embodiments 1 to 91, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 95% identical to the amino acid sequence of the REC1_2 domain of the reference protein sequence. 102. The Type II Cas protein of any one of embodiments 1 to 91, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 96% identical to the amino acid sequence of the REC1_2 domain of the reference protein sequence. 103. The Type II Cas protein of any one of embodiments 1 to 91, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 97% identical to the amino acid sequence of the REC1_2 domain of the reference protein sequence. 104. The Type II Cas protein of any one of embodiments 1 to 91, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 98% identical to the amino acid sequence of the REC1_2 domain of the reference protein sequence. 105. The Type II Cas protein of any one of embodiments 1 to 91, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 99% identical to the amino acid sequence of the REC1_2 domain of the reference protein sequence. 106. The Type II Cas protein of any one of embodiments 1 to 91, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is identical to the amino acid sequence of the REC1_2 domain of the reference protein sequence. 107. The Type II Cas protein of any one of embodiments 1 to 106, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 50% identical to the amino acid sequence of the HNH domain of the reference protein sequence. 108. The Type II Cas protein of any one of embodiments 1 to 106, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 55% identical to the amino acid sequence of the HNH domain of the reference protein sequence. 109. The Type II Cas protein of any one of embodiments 1 to 106, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 60% identical to the amino acid sequence of the HNH domain of the reference protein sequence. 110. The Type II Cas protein of any one of embodiments 1 to 106, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 65% identical to the amino acid sequence of the HNH domain of the reference protein sequence. 111. The Type II Cas protein of any one of embodiments 1 to 106, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 70% identical to the amino acid sequence of the HNH domain of the reference protein sequence. 112. The Type II Cas protein of any one of embodiments 1 to 106, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 75% identical to the amino acid sequence of the HNH domain of the reference protein sequence. 113. The Type II Cas protein of any one of embodiments 1 to 106, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 80% identical to the amino acid sequence of the HNH domain of the reference protein sequence. 114. The Type II Cas protein of any one of embodiments 1 to 106, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 85% identical to the amino acid sequence of the HNH domain of the reference protein sequence. 115. The Type II Cas protein of any one of embodiments 1 to 106, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 90% identical to the amino acid sequence of the HNH domain of the reference protein sequence. 116. The Type II Cas protein of any one of embodiments 1 to 106, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 95% identical to the amino acid sequence of the HNH domain of the reference protein sequence. 117. The Type II Cas protein of any one of embodiments 1 to 106, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 96% identical to the amino acid sequence of the HNH domain of the reference protein sequence. 118. The Type II Cas protein of any one of embodiments 1 to 106, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 97% identical to the amino acid sequence of the HNH domain of the reference protein sequence. 119. The Type II Cas protein of any one of embodiments 1 to 106, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 98% identical to the amino acid sequence of the HNH domain of the reference protein sequence. 120. The Type II Cas protein of any one of embodiments 1 to 106, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 99% identical to the amino acid sequence of the HNH domain of the reference protein sequence. 121. The Type II Cas protein of any one of embodiments 1 to 106, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is identical to the amino acid sequence of the HNH domain of the reference protein sequence. 122. The Type II Cas protein of any one of embodiments 1 to 121, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 50% identical to the amino acid sequence of the Topo domain of the reference protein sequence. 123. The Type II Cas protein of any one of embodiments 1 to 121, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 55% identical to the amino acid sequence of the Topo domain of the reference protein sequence. 124. The Type II Cas protein of any one of embodiments 1 to 121, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 60% identical to the amino acid sequence of the Topo domain of the reference protein sequence. 125. The Type II Cas protein of any one of embodiments 1 to 121, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 65% identical to the amino acid sequence of the Topo domain of the reference protein sequence. 126. The Type II Cas protein of any one of embodiments 1 to 121, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 70% identical to the amino acid sequence of the Topo domain of the reference protein sequence. 127. The Type II Cas protein of any one of embodiments 1 to 121, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 75% identical to the amino acid sequence of the Topo domain of the reference protein sequence. 128. The Type II Cas protein of any one of embodiments 1 to 121, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 80% identical to the amino acid sequence of the Topo domain of the reference protein sequence. 129. The Type II Cas protein of any one of embodiments 1 to 121, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 85% identical to the amino acid sequence of the Topo domain of the reference protein sequence. 130. The Type II Cas protein of any one of embodiments 1 to 121, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 90% identical to the amino acid sequence of the Topo domain of the reference protein sequence. 131. The Type II Cas protein of any one of embodiments 1 to 121, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 95% identical to the amino acid sequence of the Topo domain of the reference protein sequence. 132. The Type II Cas protein of any one of embodiments 1 to 121, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 96% identical to the amino acid sequence of the Topo domain of the reference protein sequence. 133. The Type II Cas protein of any one of embodiments 1 to 121, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 97% identical to the amino acid sequence of the Topo domain of the reference protein sequence. 134. The Type II Cas protein of any one of embodiments 1 to 121, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 98% identical to the amino acid sequence of the Topo domain of the reference protein sequence. 135. The Type II Cas protein of any one of embodiments 1 to 121, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 99% identical to the amino acid sequence of the Topo domain of the reference protein sequence. 136. The Type II Cas protein of any one of embodiments 1 to 121, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is identical to the amino acid sequence of the Topo domain of the reference protein sequence. 137. The Type II Cas protein of any one of embodiments 1 to 136, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 50% identical to the amino acid sequence of the CTD domain of the reference protein sequence. 138. The Type II Cas protein of any one of embodiments 1 to 136, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 55% identical to the amino acid sequence of the CTD domain of the reference protein sequence. 139. The Type II Cas protein of any one of embodiments 1 to 136, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 60% identical to the amino acid sequence of the CTD domain of the reference protein sequence. 140. The Type II Cas protein of any one of embodiments 1 to 136, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 65% identical to the amino acid sequence of the CTD domain of the reference protein sequence. 141. The Type II Cas protein of any one of embodiments 1 to 136, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 70% identical to the amino acid sequence of the CTD domain of the reference protein sequence. 142. The Type II Cas protein of any one of embodiments 1 to 136, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 75% identical to the amino acid sequence of the CTD domain of the reference protein sequence. 143. The Type II Cas protein of any one of embodiments 1 to 136, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 80% identical to the amino acid sequence of the CTD domain of the reference protein sequence. 144. The Type II Cas protein of any one of embodiments 1 to 136, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 85% identical to the amino acid sequence of the CTD domain of the reference protein sequence. 145. The Type II Cas protein of any one of embodiments 1 to 136, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 90% identical to the amino acid sequence of the CTD domain of the reference protein sequence. 146. The Type II Cas protein of any one of embodiments 1 to 136, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 95% identical to the amino acid sequence of the CTD domain of the reference protein sequence. 147. The Type II Cas protein of any one of embodiments 1 to 136, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 96% identical to the amino acid sequence of the CTD domain of the reference protein sequence. 148. The Type II Cas protein of any one of embodiments 1 to 136, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 97% identical to the amino acid sequence of the CTD domain of the reference protein sequence. 149. The Type II Cas protein of any one of embodiments 1 to 136, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 98% identical to the amino acid sequence of the CTD domain of the reference protein sequence. 150. The Type II Cas protein of any one of embodiments 1 to 136, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 99% identical to the amino acid sequence of the CTD domain of the reference protein sequence. 151. The Type II Cas protein of any one of embodiments 1 to 136, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is identical to the amino acid sequence of the CTD domain of the reference protein sequence. 152. The Type II Cas protein of any one of embodiments 1 to 151, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 55% identical to the full length of the reference protein sequence. 153. The Type II Cas protein of any one of embodiments 1 to 151, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 60% identical to the full length of the reference protein sequence. 154. The Type II Cas protein of any one of embodiments 1 to 151, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 65% identical to the full length of the reference protein sequence. 155. The Type II Cas protein of any one of embodiments 1 to 151, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 70% identical to the full length of the reference protein sequence. 156. The Type II Cas protein of any one of embodiments 1 to 151, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 75% identical to the full length of the reference protein sequence. 157. The Type II Cas protein of any one of embodiments 1 to 151, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 80% identical to the full length of the reference protein sequence. 158. The Type II Cas protein of any one of embodiments 1 to 151, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 85% identical to the full length of the reference protein sequence. 159. The Type II Cas protein of any one of embodiments 1 to 151, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 90% identical to the full length of the reference protein sequence. 160. The Type II Cas protein of any one of embodiments 1 to 151, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 95% identical to the full length of the reference protein sequence. 161. The Type II Cas protein of any one of embodiments 1 to 151, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 96% identical to the full length of the reference protein sequence. 162. The Type II Cas protein of any one of embodiments 1 to 151, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 97% identical to the full length of the reference protein sequence. 163. The Type II Cas protein of any one of embodiments 1 to 151, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 98% identical to the full length of the reference protein sequence. 164. The Type II Cas protein of any one of embodiments 1 to 151, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 99% identical to the full length of the reference protein sequence. 165. The Type II Cas protein of any one of embodiments 1 to 151, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is identical to the full length of the reference protein sequence. 166. The Type II Cas protein of any one of embodiments 1 to 165, which is a chimeric Type II Cas protein. 167. The Type II Cas protein of any one of embodiments 1 to 166, which is a fusion protein. 168. The Type II Cas protein of embodiment 167, which comprises one or more nuclear localization signals. 169. The Type II Cas protein of embodiment 168, which comprises two or more nuclear localization signals. 170. The Type II Cas protein of embodiment 168 or embodiment 169, which comprises an N- terminal nuclear localization signal. 171. The Type II Cas protein of any one of embodiments 168 to 170, which comprises a C- terminal nuclear localization signal. 172. The Type II Cas protein of any one of embodiments 168 to 171, which comprises an N- terminal nuclear localization signal and a C-terminal nuclear localization signal. 173. The Type II Cas protein of any one of embodiments 168 to 172, wherein the amino acid sequence of one or more of the nuclear localization signals comprises the amino acid sequence KRTADGSEFESPKKKRKV (SEQ ID NO:116), PKKKRKV (SEQ ID NO:117), PKKKRRV (SEQ ID NO:118), KRPAATKKAGQAKKKK (SEQ ID NO:119), YGRKKRRQRRR (SEQ ID NO:120), RKKRRQRRR (SEQ ID NO:121), PAAKRVKLD (SEQ ID NO:122), RQRRNELKRSP (SEQ ID NO:123), VSRKRPRP (SEQ ID NO:124), PPKKARED (SEQ ID NO:125), PQPKKKPL (SEQ ID NO:126), SALIKKKKKMAP (SEQ ID NO:127), PKQKKRK (SEQ ID NO:128), RKLKKKIKKL (SEQ ID NO:129), REKKKFLKRR (SEQ ID NO:130), KRKGDEVDGVDEVAKKKSKK (SEQ ID NO:131), RKCLQAGMNLEARKTKK (SEQ ID NO:132), NQSSNFGPMKGGNFGGRSSGPYGGGGQYFAKPRNQGGY (SEQ ID NO:133), RMRIZFKNKGKDTAELRRRRVEVSVELRKAKKDEQILKRRNV (SEQ ID NO:134) PAAKKKKLD (SEQ ID NO:582) or KRPAATKKAGQAKKKKLD (SEQ ID NO:583). 174. The Type II Cas protein of embodiment 173, wherein the amino acid sequence of one or more of the nuclear localization signals comprises the amino acid sequence KRTADGSEFESPKKKRKV (SEQ ID NO:116). 175. The Type II Cas protein of embodiment 173, wherein the amino acid sequence of one or more of the nuclear localization signals comprises the amino acid sequence PKKKRKV (SEQ ID NO:117). 176. The Type II Cas protein of embodiment 173, wherein the amino acid sequence of one or more of the nuclear localization signals comprises the amino acid sequence PKKKRRV (SEQ ID NO:118). 177. The Type II Cas protein of embodiment 173, wherein the amino acid sequence of one or more of the nuclear localization signals comprises the amino acid sequence KRPAATKKAGQAKKKK (SEQ ID NO:119). 178. The Type II Cas protein of embodiment 173, wherein the amino acid sequence of one or more of the nuclear localization signals comprises the amino acid sequence YGRKKRRQRRR (SEQ ID NO:120). 179. The Type II Cas protein of embodiment 173, wherein the amino acid sequence of one or more of the nuclear localization signals comprises the amino acid sequence RKKRRQRRR (SEQ ID NO:121). 180. The Type II Cas protein of embodiment 173, wherein the amino acid sequence of one or more of the nuclear localization signals comprises the amino acid sequence PAAKRVKLD (SEQ ID NO:122). 181. The Type II Cas protein of embodiment 173, wherein the amino acid sequence of one or more of the nuclear localization signals comprises the amino acid sequence RQRRNELKRSP (SEQ ID NO:123). 182. The Type II Cas protein of embodiment 173, wherein the amino acid sequence of one or more of the nuclear localization signals comprises the amino acid sequence VSRKRPRP (SEQ ID NO:124). 183. The Type II Cas protein of embodiment 173, wherein the amino acid sequence of one or more of the nuclear localization signals comprises the amino acid sequence PPKKARED (SEQ ID NO:125). 184. The Type II Cas protein of embodiment 173, wherein the amino acid sequence of one or more of the nuclear localization signals comprises the amino acid sequence PQPKKKPL (SEQ ID NO:126). 185. The Type II Cas protein of embodiment 173, wherein the amino acid sequence of one or more of the nuclear localization signals comprises the amino acid sequence SALIKKKKKMAP (SEQ ID NO:127). 186. The Type II Cas protein of embodiment 173, wherein the amino acid sequence of one or more of the nuclear localization signals comprises the amino acid sequence PKQKKRK (SEQ ID NO:128). 187. The Type II Cas protein of embodiment 173, wherein the amino acid sequence of one or more of the nuclear localization signals comprises the amino acid sequence RKLKKKIKKL (SEQ ID NO:129). 188. The Type II Cas protein of embodiment 173, wherein the amino acid sequence of one or more of the nuclear localization signals comprises the amino acid sequence REKKKFLKRR (SEQ ID NO:130). 189. The Type II Cas protein of embodiment 173, wherein the amino acid sequence of one or more of the nuclear localization signals comprises the amino acid sequence KRKGDEVDGVDEVAKKKSKK (SEQ ID NO:131). 190. The Type II Cas protein of embodiment 173, wherein the amino acid sequence of one or more of the nuclear localization signals comprises the amino acid sequence RKCLQAGMNLEARKTKK (SEQ ID NO:132). 191. The Type II Cas protein of embodiment 173, wherein the amino acid sequence of one or more of the nuclear localization signals comprises the amino acid sequence NQSSNFGPMKGGNFGGRSSGPYGGGGQYFAKPRNQGGY (SEQ ID NO:133). 192. The Type II Cas protein of embodiment 173, wherein the amino acid sequence of one or more of the nuclear localization signals comprises the amino acid sequence RMRIZFKNKGKDTAELRRRRVEVSVELRKAKKDEQILKRRNV (SEQ ID NO:134). 193. The Type II Cas protein of embodiment 173, wherein the amino acid sequence of one or more of the nuclear localization signals comprises the amino acid sequence PAAKKKKLD (SEQ ID NO:582). 194. The Type II Cas protein of embodiment 173, wherein the amino acid sequence of one or more of the nuclear localization signals comprises the amino acid sequence KRPAATKKAGQAKKKKLD (SEQ ID NO:583). 195. The Type II Cas protein of any one of embodiments 168 to 194, wherein the amino acid sequence of each nuclear localization signal is the same. 196. The Type II Cas protein of any one of embodiments 166 to 195, which comprises a fusion partner which is a DNA, RNA or protein modification enzyme, optionally wherein the DNA, RNA or protein modification enzyme is an adenosine deaminase, a cytidine deaminase, a reverse transcriptase, a guanosyl transferase, a DNA methyltransferase, a RNA methyltransferase, a DNA demethylase, a RNA demethylase, a dioxygenase, a polyadenylate polymerase, a pseudouridine synthase, an acetyltransferase, a deacetylase, a ubiquitin-ligase, a deubiquitinase, a kinase, a phosphatase, a NEDD8-ligase, a de-NEDDylase, a SUMO-ligase, a deSUMOylase, a histone deacetylase, a histone acetyltransferase, a histone methyltransferase, or a histone demethylase. 197. The Type II Cas protein of any one of embodiments 166 to 196, which comprises a means for deaminating adenosine, optionally wherein the means for deaminating adenosine is an adenosine deaminase. 198. The Type II Cas protein of any one of embodiments 166 to 196, which comprises a fusion partner which is an adenosine deaminase, optionally wherein the amino acid sequence of the adenosine deaminase comprises an amino acid sequence having at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, at least 95%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with SEQ ID NO:137, optionally wherein the adenosine deaminase is the adenosine deaminase moiety contained in the adenine base editor ABE8e. 199. The Type II Cas protein of any one of embodiments 166 to 196, which comprises a means for deaminating cytidine, optionally wherein the means for deaminating cytidine is a cytidine deaminase. 200. The Type II Cas protein of any one of embodiments 166 to 196, which comprises a fusion partner which is a cytidine deaminase. 201. The Type II Cas protein of any one of embodiments 166 to 196, which comprises a means for synthesizing DNA from a single-stranded template, optionally wherein the means for synthesizing DNA from a single-stranded template is a reverse transcriptase. 202. The Type II Cas protein of any one of embodiments 166 to 196, which comprises a fusion partner which is a reverse transcriptase. 203. The Type II Cas protein of any one of embodiments 166 to 202, which comprises a tag. 204. The Type II Cas protein of embodiment 203, wherein the tag is a SV5 tag, optionally wherein the SV5 tag comprises the amino acid sequence GKPIPNPLLGLDST (SEQ ID NO:135) or IPNPLLGLD (SEQ ID NO:136). 205. The Type II Cas protein of any one of embodiments 1 to 204, wherein the reference protein sequence is SEQ ID NO:1. 206. The Type II Cas protein of embodiment 205, whose amino acid sequence comprises the amino acid sequence of SEQ ID NO:1. 207. The Type II Cas protein of any one of embodiments 1 to 204, wherein the reference protein sequence is SEQ ID NO:2. 208. The Type II Cas protein of any one of embodiments 205 to 207, whose amino acid sequence comprises the amino acid sequence of SEQ ID NO:2. 209. The Type II Cas protein of embodiment 205 or embodiment 206, whose amino acid sequence comprises the amino acid sequence of SEQ ID NO:3. 210. The Type II Cas protein of any one of embodiments 1 to 204, wherein the reference protein sequence is SEQ ID NO:7. 211. The Type II Cas protein of embodiment 210, whose amino acid sequence comprises the amino acid sequence of SEQ ID NO:7. 212. The Type II Cas protein of any one of embodiments 1 to 204, wherein the reference protein sequence is SEQ ID NO:8. 213. The Type II Cas protein of any one of embodiments 210 to 212, whose amino acid sequence comprises the amino acid sequence of SEQ ID NO:8. 214. The Type II Cas protein of embodiment 210 or embodiment 211, whose amino acid sequence comprises the amino acid sequence of SEQ ID NO:9. 215. The Type II Cas protein of any one of embodiments 1 to 204, wherein the reference protein sequence is SEQ ID NO:13. 216. The Type II Cas protein of embodiment 215, whose amino acid sequence comprises the amino acid sequence of SEQ ID NO:13. 217. The Type II Cas protein of any one of embodiments 1 to 204, wherein the reference protein sequence is SEQ ID NO:14. 218. The Type II Cas protein of any one of embodiments 215 to 217, whose amino acid sequence comprises the amino acid sequence of SEQ ID NO:14. 219. The Type II Cas protein of embodiment 215 or embodiment 216, whose amino acid sequence comprises the amino acid sequence of SEQ ID NO:15. 220. The Type II Cas protein of any one of embodiments 1 to 204, wherein the reference protein sequence is SEQ ID NO:19. 221. The Type II Cas protein of embodiment 220, whose amino acid sequence comprises the amino acid sequence of SEQ ID NO:19. 222. The Type II Cas protein of any one of embodiments 1 to 204, wherein the reference protein sequence is SEQ ID NO:20. 223. The Type II Cas protein of any one of embodiments 220 to 222, whose amino acid sequence comprises the amino acid sequence of SEQ ID NO:20. 224. The Type II Cas protein of embodiment 220 or embodiment 221, whose amino acid sequence comprises the amino acid sequence of SEQ ID NO:21. 225. The Type II Cas protein of any one of embodiments 1 to 204, wherein the reference protein sequence is SEQ ID NO:25. 226. The Type II Cas protein of embodiment 225, whose amino acid sequence comprises the amino acid sequence of SEQ ID NO:25. 227. The Type II Cas protein of any one of embodiments 1 to 204, wherein the reference protein sequence is SEQ ID NO:26. 228. The Type II Cas protein of any one of embodiments 225 to 227, whose amino acid sequence comprises the amino acid sequence of SEQ ID NO:26. 229. The Type II Cas protein of embodiment 226 or embodiment 227, whose amino acid sequence comprises the amino acid sequence of SEQ ID NO:27. 230. The Type II Cas protein of any one of embodiments 1 to 204, wherein the reference protein sequence is SEQ ID NO:31. 231. The Type II Cas protein of embodiment 230, whose amino acid sequence comprises the amino acid sequence of SEQ ID NO:31. 232. The Type II Cas protein of any one of embodiments 1 to 204, wherein the reference protein sequence is SEQ ID NO:32. 233. The Type II Cas protein of any one of embodiments 231 to 232, whose amino acid sequence comprises the amino acid sequence of SEQ ID NO:32. 234. The Type II Cas protein of embodiment 230 or embodiment 231, whose amino acid sequence comprises the amino acid sequence of SEQ ID NO:33. 235. The Type II Cas protein of any one of embodiments 1 to 204, wherein the reference protein sequence is SEQ ID NO:37. 236. The Type II Cas protein of embodiment 235, whose amino acid sequence comprises the amino acid sequence of SEQ ID NO:37. 237. The Type II Cas protein of any one of embodiments 1 to 204, wherein the reference protein sequence is SEQ ID NO:38. 238. The Type II Cas protein of any one of embodiments 235 to 237, whose amino acid sequence comprises the amino acid sequence of SEQ ID NO:38. 239. The Type II Cas protein of embodiment 235 or embodiment 236, whose amino acid sequence comprises the amino acid sequence of SEQ ID NO:39. 240. The Type II Cas protein of any one of embodiments 1 to 204, wherein the reference protein sequence is SEQ ID NO:43. 241. The Type II Cas protein of embodiment 240, whose amino acid sequence comprises the amino acid sequence of SEQ ID NO:43. 242. The Type II Cas protein of any one of embodiments 1 to 204, wherein the reference protein sequence is SEQ ID NO:44. 243. The Type II Cas protein of any one of embodiments 240 to 242, whose amino acid sequence comprises the amino acid sequence of SEQ ID NO:44. 244. The Type II Cas protein of embodiment 240 or embodiment 241, whose amino acid sequence comprises the amino acid sequence of SEQ ID NO:45. 245. The Type II Cas protein of any one of embodiments 1 to 204, wherein the reference protein sequence is SEQ ID NO:49. 246. The Type II Cas protein of embodiment 245, whose amino acid sequence comprises the amino acid sequence of SEQ ID NO:49. 247. The Type II Cas protein of any one of embodiments 1 to 204, wherein the reference protein sequence is SEQ ID NO:50. 248. The Type II Cas protein of any one of embodiments 245 to 247, whose amino acid sequence comprises the amino acid sequence of SEQ ID NO:50. 249. The Type II Cas protein of embodiment 245 or embodiment 246, whose amino acid sequence comprises the amino acid sequence of SEQ ID NO:51. 250. The Type II Cas protein of any one of embodiments 1 to 204, wherein the reference protein sequence is SEQ ID NO:55. 251. The Type II Cas protein of embodiment 250, whose amino acid sequence comprises the amino acid sequence of SEQ ID NO:55. 252. The Type II Cas protein of any one of embodiments 1 to 204, wherein the reference protein sequence is SEQ ID NO:56. 253. The Type II Cas protein of any one of embodiments 250 to 252, whose amino acid sequence comprises the amino acid sequence of SEQ ID NO:56. 254. The Type II Cas protein of embodiment 250 or embodiment 251, whose amino acid sequence comprises the amino acid sequence of SEQ ID NO:57. 255. A Type II Cas protein whose amino acid sequence is identical to a Type II Cas protein of any one of embodiments 1 to 254 except for one or more amino acid substitutions relative to the reference sequence that provide nickase activity, optionally wherein the one or more amino acid substitutions comprise a substitution (e.g., alanine substitution) at a position corresponding to position D10 of SpCas9 or H840A of SpCas9 (e.g., as shown in Table 3). 256. The Type II Cas of embodiment 255, wherein the one or more amino acid substitutions that provide nickase activity are in a RuvC or HNH domain. 257. The Type II Cas of embodiment 255, wherein the one or more amino acid substitutions relative to the reference sequence that provide nickase activity comprise a D12A substitution, wherein the position of the D12A substitution is defined with respect to the amino acid numbering of SEQ ID NO:2. 258. The Type II Cas of embodiment 255, wherein the one or more amino acid substitutions relative to the reference sequence that provide nickase activity comprise an H868A substitution, wherein the position of the H868A substitution is defined with respect to the amino acid numbering of SEQ ID NO:2. 259. The Type II Cas of embodiment 255, wherein the one or more amino acid substitutions relative to the reference sequence that provide nickase activity comprise a D18A substitution, wherein the position of the D18A substitution is defined with respect to the amino acid numbering of SEQ ID NO:8. 260. The Type II Cas of embodiment 255, wherein the one or more amino acid substitutions relative to the reference sequence that provide nickase activity comprise an H845A substitution, wherein the position of the H845A substitution is defined with respect to the amino acid numbering of SEQ ID NO:8. 261. The Type II Cas of embodiment 255, wherein the one or more amino acid substitutions relative to the reference sequence that provide nickase activity comprise a D14A substitution, wherein the position of the D14A substitution is defined with respect to the amino acid numbering of SEQ ID NO:14. 262. The Type II Cas of embodiment 255, wherein the one or more amino acid substitutions relative to the reference sequence that provide nickase activity comprise an H900A substitution, wherein the position of the H900A substitution is defined with respect to the amino acid numbering of SEQ ID NO:14. 263. The Type II Cas of embodiment 255, wherein the one or more amino acid substitutions relative to the reference sequence that provide nickase activity comprise a D15A substitution, wherein the position of the D15A substitution is defined with respect to the amino acid numbering of SEQ ID NO:20. 264. The Type II Cas of embodiment 255, wherein the one or more amino acid substitutions relative to the reference sequence that provide nickase activity comprise an H890A substitution, wherein the position of the H890A substitution is defined with respect to the amino acid numbering of SEQ ID NO:20. 265. The Type II Cas of embodiment 255, wherein the one or more amino acid substitutions relative to the reference sequence that provide nickase activity comprise a D15A substitution, wherein the position of the D15A substitution is defined with respect to the amino acid numbering of SEQ ID NO:26. 266. The Type II Cas of embodiment 255, wherein the one or more amino acid substitutions relative to the reference sequence that provide nickase activity comprise an H898A substitution, wherein the position of the H898A substitution is defined with respect to the amino acid numbering of SEQ ID NO:26. 267. The Type II Cas of embodiment 255, wherein the one or more amino acid substitutions relative to the reference sequence that provide nickase activity comprise a D17A substitution, wherein the position of the D17A substitution is defined with respect to the amino acid numbering of SEQ ID NO:32. 268. The Type II Cas of embodiment 255, wherein the one or more amino acid substitutions relative to the reference sequence that provide nickase activity comprise an H836A substitution, wherein the position of the H836A substitution is defined with respect to the amino acid numbering of SEQ ID NO:32. 269. The Type II Cas of embodiment 255, wherein the one or more amino acid substitutions relative to the reference sequence that provide nickase activity comprise a D15A substitution, wherein the position of the D15A substitution is defined with respect to the amino acid numbering of SEQ ID NO:38. 270. The Type II Cas of embodiment 255, wherein the one or more amino acid substitutions relative to the reference sequence that provide nickase activity comprise an H893A substitution, wherein the position of the H893A substitution is defined with respect to the amino acid numbering of SEQ ID NO:38. 271. The Type II Cas of embodiment 255, wherein the one or more amino acid substitutions relative to the reference sequence that provide nickase activity comprise a D14A substitution, wherein the position of the D14A substitution is defined with respect to the amino acid numbering of SEQ ID NO:44. 272. The Type II Cas of embodiment 255, wherein the one or more amino acid substitutions relative to the reference sequence that provide nickase activity comprise an H878A substitution, wherein the position of the H878A substitution is defined with respect to the amino acid numbering of SEQ ID NO:44. 273. The Type II Cas of embodiment 255, wherein the one or more amino acid substitutions relative to the reference sequence that provide nickase activity comprise a D18A substitution, wherein the position of the D18A substitution is defined with respect to the amino acid numbering of SEQ ID NO:50. 274. The Type II Cas of embodiment 255, wherein the one or more amino acid substitutions relative to the reference sequence that provide nickase activity comprise a H857A substitution, wherein the position of the H857A substitution is defined with respect to the amino acid numbering of SEQ ID NO:50. 275. The Type II Cas of embodiment 255, wherein the one or more amino acid substitutions relative to the reference sequence that provide nickase activity comprise a D15A substitution, wherein the position of the D15A substitution is defined with respect to the amino acid numbering of SEQ ID NO:56. 276. The Type II Cas of embodiment 255, wherein the one or more amino acid substitutions relative to the reference sequence that provide nickase activity comprise an H890A substitution, wherein the position of the H890A substitution is defined with respect to the amino acid numbering of SEQ ID NO:56. 277. A Type II Cas protein whose amino acid sequence is identical to a Type II Cas protein of any one of embodiments 1 to 254 except for one or more amino acid substitutions relative to the reference sequence that render the Type II Cas protein catalytically inactive, optionally wherein the one or more amino acid substitutions comprise a substitution (e.g., alanine substitution) at a position corresponding to position D10 of SpCas9 or H840 of SpCas9 (e.g., as shown in Table 3), or a combination thereof. 278. The Type II Cas protein of embodiment 277, wherein the one or more amino acid substitutions that render the Type II Cas protein catalytically inactive comprise D12A and H868A substitutions, wherein the positions of the D12A and H868A substitutions are defined with respect to the amino acid numbering of SEQ ID NO:2. 279. The Type II Cas protein of embodiment 277, wherein the one or more amino acid substitutions that render the Type II Cas protein catalytically inactive comprise D18A and H845A substitutions, wherein the positions of the D18A and H845A substitutions are defined with respect to the amino acid numbering of SEQ ID NO:8. 280. The Type II Cas protein of embodiment 277, wherein the one or more amino acid substitutions that render the Type II Cas protein catalytically inactive comprise D14A and H900A substitutions, wherein the positions of the D14A and H900A substitutions are defined with respect to the amino acid numbering of SEQ ID NO:14. 281. The Type II Cas protein of embodiment 277, wherein the one or more amino acid substitutions that render the Type II Cas protein catalytically inactive comprise D15A and H890A substitutions, wherein the positions of the D15A and H890A substitutions are defined with respect to the amino acid numbering of SEQ ID NO:20. 282. The Type II Cas protein of embodiment 277, wherein the one or more amino acid substitutions that render the Type II Cas protein catalytically inactive comprise D15A and H898A substitutions, wherein the positions of the D15A and H898A substitutions are defined with respect to the amino acid numbering of SEQ ID NO:26. 283. The Type II Cas protein of embodiment 277, wherein the one or more amino acid substitutions that render the Type II Cas protein catalytically inactive comprise D17A and H836A substitutions, wherein the positions of the D17A and H836A substitutions are defined with respect to the amino acid numbering of SEQ ID NO:32 284. The Type II Cas protein of embodiment 277, wherein the one or more amino acid substitutions that render the Type II Cas protein catalytically inactive comprise D15A and H893A substitutions, wherein the positions of the D15A and H893A substitutions are defined with respect to the amino acid numbering of SEQ ID NO:38. 285. The Type II Cas protein of embodiment 277, wherein the one or more amino acid substitutions that render the Type II Cas protein catalytically inactive comprise D14A and H878A substitutions, wherein the positions of the D14A and H878A substitutions are defined with respect to the amino acid numbering of SEQ ID NO:44. 286. The Type II Cas protein of embodiment 277, wherein the one or more amino acid substitutions that render the Type II Cas protein catalytically inactive comprise D18A and H857A substitutions, wherein the positions of the D18A and H857A substitutions are defined with respect to the amino acid numbering of SEQ ID NO:50. 287. The Type II Cas protein of embodiment 277, wherein the one or more amino acid substitutions that render the Type II Cas protein catalytically inactive comprise D15A and H890A substitutions, wherein the positions of the D15A and H890A substitutions are defined with respect to the amino acid numbering of SEQ ID NO:56. 288. A DGHJ Type II Cas guide RNA (gRNA) molecule. 289. A CUAZ Type II Cas guide RNA (gRNA) molecule. 290. An AHWY Type II Cas guide RNA (gRNA) molecule. 291. A CBGI Type II Cas guide RNA (gRNA) molecule. 292. An ASDR Type II Cas guide RNA (gRNA) molecule. 293. A BCZZ Type II Cas guide RNA (gRNA) molecule. 294. A DRCY Type II Cas guide RNA (gRNA) molecule. 295. A DRPY Type II Cas guide RNA (gRNA) molecule. 296. An EAJR Type II Cas guide RNA (gRNA) molecule. 297. A CFDE Type II Cas guide RNA (gRNA) molecule. 298. The gRNA of any one of embodiments 288 to 297, which is a gRNA for editing a human RHO gene. 299. The gRNA of any one of embodiments 288 to 297, which is a gRNA for editing a human B2M gene. 300. The gRNA of any one of embodiments 288 to 297, which is a gRNA for editing a human TRAC gene. 301. The gRNA of any one of embodiments 288 to 297, which is a gRNA for editing a human LAG3 gene. 302. The gRNA of any one of embodiments 288 to 297, which is a gRNA for editing a human PD1 gene. 303. A guide RNA (gRNA) molecule for editing a human B2M gene comprising a spacer whose nucleotide sequence comprises 15 or more consecutive nucleotides of a reference sequence or comprises a nucleotide sequence that is at least 85% identical to the reference sequence, wherein the reference sequence is selected from SEQ ID NOs:138-162. 304. A guide RNA (gRNA) molecule for editing a human TRAC gene comprising a spacer whose nucleotide sequence comprises 15 or more consecutive nucleotides of a reference sequence or comprises a nucleotide sequence that is at least 85% identical to the reference sequence, wherein the reference sequence is selected from SEQ ID NOs:163-194. 305. A guide RNA (gRNA) molecule for editing a human PD1 gene comprising a spacer whose nucleotide sequence comprises 15 or more consecutive nucleotides of a reference sequence or comprises a nucleotide sequence that is at least 85% identical to the reference sequence, wherein the reference sequence is selected from SEQ ID NOs:195-217. 306. The gRNA of any one of embodiments 303 to 305, which comprises a spacer that is 15 to 30 nucleotides in length. 307. The gRNA of embodiment 306, wherein the spacer is 18 to 30 nucleotides in length. 308. The gRNA of embodiment 306, wherein the spacer is 20 to 28 nucleotides in length. 309. The gRNA of embodiment 306, wherein the spacer is 22 to 26 nucleotides in length. 310. The gRNA of embodiment 306, wherein the spacer is 23 to 25 nucleotides in length. 311. The gRNA of embodiment 306, wherein the spacer is 22 to 25 nucleotides in length. 312. The gRNA of embodiment 306, wherein the spacer is 15 to 25 nucleotides in length. 313. The gRNA of embodiment 306, wherein the spacer is 16 to 24 nucleotides in length. 314. The gRNA of embodiment 306, wherein the spacer is 17 to 23 nucleotides in length. 315. The gRNA of embodiment 306, wherein the spacer is 18 to 22 nucleotides in length. 316. The gRNA of embodiment 306, wherein the spacer is 19 to 21 nucleotides in length. 317. The gRNA of embodiment 306, wherein the spacer is 25 nucleotides in length. 318. The gRNA of embodiment 306, wherein the spacer is 24 nucleotides in length. 319. The gRNA of embodiment 306, wherein the spacer is 23 nucleotides in length. 320. The gRNA of embodiment 306, wherein the spacer is 22 nucleotides in length. 321. The gRNA of embodiment 306, wherein the spacer is 21 nucleotides in length. 322. The gRNA of embodiment 306, wherein the spacer is 20 nucleotides in length. 323. The gRNA of any one of embodiments 303 to 322, wherein the spacer comprises 16 or more consecutive nucleotides of the reference sequence. 324. The gRNA of any one of embodiments 303 to 322, wherein the spacer comprises 17 or more consecutive nucleotides of the reference sequence. 325. The gRNA of any one of embodiments 303 to 322, wherein the spacer comprises 18 or more consecutive nucleotides of the reference sequence. 326. The gRNA of any one of embodiments 303 to 322, wherein the spacer comprises 19 or more consecutive nucleotides of the reference sequence. 327. The gRNA of any one of embodiments 303 to 322, wherein the spacer comprises 20 consecutive nucleotides of the reference sequence. 328. The gRNA of any one of embodiments 303 to 321, wherein the reference sequence is a reference sequence having at least 21 and the spacer comprises 21 consecutive nucleotides of the reference sequence. 329. The gRNA of any one of embodiments 303 to 320, wherein the reference sequence is a reference sequence having at least 22 and the spacer comprises 22 consecutive nucleotides of the reference sequence. 330. The gRNA of any one of embodiments 303 to 319, wherein the reference sequence is a reference sequence having at least 23 nucleotides and the spacer comprises 23 consecutive nucleotides of the reference sequence. 331. The gRNA of any one of embodiments 303 to 318, wherein the reference sequence is a reference sequence having at least 24 nucleotides and the spacer comprises 24 consecutive nucleotides of the reference sequence. 332. The gRNA of any one of embodiments 303 to 322, wherein the spacer comprises a nucleotide sequence that is at least 90% identical to the reference sequence. 333. The gRNA of embodiment 332, wherein the spacer comprises a nucleotide sequence that is at least 95% identical to the reference sequence. 334. The gRNA of any one of embodiments 303 to 322, wherein the spacer comprises a nucleotide sequence that has one mismatch relative to the reference sequence. 335. The gRNA of any one of embodiments 303 to 322, wherein the spacer comprises a nucleotide sequence that has two mismatches relative to the reference sequence. 336. The gRNA of any one of embodiments 303 to 322, wherein the spacer comprises the reference sequence. 337. The gRNA of any one of embodiments 303 and 306 to 336 when depending from embodiment 303, wherein the reference sequence is CAGAAAGAGAGAGUAGCGCGAGC (SEQ ID NO:138). 338. The gRNA of any one of embodiments 303 and 306 to 336 when depending from embodiment 303, wherein the reference sequence is CCGUGGCCUUAGCUGUGCUCGCG (SEQ ID NO:139). 339. The gRNA of any one of embodiments 303 and 306 to 336 when depending from embodiment 303, wherein the reference sequence is UCAGGUACUCCAAAGAUUCAGGU (SEQ ID NO:140). 340. The gRNA of any one of embodiments 303 and 306 to 336 when depending from embodiment 303, wherein the reference sequence is GUACUCCAAAGAUUCAGGUUUAC (SEQ ID NO:141). 341. The gRNA of any one of embodiments 303 and 306 to 336 when depending from embodiment 303, wherein the reference sequence is GACACAUAGCAAUUCAGGAAAUU (SEQ ID NO:142). 342. The gRNA of any one of embodiments 303 and 306 to 336 when depending from embodiment 303, wherein the reference sequence is ACUCUCUCUUUCUGGCCUGGAGG (SEQ ID NO:143). 343. The gRNA of any one of embodiments 303 and 306 to 336 when depending from embodiment 303, wherein the reference sequence is CCAAAGAUUCAGGUUUACUCACG (SEQ ID NO:144). 344. The gRNA of any one of embodiments 303 and 306 to 336 when depending from embodiment 303, wherein the reference sequence is CUGAAUUGCUAUGUGUCUGGGUU (SEQ ID NO:145). 345. The gRNA of any one of embodiments 303 and 306 to 336 when depending from embodiment 303, wherein the reference sequence is AUUGCUAUGUGUCUGGGUUUCAU (SEQ ID NO:146). 346. The gRNA of any one of embodiments 303 and 306 to 336 when depending from embodiment 303, wherein the reference sequence is GUCAACUUCAAUGUCGGAUGGAUG (SEQ ID NO:147). 347. The gRNA of any one of embodiments 303 and 306 to 336 when depending from embodiment 303, wherein the reference sequence is ACUUCAAUGUCGGAUGGAUG (SEQ ID NO:148). 348. The gRNA of any one of embodiments 303 and 306 to 336 when depending from embodiment 303, wherein the reference sequence is GAUAGCCUCCAGGCCAGAAAGAGA (SEQ ID NO:149). 349. The gRNA of any one of embodiments 303 and 306 to 336 when depending from embodiment 303, wherein the reference sequence is GGAGGGUAGGAGAGACUCACGCU (SEQ ID NO:150). 350. The gRNA of any one of embodiments 303 and 306 to 336 when depending from embodiment 303, wherein the reference sequence is GACGUGAGUAAACCUGAAUCUUUG (SEQ ID NO:151). 351. The gRNA of any one of embodiments 303 and 306 to 336 when depending from embodiment 303, wherein the reference sequence is UUCCAUUCUCUGCUGGAUGACGU (SEQ ID NO:152). 352. The gRNA of any one of embodiments 303 and 306 to 336 when depending from embodiment 303, wherein the reference sequence is GUCGGAUGGAUGAAACCCAGAC (SEQ ID NO:153). 353. The gRNA of any one of embodiments 303 and 306 to 336 when depending from embodiment 303, wherein the reference sequence is GAUAGCCUCCAGGCCAGAAAGAG (SEQ ID NO:154). 354. The gRNA of any one of embodiments 303 and 306 to 336 when depending from embodiment 303, wherein the reference sequence is UCUGGCCUGGAGGCUAUCCAGCG (SEQ ID NO:155). 355. The gRNA of any one of embodiments 303 and 306 to 336 when depending from embodiment 303, wherein the reference sequence is GACGUGAGUAAACCUGAAUCUUU (SEQ ID NO:156). 356. The gRNA of any one of embodiments 303 and 306 to 336 when depending from embodiment 303, wherein the reference sequence is GUUUACUCACGUCAUCCAGCAG (SEQ ID NO:157). 357. The gRNA of any one of embodiments 303 and 306 to 336 when depending from embodiment 303, wherein the reference sequence is GAGAAUGGAAAGUCAAAUUUCC (SEQ ID NO:158). 358. The gRNA of any one of embodiments 303 and 306 to 336 when depending from embodiment 303, wherein the reference sequence is UUCAGUGGGGGUGAAUUCAGUGU (SEQ ID NO:159). 359. The gRNA of any one of embodiments 303 and 306 to 336 when depending from embodiment 303, wherein the reference sequence is GCCAGAAAGAGAGAGUAGCGCG (SEQ ID NO:160). 360. The gRNA of any one of embodiments 303 and 306 to 336 when depending from embodiment 303, wherein the reference sequence is CAAUGUCGGAUGGAUGAAACCCA (SEQ ID NO:161). 361. The gRNA of any one of embodiments 303 and 306 to 336 when depending from embodiment 303, wherein the reference sequence is GCAAGGACUGGUCUUUCUAUCUCU (SEQ ID NO:162). 362. The gRNA of any one of embodiments 304 and 306 to 336 when depending from embodiment 304, wherein the reference sequence is AUCAAAAUCGGUGAAUAGGCAGAC (SEQ ID NO:163). 363. The gRNA of any one of embodiments 304 and 306 to 336 when depending from embodiment 304, wherein the reference sequence is ACACAUCAGAAUCCUUACUUUGUG (SEQ ID NO:164). 364. The gRNA of any one of embodiments 304 and 306 to 336 when depending from embodiment 304, wherein the reference sequence is GCACUGUUGCUCUUGAAGUCCAU (SEQ ID NO:165). 365. The gRNA of any one of embodiments 304 and 306 to 336 when depending from embodiment 304, wherein the reference sequence is AACUGUGCUAGACAUGAGGUCUAU (SEQ ID NO:166). 366. The gRNA of any one of embodiments 304 and 306 to 336 when depending from embodiment 304, wherein the reference sequence is GUAUAUCACAGACAAAACUGUGCU (SEQ ID NO:167). 367. The gRNA of any one of embodiments 304 and 306 to 336 when depending from embodiment 304, wherein the reference sequence is UCUUGUCCCACAGAUAUCCA (SEQ ID NO:168). 368. The gRNA of any one of embodiments 304 and 306 to 336 when depending from embodiment 304, wherein the reference sequence is CCCACAGAUAUCCAGAACCC (SEQ ID NO:169). 369. The gRNA of any one of embodiments 304 and 306 to 336 when depending from embodiment 304, wherein the reference sequence is GUGUACCAGCUGAGAGACUCU (SEQ ID NO:170). 370. The gRNA of any one of embodiments 304 and 306 to 336 when depending from embodiment 304, wherein the reference sequence is UGUCUGUGAUAUACACAUCA (SEQ ID NO:171). 371. The gRNA of any one of embodiments 304 and 306 to 336 when depending from embodiment 304, wherein the reference sequence is GCCACAGCACUGUUGCUCUUG (SEQ ID NO:172). 372. The gRNA of any one of embodiments 304 and 306 to 336 when depending from embodiment 304, wherein the reference sequence is GAUAUCCAGAACCCUGACCCUGCC (SEQ ID NO:173). 373. The gRNA of any one of embodiments 304 and 306 to 336 when depending from embodiment 304, wherein the reference sequence is CACUGGAUUUAGAGUCUCUCAGC (SEQ ID NO:174). 374. The gRNA of any one of embodiments 304 and 306 to 336 when depending from embodiment 304, wherein the reference sequence is UUUGUUUGAGAAUCAAAAUCGGU (SEQ ID NO:175). 375. The gRNA of any one of embodiments 304 and 306 to 336 when depending from embodiment 304, wherein the reference sequence is GAUUCUCAAACAAAUGUGUCACA (SEQ ID NO:176). 376. The gRNA of any one of embodiments 304 and 306 to 336 when depending from embodiment 304, wherein the reference sequence is GUCACAAAGUAAGGAUUCUGAU (SEQ ID NO:177). 377. The gRNA of any one of embodiments 304 and 306 to 336 when depending from embodiment 304, wherein the reference sequence is GCCGUGUACCAGCUGAGAGACUC (SEQ ID NO:178). 378. The gRNA of any one of embodiments 304 and 306 to 336 when depending from embodiment 304, wherein the reference sequence is GCUGAGAGACUCUAAAUCCAGUGA (SEQ ID NO:179). 379. The gRNA of any one of embodiments 304 and 306 to 336 when depending from embodiment 304, wherein the reference sequence is GCAGACAGACUUGUCACUGGAUUU (SEQ ID NO:180). 380. The gRNA of any one of embodiments 304 and 306 to 336 when depending from embodiment 304, wherein the reference sequence is AUUCACCGAUUUUGAUUCUCAAA (SEQ ID NO:181). 381. The gRNA of any one of embodiments 304 and 306 to 336 when depending from embodiment 304, wherein the reference sequence is AUUUGUUUGAGAAUCAAAAUCGG (SEQ ID NO:182). 382. The gRNA of any one of embodiments 304 and 306 to 336 when depending from embodiment 304, wherein the reference sequence is UGAUUCUCAAACAAAUGUGUCAC (SEQ ID NO:183). 383. The gRNA of any one of embodiments 304 and 306 to 336 when depending from embodiment 304, wherein the reference sequence is UUACUUUGUGACACAUUUGUUUG (SEQ ID NO:184). 384. The gRNA of any one of embodiments 304 and 306 to 336 when depending from embodiment 304, wherein the reference sequence is GUUUUGUCUGUGAUAUACACAUC (SEQ ID NO:185). 385. The gRNA of any one of embodiments 304 and 306 to 336 when depending from embodiment 304, wherein the reference sequence is CAGGCCACAGCACUGUUGCUCUU (SEQ ID NO:186). 386. The gRNA of any one of embodiments 304 and 306 to 336 when depending from embodiment 304, wherein the reference sequence is gAACAGUGCUGUGGCCUGGAGCAA (SEQ ID NO:187). 387. The gRNA of any one of embodiments 304 and 306 to 336 when depending from embodiment 304, wherein the reference sequence is ACUGGAUUUAGAGUCUCUCAGCU (SEQ ID NO:188). 388. The gRNA of any one of embodiments 304 and 306 to 336 when depending from embodiment 304, wherein the reference sequence is GCCUAUUCACCGAUUUUGAUUCU (SEQ ID NO:189). 389. The gRNA of any one of embodiments 304 and 306 to 336 when depending from embodiment 304, wherein the reference sequence is ACACAUCAGAAUCCUUACUUUGU (SEQ ID NO:190). 390. The gRNA of any one of embodiments 304 and 306 to 336 when depending from embodiment 304, wherein the reference sequence is CUAGCACAGUUUUGUCUGUGAUA (SEQ ID NO:191). 391. The gRNA of any one of embodiments 304 and 306 to 336 when depending from embodiment 304, wherein the reference sequence is UGAAGUCCAUAGACCUCAUGUCU (SEQ ID NO:192). 392. The gRNA of any one of embodiments 304 and 306 to 336 when depending from embodiment 304, wherein the reference sequence is GUCUAGCACAGUUUUGUCUGUGA (SEQ ID NO:193). 393. The gRNA of any one of embodiments 304 and 306 to 336 when depending from embodiment 304, wherein the reference sequence is AUAAUGCUGUUGUUGAAGGCGUU (SEQ ID NO:194). 394. The gRNA of any one of embodiments 305 and 306 to 336 when depending from embodiment 305, wherein the reference sequence is AUCCUGGCCGCCAGCCCAGUUGUA (SEQ ID NO:195). 395. The gRNA of any one of embodiments 305 and 306 to 336 when depending from embodiment 305, wherein the reference sequence is GGGGCUGCUCCAGGCAUGCAGAUC (SEQ ID NO:196). 396. The gRNA of any one of embodiments 305 and 306 to 336 when depending from embodiment 305, wherein the reference sequence is GCCCAGUUGUAGCACCGCCCAGA (SEQ ID NO:197). 397. The gRNA of any one of embodiments 305 and 306 to 336 when depending from embodiment 305, wherein the reference sequence is GGCCAGUCGUCUGGGCGGUGCUA (SEQ ID NO:198). 398. The gRNA of any one of embodiments 305 and 306 to 336 when depending from embodiment 305, wherein the reference sequence is GUGGGGCUGCUCCAGGCAUGC (SEQ ID NO:199). 399. The gRNA of any one of embodiments 305 and 306 to 336 when depending from embodiment 305, wherein the reference sequence is CGACCCCACCUACCUAAGAA (SEQ ID NO:200). 400. The gRNA of any one of embodiments 305 and 306 to 336 when depending from embodiment 305, wherein the reference sequence is CAGGCAUGCAGAUCCCACAG (SEQ ID NO:201). 401. The gRNA of any one of embodiments 305 and 306 to 336 when depending from embodiment 305, wherein the reference sequence is GCCGCCAGCCCAGUUGUAGCA (SEQ ID NO:202). 402. The gRNA of any one of embodiments 305 and 306 to 336 when depending from embodiment 305, wherein the reference sequence is CUAAGAACCAUCCUGGCCGC (SEQ ID NO:203). 403. The gRNA of any one of embodiments 305 and 306 to 336 when depending from embodiment 305, wherein the reference sequence is UCAGACUCCCCAGACAGGCCCUG (SEQ ID NO:204). 404. The gRNA of any one of embodiments 305 and 306 to 336 when depending from embodiment 305, wherein the reference sequence is CGGGGCCUCCGAGGCCGCACCUG (SEQ ID NO:205). 405. The gRNA of any one of embodiments 305 and 306 to 336 when depending from embodiment 305, wherein the reference sequence is GAACCAUCCUGGCCGCCAGCCCAG (SEQ ID NO:206). 406. The gRNA of any one of embodiments 305 and 306 to 336 when depending from embodiment 305, wherein the reference sequence is GGCUGGCGGCCAGGAUGGUUCUU (SEQ ID NO:207). 407. The gRNA of any one of embodiments 305 and 306 to 336 when depending from embodiment 305, wherein the reference sequence is CAUCGGAGAGCUUCGUGCUAAAC (SEQ ID NO:208). 408. The gRNA of any one of embodiments 305 and 306 to 336 when depending from embodiment 305, wherein the reference sequence is CCGUCUGGUUGCUGGGGCUCAUG (SEQ ID NO:209). 409. The gRNA of any one of embodiments 305 and 306 to 336 when depending from embodiment 305, wherein the reference sequence is GGGCCAGGGAGAUGGCCCCACAG (SEQ ID NO:210). 410. The gRNA of any one of embodiments 305 and 306 to 336 when depending from embodiment 305, wherein the reference sequence is GGGGGGUUCCAGGGCCUGUCUGG (SEQ ID NO:211). 411. The gRNA of any one of embodiments 305 and 306 to 336 when depending from embodiment 305, wherein the reference sequence is CGGGCCCUGACCACGCUCAUGUG (SEQ ID NO:212). 412. The gRNA of any one of embodiments 305 and 306 to 336 when depending from embodiment 305, wherein the reference sequence is GCCACCUUCACCUGCAGCUUCUCC (SEQ ID NO:213). 413. The gRNA of any one of embodiments 305 and 306 to 336 when depending from embodiment 305, wherein the reference sequence is GGCCAGGACUGCCGCUUCCGUGU (SEQ ID NO:214). 414. The gRNA of any one of embodiments 305 and 306 to 336 when depending from embodiment 305, wherein the reference sequence is UGGGGCUCAUGCGGUACCAGUUU (SEQ ID NO:215). 415. The gRNA of any one of embodiments 305 and 306 to 336 when depending from embodiment 305, wherein the reference sequence is CAACACAUCGGAGAGCUUCGUGC (SEQ ID NO:216). 416. The gRNA of any one of embodiments 305 and 306 to 336 when depending from embodiment 305, wherein the reference sequence is GUCACGCCCGUUGGGCAGUUGUGU (SEQ ID NO:217). 417. The gRNA of any one of embodiments 288 to 416, which is a single guide RNA (sgRNA). 418. A gRNA comprising a spacer and a sgRNA scaffold, which is optionally a gRNA according to any one of embodiments 288 to 417, wherein: (a) WKH^VSDFHU^LV^SRVLWLRQHG^^¶^WR^WKH^VJ51$^VFDIIROG^^DQG (b) the nucleotide sequence of the sgRNA scaffold comprises a nucleotide sequence that is at least 50% identical to a reference scaffold sequence, wherein the reference scaffold sequence is any one of SEQ ID NOS:81-102. 419. A gRNA comprising a means for binding a target mammalian genomic sequence and a sgRNA scaffold, optionally wherein the means for binding a target mammalian genomic sequence is a spacer, wherein: (a) WKH^PHDQV^IRU^ELQGLQJ^D^WDUJHW^JHQRPLF^VHTXHQFH^LV^SRVLWLRQHG^^¶^WR^WKH^VJ51$^VFDIIROG^^ and (b) the nucleotide sequence of the sgRNA scaffold comprises a nucleotide sequence that is at least 50% identical to a reference scaffold sequence, wherein the reference scaffold sequence is any one of SEQ ID NOS:81-102. 420. The gRNA of embodiment 418 or embodiment 419, wherein the sgRNA scaffold comprises one or more G:C couples not present in the reference scaffold sequence. 421. The gRNA of any one of embodiments 418 to 419, wherein the sgRNA scaffold comprises one or more U to A substitutions relative to the reference scaffold sequence. 422. The gRNA of any one of embodiments 418 to 421, wherein the sgRNA scaffold comprises one or more trimmed stem loop sequences in place of one or more longer stem loop sequences in the reference scaffold sequence. 423. The gRNA of embodiment 422, wherein the trimmed stem loop sequence comprises a GAAA tetraloop in place of a longer stem loop sequence in the reference scaffold sequence. 424. The gRNA of any one of embodiments 418 to 423, wherein the sgRNA scaffold comprises one or more trimmed loop sequences in place of one or more longer loop sequences in the reference scaffold sequence. 425. The gRNA of embodiment 424, wherein the sgRNA scaffold comprises a GAAA tetraloop in place of a longer loop sequence in the reference scaffold sequence. 426. The gRNA of any one of embodiments 418 to 425, wherein the sgRNA scaffold comprises a nucleotide sequence that is at least 55% identical to the reference scaffold sequence. 427. The gRNA of embodiment 426, wherein the sgRNA scaffold comprises a nucleotide sequence that is at least 60% identical to the reference scaffold sequence. 428. The gRNA of embodiment 426, wherein the sgRNA scaffold comprises a nucleotide sequence that is at least 65% identical to the reference scaffold sequence. 429. The gRNA of embodiment 426, wherein the sgRNA scaffold comprises a nucleotide sequence that is at least 70% identical to the reference scaffold sequence. 430. The gRNA of embodiment 426, wherein the sgRNA scaffold comprises a nucleotide sequence that is at least 75% identical to the reference scaffold sequence. 431. The gRNA of embodiment 426, wherein the sgRNA scaffold comprises a nucleotide sequence that is at least 80% identical to the reference scaffold sequence. 432. The gRNA of embodiment 426, wherein the sgRNA scaffold comprises a nucleotide sequence that is at least 85% identical to the reference scaffold sequence. 433. The gRNA of embodiment 426, wherein the sgRNA scaffold comprises a nucleotide sequence that is at least 90% identical to the reference scaffold sequence. 434. The gRNA of embodiment 426, wherein the sgRNA scaffold comprises a nucleotide sequence that is at least 95% identical to the reference scaffold sequence. 435. The gRNA of embodiment 426, wherein the sgRNA scaffold comprises a nucleotide sequence that is at least 96% identical to the reference scaffold sequence. 436. The gRNA of embodiment 426, wherein the sgRNA scaffold comprises a nucleotide sequence that is at least 97% identical to the reference scaffold sequence. 437. The gRNA of embodiment 426, wherein the sgRNA scaffold comprises a nucleotide sequence that is at least 98% identical to the reference scaffold sequence. 438. The gRNA of embodiment 426, wherein the sgRNA scaffold comprises a nucleotide sequence that is at least 99% identical to the reference scaffold sequence. 439. The gRNA of embodiment 426, wherein the sgRNA scaffold comprises a nucleotide sequence that has no more than 5 nucleotide mismatches with the reference scaffold sequence. 440. The gRNA of embodiment 426, wherein the sgRNA scaffold comprises a nucleotide sequence that has no more than 4 nucleotide mismatches with the reference scaffold sequence. 441. The gRNA of embodiment 426, wherein the sgRNA scaffold comprises a nucleotide sequence that has no more than 3 nucleotide mismatches with the reference scaffold sequence. 442. The gRNA of embodiment 426, wherein the sgRNA scaffold comprises a nucleotide sequence that has no more than 2 nucleotide mismatches with the reference scaffold sequence. 443. The gRNA of embodiment 426, wherein the sgRNA scaffold comprises a nucleotide sequence that has no more than 1 nucleotide mismatches with the reference scaffold sequence. 444. The gRNA of embodiment 418 or embodiment 419, wherein the sgRNA scaffold comprises a nucleotide sequence that is 100% identical to the reference scaffold sequence. 445. The gRNA of any one of embodiments 418 to 444, wherein the reference scaffold sequence is SEQ ID NO:81, SEQ ID NO: 82 or SEQ ID NO:83. 446. The gRNA of embodiment 445, wherein the reference scaffold sequence is SEQ ID NO:81. 447. The gRNA of embodiment 445, wherein the reference scaffold sequence is SEQ ID NO:82. 448. The gRNA of embodiment 445, wherein the reference scaffold sequence is SEQ ID NO:83. 449. The gRNA of any one of embodiments 418 to 444, wherein the reference scaffold sequence is SEQ ID NO:84 or SEQ ID NO:85. 450. The gRNA of embodiment 449, wherein the reference scaffold sequence is SEQ ID NO:84. 451. The gRNA of embodiment 449, wherein the reference scaffold sequence is SEQ ID NO:85. 452. The gRNA of any one of embodiments 418 to 444, wherein the reference scaffold sequence is SEQ ID NO:86, SEQ ID NO:87, or SEQ ID NO:88. 453. The gRNA of embodiment 452, wherein the reference scaffold sequence is SEQ ID NO:86. 454. The gRNA of embodiment 452, wherein the reference scaffold sequence is SEQ ID NO:87. 455. The gRNA of embodiment 452, wherein the reference scaffold sequence is SEQ ID NO:88. 456. The gRNA of any one of embodiments 418 to 444, wherein the reference scaffold sequence is SEQ ID NO:89 or SEQ ID NO:90. 457. The gRNA of embodiment 456, wherein the reference scaffold sequence is SEQ ID NO:89. 458. The gRNA of embodiment 456, wherein the reference scaffold sequence is SEQ ID NO:90. 459. The gRNA of any one of embodiments 418 to 444, wherein the reference scaffold sequence is SEQ ID NO:91 or SEQ ID NO:92. 460. The gRNA of embodiment 459, wherein the reference scaffold sequence is SEQ ID NO:91. 461. The gRNA of embodiment 459, wherein the reference scaffold sequence is SEQ ID NO:92. 462. The gRNA of any one of embodiments 418 to 444, wherein the reference scaffold sequence is SEQ ID NO:93 or SEQ ID NO:94. 463. The gRNA of embodiment 462, wherein the reference scaffold sequence is SEQ ID NO:93. 464. The gRNA of embodiment 462, wherein the reference scaffold sequence is SEQ ID NO:94. 465. The gRNA of any one of embodiments 418 to 444, wherein the reference scaffold sequence is SEQ ID NO:95 or SEQ ID NO:96. 466. The gRNA of embodiment 465, wherein the reference scaffold sequence is SEQ ID NO:95. 467. The gRNA of embodiment 465, wherein the reference scaffold sequence is SEQ ID NO:96. 468. The gRNA of any one of embodiments 418 to 444, wherein the reference scaffold sequence is SEQ ID NO:97 or SEQ ID NO:98. 469. The gRNA of embodiment 471, wherein the reference scaffold sequence is SEQ ID NO:97. 470. The gRNA of embodiment 471, wherein the reference scaffold sequence is SEQ ID NO:98. 471. The gRNA of any one of embodiments 418 to 444, wherein the reference scaffold sequence is SEQ ID NO:99 or SEQ ID NO:100. 472. The gRNA of embodiment 471, wherein the reference scaffold sequence is SEQ ID NO:99. 473. The gRNA of embodiment 471, wherein the reference scaffold sequence is SEQ ID NO:100. 474. The gRNA of any one of embodiments 418 to 444, wherein the reference scaffold sequence is SEQ ID NO:101 or SEQ ID NO:102. 475. The gRNA of embodiment 474, wherein the reference scaffold sequence is SEQ ID NO:101. 476. The gRNA of embodiment 474, wherein the reference scaffold sequence is SEQ ID NO:102. 477. The gRNA of embodiment 418 or embodiment 419, wherein the nucleotide sequence of the sgRNA scaffold comprises the nucleotide sequence of SEQ ID NO:81. 478. The gRNA of embodiment 418 or embodiment 419, wherein the nucleotide sequence of the sgRNA scaffold comprises the nucleotide sequence of SEQ ID NO:82. 479. The gRNA of embodiment 418 or embodiment 419, wherein the nucleotide sequence of the sgRNA scaffold comprises the nucleotide sequence of SEQ ID NO:83. 480. The gRNA of embodiment 418 or embodiment 419, wherein the nucleotide sequence of the sgRNA scaffold comprises the nucleotide sequence of SEQ ID NO:84. 481. The gRNA of embodiment 418 or embodiment 419, wherein the nucleotide sequence of the sgRNA scaffold comprises the nucleotide sequence of SEQ ID NO:85. 482. The gRNA of embodiment 418 or embodiment 419, wherein the nucleotide sequence of the sgRNA scaffold comprises the nucleotide sequence of SEQ ID NO:86. 483. The gRNA of embodiment 418 or embodiment 419, wherein the nucleotide sequence of the sgRNA scaffold comprises the nucleotide sequence of SEQ ID NO:87. 484. The gRNA of embodiment 418 or embodiment 419, wherein the nucleotide sequence of the sgRNA scaffold comprises the nucleotide sequence of SEQ ID NO:88. 485. The gRNA of embodiment 418 or embodiment 419, wherein the nucleotide sequence of the sgRNA scaffold comprises the nucleotide sequence of SEQ ID NO:89. 486. The gRNA of embodiment 418 or embodiment 419, wherein the nucleotide sequence of the sgRNA scaffold comprises the nucleotide sequence of SEQ ID NO:90. 487. The gRNA of embodiment 418 or embodiment 419, wherein the nucleotide sequence of the sgRNA scaffold comprises the nucleotide sequence of SEQ ID NO:91. 488. The gRNA of embodiment 418 or embodiment 419, wherein the nucleotide sequence of the sgRNA scaffold comprises the nucleotide sequence of SEQ ID NO:92. 489. The gRNA of embodiment 418 or embodiment 419, wherein the nucleotide sequence of the sgRNA scaffold comprises the nucleotide sequence of SEQ ID NO:93. 490. The gRNA of embodiment 418 or embodiment 419, wherein the nucleotide sequence of the sgRNA scaffold comprises the nucleotide sequence of SEQ ID NO:94. 491. The gRNA of embodiment 418 or embodiment 419, wherein the nucleotide sequence of the sgRNA scaffold comprises the nucleotide sequence of SEQ ID NO:95. 492. The gRNA of embodiment 418 or embodiment 419, wherein the nucleotide sequence of the sgRNA scaffold comprises the nucleotide sequence of SEQ ID NO:96. 493. The gRNA of embodiment 418 or embodiment 419, wherein the nucleotide sequence of the sgRNA scaffold comprises the nucleotide sequence of SEQ ID NO:97. 494. The gRNA of embodiment 418 or embodiment 419, wherein the nucleotide sequence of the sgRNA scaffold comprises the nucleotide sequence of SEQ ID NO:98. 495. The gRNA of embodiment 418 or embodiment 419, wherein the nucleotide sequence of the sgRNA scaffold comprises the nucleotide sequence of SEQ ID NO:99. 496. The gRNA of embodiment 418 or embodiment 419, wherein the nucleotide sequence of the sgRNA scaffold comprises the nucleotide sequence of SEQ ID NO:100. 497. The gRNA of embodiment 418 or embodiment 419, wherein the nucleotide sequence of the sgRNA scaffold comprises the nucleotide sequence of SEQ ID NO:101. 498. The gRNA of embodiment 418 or embodiment 419, wherein the nucleotide sequence of the sgRNA scaffold comprises the nucleotide sequence of SEQ ID NO:102. 499. The gRNA of any one of embodiments 418 to 498, wherein the sgRNA scaffold FRPSULVHV^^^WR^^^XUDFLOV^DW^LWV^^¶^HQG^ 500. The gRNA of embodiment 499, wheUHLQ^WKH^VJ51$^VFDIIROG^FRPSULVHV^^^XUDFLO^DW^LWV^^¶^ end. 501. The gRNA of embodiment 499^^ZKHUHLQ^WKH^VJ51$^VFDIIROG^FRPSULVHV^^^XUDFLOV^DW^LWV^^¶^ end. 502. The gRNA of embodiment 499^^ZKHUHLQ^WKH^VJ51$^VFDIIROG^FRPSULVHV^^^XUDFLOV^DW^LWV^^¶^ end. 503. The gRNA of embodiment 499^^ZKHUHLQ^WKH^VJ51$^VFDIIROG^FRPSULVHV^^^XUDFLOV^DW^LWV^^¶^ end. 504. The gRNA of embodiment 499^^ZKHUHLQ^WKH^VJ51$^VFDIIROG^FRPSULVHV^^^XUDFLOV^DW^LWV^^¶^ end. 505. The gRNA of embodiment 499^^ZKHUHLQ^WKH^VJ51$^VFDIIROG^FRPSULVHV^^^XUDFLOV^DW^LWV^^¶^ end. 506. The gRNA of embodiment 499^^ZKHUHLQ^WKH^VJ51$^VFDIIROG^FRPSULVHV^^^XUDFLOV^DW^LWV^^¶^ end. 507. The gRNA of embodiment 499^^ZKHUHLQ^WKH^VJ51$^VFDIIROG^FRPSULVHV^^^XUDFLOV^DW^LWV^^¶^ end. 508. The gRNA of embodiment 418 or embodiment 419, wherein the nucleotide sequence of the sgRNA scaffold comprises the nucleotide sequence of any one of SEQ ID NOS:103-115. 509. The gRNA of embodiment 508, wherein the nucleotide sequence of the sgRNA scaffold comprises the nucleotide sequence of SEQ ID NO:103. 510. The gRNA of embodiment 508, wherein the nucleotide sequence of the sgRNA scaffold comprises the nucleotide sequence of SEQ ID NO:104. 511. The gRNA of embodiment 508, wherein the nucleotide sequence of the sgRNA scaffold comprises the nucleotide sequence of SEQ ID NO:105. 512. The gRNA of embodiment 508, wherein the nucleotide sequence of the sgRNA scaffold comprises the nucleotide sequence of SEQ ID NO:106. 513. The gRNA of embodiment 508, wherein the nucleotide sequence of the sgRNA scaffold comprises the nucleotide sequence of SEQ ID NO:107. 514. The gRNA of embodiment 508, wherein the nucleotide sequence of the sgRNA scaffold comprises the nucleotide sequence of SEQ ID NO:108. 515. The gRNA of embodiment 508, wherein the nucleotide sequence of the sgRNA scaffold comprises the nucleotide sequence of SEQ ID NO:109. 516. The gRNA of embodiment 508, wherein the nucleotide sequence of the sgRNA scaffold comprises the nucleotide sequence of SEQ ID NO:110. 517. The gRNA of embodiment 508, wherein the nucleotide sequence of the sgRNA scaffold comprises the nucleotide sequence of SEQ ID NO:111. 518. The gRNA of embodiment 508, wherein the nucleotide sequence of the sgRNA scaffold comprises the nucleotide sequence of SEQ ID NO:112. 519. The gRNA of embodiment 508, wherein the nucleotide sequence of the sgRNA scaffold comprises the nucleotide sequence of SEQ ID NO:113. 520. The gRNA of embodiment 508, wherein the nucleotide sequence of the sgRNA scaffold comprises the nucleotide sequence of SEQ ID NO:114. 521. The gRNA of embodiment 508, wherein the nucleotide sequence of the sgRNA scaffold comprises the nucleotide sequence of SEQ ID NO:115. 522. The gRNA of any one of embodiments 418 to 521, wherein the nucleotide sequence of the spacer is partially or fully complementary to a target mammalian genomic sequence. 523. A gRNA comprising (i) a crRNA comprising a spacer (optionally wherein the spacer is a spacer described in any one of embodiments 303 to 416) DQG^D^FU51$^VFDIIROG^^ZKHUHLQ^WKH^VSDFHU^LV^^¶^ to the crRNA scaffold, and (ii) a tracrRNA, wherein the nucleotide sequence of the spacer is partially or fully complementary to a target mammalian genomic sequence and the nucleotide sequence of the crRNA scaffold comprises the nucleotide sequence of SEQ ID NO:61, SEQ ID NO:63, SEQ ID NO:65, SEQ ID NO:67, SEQ ID NO:69, SEQ ID NO:71, SEQ ID NO:73, SEQ ID NO:75, SEQ ID NO:77, or SEQ ID NO:79. 524. A gRNA comprising (i) a crRNA comprising a means for binding a target mammalian genomic sequence (which is optionally a spacer) and a crRNA scaffold, wherein the means for binding a target mammalian JHQRPLF^VHTXHQFH^LV^^¶^WR^WKH^FU51$^VFDIIROG^^DQG^^LL^^D^WUDFU51$^^ZKHUHLQ^WKH^ nucleotide sequence of the crRNA scaffold comprises the nucleotide sequence of SEQ ID NO:61, SEQ ID NO:63, SEQ ID NO:65, SEQ ID NO:67, SEQ ID NO:69, SEQ ID NO:71, SEQ ID NO:73, SEQ ID NO:75, SEQ ID NO:77 , or SEQ ID NO:79. 525. The gRNA of embodiment 523 or 524, wherein the nucleotide sequence of the crRNA scaffold comprises the nucleotide sequence of SEQ ID NO:61. 526. The gRNA of any one of embodiments 523 to 525, wherein the nucleotide sequence of the tracrRNA comprises the nucleotide sequence of SEQ ID NO:62. 527. The gRNA of embodiment 523 or 524, wherein the nucleotide sequence of the crRNA scaffold comprises the nucleotide sequence of SEQ ID NO:63. 528. The gRNA of embodiment 523, embodiment 524, or embodiment 527, wherein the nucleotide sequence of the tracrRNA comprises the nucleotide sequence of SEQ ID NO:64. 529. The gRNA of embodiment 523 or 524, wherein the nucleotide sequence of the crRNA scaffold comprises the nucleotide sequence of SEQ ID NO:65. 530. The gRNA of embodiment 523, embodiment 524, or embodiment 529, wherein the nucleotide sequence of the tracrRNA comprises the nucleotide sequence of SEQ ID NO:66. 531. The gRNA of embodiment 523 or 524, wherein the nucleotide sequence of the crRNA scaffold comprises the nucleotide sequence of SEQ ID NO:67. 532. The gRNA of embodiment 523, embodiment 524, or embodiment 531, wherein the nucleotide sequence of the tracrRNA comprises the nucleotide sequence of SEQ ID NO:68. 533. The gRNA of embodiment 523 or 524, wherein the nucleotide sequence of the crRNA scaffold comprises the nucleotide sequence of SEQ ID NO:69. 534. The gRNA of embodiment 523, embodiment 524, or embodiment 533, wherein the nucleotide sequence of the tracrRNA comprises the nucleotide sequence of SEQ ID NO:70. 535. The gRNA of embodiment 523 or 524, wherein the nucleotide sequence of the crRNA scaffold comprises the nucleotide sequence of SEQ ID NO:71. 536. The gRNA of embodiment 523, embodiment 524, or embodiment 535, wherein the nucleotide sequence of the tracrRNA comprises the nucleotide sequence of SEQ ID NO:72. 537. The gRNA of embodiment 523 or 524, wherein the nucleotide sequence of the crRNA scaffold comprises the nucleotide sequence of SEQ ID NO:73. 538. The gRNA of embodiment 523, embodiment 524, or embodiment 537, wherein the nucleotide sequence of the tracrRNA comprises the nucleotide sequence of SEQ ID NO:74. 539. The gRNA of embodiment 523 or 524, wherein the nucleotide sequence of the crRNA scaffold comprises the nucleotide sequence of SEQ ID NO:75. 540. The gRNA of embodiment 523, embodiment 524, or embodiment 539, wherein the nucleotide sequence of the tracrRNA comprises the nucleotide sequence of SEQ ID NO:76. 541. The gRNA of embodiment 523 or 524, wherein the nucleotide sequence of the crRNA scaffold comprises the nucleotide sequence of SEQ ID NO:77. 542. The gRNA of embodiment 523, embodiment 524, or embodiment 541, wherein the nucleotide sequence of the tracrRNA comprises the nucleotide sequence of SEQ ID NO:78. 543. The gRNA of embodiment 523 or 524, wherein the nucleotide sequence of the crRNA scaffold comprises the nucleotide sequence of SEQ ID NO:79. 544. The gRNA of embodiment 523, embodiment 524, or embodiment 543, wherein the nucleotide sequence of the tracrRNA comprises the nucleotide sequence of SEQ ID NO:80. 545. The gRNA of any one of embodiments 523 to 544, wherein the gRNA comprises separate crRNA and tracrRNA molecules. 546. The gRNA of any one of embodiments 523 to 544, wherein the gRNA is a single guide RNA (sgRNA). 547. The gRNA of any one of embodiments 522 to 546, wherein the target mammalian genomic sequence is a human genomic sequence. 548. The gRNA of embodiment 547, wherein the target mammalian genomic sequence is a CCR5, EMX1, Fas, FANCF, HBB, ZSCAN2, Chr6, ADAMTSL1, B2M, CXCR4, PD1, DNMT1, Match8, TRAC, TRBC, VEGFAsite2, VEGFAsite3, CACNA, HEKsite3, HEKsite4, Chr8, BCR, ATM, HBG1, HPRT, IL2RG, NF1, USH2A, RHO, BcLenh, or CTFR genomic sequence. 549. The gRNA of embodiment 547, wherein the target mammalian genomic sequence is a RHO genomic sequence. 550. The gRNA of embodiment 547, wherein the target mammalian genomic sequence is a TRAC genomic sequence. 551. The gRNA of embodiment 547, wherein the target mammalian genomic sequence is a B2M genomic sequence. 552. The gRNA of embodiment 547, wherein the target mammalian genomic sequence is a PD1 genomic sequence. 553. The gRNA of embodiment 548, wherein the target mammalian genomic sequence is a CCR5, EMX1, Fas, FANCF, HBB, ZSCAN, Chr6, ADAMTSL1, B2M, CXCR4, PD1, DNMT1, Match8, TRAC, TRBC, VEGFAsite2, VEGFAsite3, CACNA, HEKsite3, HEKsite4, Chr8, or BCR genomic sequence. 554. The gRNA of any one of embodiments 418 to 553, wherein the target mammalian genomic sequence is upstream of a protospacer adjacent motif (PAM) sequence in the non-target strand recognized by a Type II Cas protein, optionally wherein the Type II Cas protein is a Type II Cas protein according to any one of embodiments 1 to 286. 555. The gRNA of embodiment 554, wherein the PAM sequence is NGG. 556. The gRNA of embodiment 554, wherein the PAM sequence is NNGNC. 557. The gRNA of embodiment 554, wherein the PAM sequence is NNGNCNY. 558. The gRNA of embodiment 554, wherein the PAM sequence is NNRC. 559. The gRNA of embodiment 554, wherein the PAM sequence is^NNAC. 560. The gRNA of embodiment 554, wherein the PAM sequence is NNACNT. 561. The gRNA of embodiment 554, wherein the PAM sequence is NNRTA. 562. The gRNA of embodiment 554, wherein the PAM sequence is NNNNCC. 563. The gRNA of embodiment 554, wherein the PAM sequence is NNANCC. 564. The gRNA of embodiment 554, wherein the PAM sequence is NNACCC. 565. The gRNA of embodiment 554, wherein the PAM sequence is NNAT. 566. The gRNA of embodiment 554, wherein the PAM sequence is NNATM. 567. The gRNA of embodiment 554, wherein the PAM sequence is NNATA. 568. The gRNA of embodiment 554, wherein the PAM sequence is NVAT. 569. The gRNA of embodiment 554, wherein the PAM sequence is NVATM. 570. The gRNA of embodiment 554, wherein the PAM sequence is NVRTA. 571. The gRNA of embodiment 554, wherein the PAM sequence is NRRTA. 572. The gRNA of embodiment 554, wherein the PAM sequence is NVATA. 573. The gRNA of embodiment 554, wherein the PAM sequence is NRATA. 574. The gRNA of embodiment 554, wherein the PAM sequence is NNNAC. 575. The gRNA of embodiment 554, wherein the PAM sequence is NRNAC. 576. The gRNA of embodiment 554, wherein the PAM sequence is^NRHAC. 577. The gRNA of embodiment 554, wherein the PAM sequence is NRNACD. 578. The gRNA of embodiment 554, wherein the PAM sequence is NRHACD. 579. The gRNA of embodiment 554, wherein the PAM sequence is NRNACA. 580. The gRNA of embodiment 554, wherein the PAM sequence is NRHACA. 581. The gRNA of embodiment 554, wherein the PAM sequence is NNNRNY. 582. The gRNA of embodiment 554, wherein the PAM sequence is NRNRNY. 583. The gRNA of embodiment 554, wherein the PAM sequence is NRNRNC. 584. The gRNA of embodiment 554, wherein the PAM sequence is NANANC. 585. The gRNA of embodiment 554, wherein the PAM sequence is NNNRDC. 586. The gRNA of embodiment 554, wherein the PAM sequence is NNART. 587. The gRNA of embodiment 554, wherein the PAM sequence is NRART. 588. The gRNA of any one of embodiments 418 to 587, wherein the spacer is 15 to 30 nucleotides in length. 589. The gRNA of embodiment 588, wherein the spacer is 15 to 25 nucleotides in length. 590. The gRNA of embodiment 588, wherein the spacer is 16 to 24 nucleotides in length. 591. The gRNA of embodiment 588, wherein the spacer is 17 to 23 nucleotides in length. 592. The gRNA of embodiment 588, wherein the spacer is 18 to 22 nucleotides in length. 593. The gRNA of embodiment 588, wherein the spacer is 19 to 21 nucleotides in length. 594. The gRNA of embodiment 588, wherein the spacer is 18 to 30 nucleotides in length. 595. The gRNA of embodiment 588, wherein the spacer is 20 to 28 nucleotides in length. 596. The gRNA of embodiment 588, wherein the spacer is 22 to 26 nucleotides in length. 597. The gRNA of embodiment 588, wherein the spacer is 23 to 25 nucleotides in length. 598. The gRNA of embodiment 588, wherein the spacer is 18 nucleotides in length. 599. The gRNA of embodiment 588, wherein the spacer is 19 nucleotides in length. 600. The gRNA of embodiment 588, wherein the spacer is 20 nucleotides in length. 601. The gRNA of embodiment 588, wherein the spacer is 21 nucleotides in length. 602. The gRNA of embodiment 588, wherein the spacer is 22 nucleotides in length. 603. The gRNA of embodiment 588, wherein the spacer is 23 nucleotides in length. 604. The gRNA of embodiment 588, wherein the spacer is 24 nucleotides in length. 605. The gRNA of embodiment 588, wherein the spacer is 25 nucleotides in length. 606. The gRNA of embodiment 588, wherein the spacer is 26 nucleotides in length. 607. The gRNA of embodiment 588, wherein the spacer is 27 nucleotides in length. 608. The gRNA of embodiment 588, wherein the spacer is 28 nucleotides in length. 609. A system comprising the Type II Cas protein of any one of embodiments 1 to 287 and a guide RNA (gRNA) comprising a spacer sequence, optionally wherein the gRNA is a gRNA according to any one of embodiments 288 to 608. 610. A system comprising the Type II Cas protein of any one of embodiments 1 to 287 and a means for targeting the Type II Cas protein to a target genomic sequence, optionally wherein the means for targeting the Type II Cas protein to a target genomic sequence is a guide RNA (gRNA) molecule, optionally as described in in any one of embodiments 288 to 608, optionally wherein the gRNA molecule comprises a spacer partially or fully complementary to a target mammalian genomic sequence. 611. The system of embodiment 609, wherein the spacer sequence is partially or fully complementary to a target mammalian genomic sequence. 612. The system of any one of embodiments 610 to 611, wherein the target mammalian genomic sequence is a human genomic sequence. 613. The system of embodiment 612, wherein the target mammalian genomic sequence is a CCR5, EMX1, Fas, FANCF, HBB, ZSCAN2, Chr6, ADAMTSL1, B2M, CXCR4, PD1, DNMT1, Match8, TRAC, TRBC, VEGFAsite2, VEGFAsite3, CACNA, HEKsite3, HEKsite4, Chr8, BCR, ATM, HBG1, HPRT, IL2RG, NF1, USH2A, RHO, BcLenh, or CTFR genomic sequence. 614. The system of any one of embodiments 610 to 613, wherein the target mammalian genomic sequence is upstream of a protospacer adjacent motif (PAM) sequence in the non-target strand recognized by the Type II Cas protein. 615. The system of embodiment 614, wherein the reference protein sequence is SEQ ID NO:1 or SEQ ID NO:2 and wherein the PAM sequence is NGG. 616. The system of embodiment 614, wherein the reference protein sequence is SEQ ID NO:7 or SEQ ID NO:8 and the PAM sequence is NNGNC. 617. The system of embodiment 614, wherein the reference protein sequence is SEQ ID NO:7 or SEQ ID NO:8 and the PAM sequence is NNGNCNY. 618. The system of embodiment 614, wherein the reference protein sequence is SEQ ID NO:13 or SEQ ID NO:14 and the PAM sequence is NNRC. 619. The system of embodiment 614, wherein the reference protein sequence is SEQ ID NO:13 or SEQ ID NO:14 and the PAM sequence is NNAC. 620. The system of embodiment 614, wherein the reference protein sequence is SEQ ID NO:13 or SEQ ID NO:14 and the PAM sequence is NNACNT. 621. The system of embodiment 614, wherein the reference protein sequence is SEQ ID NO:19 or SEQ ID NO:20 and the PAM sequence is NNRTA. 622. The system of embodiment 614, wherein the reference protein sequence is SEQ ID NO:25 or SEQ ID NO:26 and the PAM sequence is NNNNCC. 623. The system of embodiment 614, wherein the reference protein sequence is SEQ ID NO:25 or SEQ ID NO:26 and the PAM sequence is NNANCC. 624. The system of embodiment 614, wherein the reference protein sequence is SEQ ID NO:25 or SEQ ID NO:26 and the PAM sequence is NNACCC. 625. The system of embodiment 614, wherein the reference protein sequence is SEQ ID NO:31 or SEQ ID NO:32 and the PAM sequence is NGG. 626. The system of embodiment 614, wherein the reference protein sequence is SEQ ID NO:37 or SEQ ID NO:38 and the PAM sequence is NNAT, NNATM, NNATA, NVAT, or NVATM. 627. The system of embodiment 614, wherein the reference protein sequence is SEQ ID NO:37 or SEQ ID NO:38 and the PAM sequence is NVRTA, NRRTA, NVATA, or NRATA. 628. The system of embodiment 626, wherein the PAM sequence is NNAT. 629. The system of embodiment 626, wherein the PAM sequence is NNATM. 630. The system of embodiment 626, wherein the PAM sequence is NNATA. 631. The system of embodiment 626, wherein the PAM sequence is NVAT. 632. The system of embodiment 626, wherein the PAM sequence is NVATM. 633. The system of embodiment 627, wherein the PAM sequence is NVRTA. 634. The system of embodiment 627, wherein the PAM sequence is NRRTA. 635. The system of embodiment 627, wherein the PAM sequence is NVATA. 636. The system of embodiment 627, wherein the PAM sequence is NRATA. 637. The system of embodiment 614, wherein the reference protein sequence is SEQ ID NO:43 or SEQ ID NO:44 and the PAM sequence is NNNAC, NRNAC, NRHAC, NRNACD, NRHACD, NRNACA, or NRHACA. 638. The system of embodiment 637, wherein the PAM sequence is NNNAC. 639. The system of embodiment 637, wherein the PAM sequence is NRNAC. 640. The system of embodiment 637, wherein the PAM sequence is NRHAC. 641. The system of embodiment 633, wherein the PAM sequence is NRNACD. 642. The system of embodiment 633, wherein the PAM sequence is NRHACD. 643. The system of embodiment 637, wherein the PAM sequence is NRNACA. 644. The system of embodiment 637, wherein the PAM sequence is NRHACA. 645. The system of embodiment 614, wherein the reference protein sequence is SEQ ID NO:49 or SEQ ID NO:50 and the PAM sequence is NNNRNY, NRNRNY,NRNRNC, NANANC, or NNNRDC. 646. The system of embodiment 645, wherein the PAM sequence is NNNRNY. 647. The system of embodiment 645, wherein the PAM sequence is NRNRNY. 648. The system of embodiment 645, wherein the PAM sequence is NRNRNC. 649. The system of embodiment 645, wherein the PAM sequence is NANANC. 650. The system of embodiment 645, wherein the PAM sequence is NNNRDC. 651. The system of embodiment 614, wherein the reference protein sequence is SEQ ID NO:55 or SEQ ID NO:56 and the PAM sequence is NNART or NNART. 652. The system of embodiment 651, wherein the PAM sequence is NNART. 653. The system of embodiment 651, wherein the PAM sequence is NRART. 654. The system of any one of embodiments 609 to 653, wherein the gRNA comprises a crRNA sequence and a tracrRNA sequence. 655. The system of embodiment 654, wherein the reference protein sequence is SEQ ID NO:1 or SEQ ID NO:2 and wherein the crRNA sequence compriseV^WKH^VSDFHU^VHTXHQFH^^¶^WR^WKH^ nucleotide sequence of SEQ ID NO:61. 656. The system of embodiment 654 or embodiment 655, wherein the reference protein sequence is SEQ ID NO:1 or SEQ ID NO:2 and wherein the tracrRNA sequence comprises the nucleotide sequence of SEQ ID NO:62. 657. The system of embodiment 654, wherein the reference protein sequence is SEQ ID 12^^^RU^6(4^,'^12^^^DQG^ZKHUHLQ^WKH^FU51$^VHTXHQFH^FRPSULVHV^WKH^VSDFHU^VHTXHQFH^^¶^WR^WKH^ nucleotide sequence of SEQ ID NO:63. 658. The system of embodiment 654 or embodiment 657, wherein the reference protein sequence is SEQ ID NO:7 or SEQ ID NO:8 and wherein the tracrRNA sequence comprises the nucleotide sequence of SEQ ID NO:64. 659. The system of embodiment 654, wherein the reference protein sequence is SEQ ID NO:13 or SEQ ID NO:14 DQG^ZKHUHLQ^WKH^FU51$^VHTXHQFH^FRPSULVHV^WKH^VSDFHU^VHTXHQFH^^¶^WR^WKH^ nucleotide sequence of SEQ ID NO:65. 660. The system of embodiment 654 or embodiment 659, wherein the reference protein sequence is SEQ ID NO:13 or SEQ ID NO:14 and wherein the tracrRNA sequence comprises the nucleotide sequence of SEQ ID NO:66. 661. The system of embodiment 654, wherein the reference protein sequence is SEQ ID NO:19 or SEQ ID NO:20 DQG^ZKHUHLQ^WKH^FU51$^VHTXHQFH^FRPSULVHV^WKH^VSDFHU^VHTXHQFH^^¶^WR^WKH^ nucleotide sequence of SEQ ID NO:67. 662. The system of embodiment 654 or embodiment 661, wherein the reference protein sequence is SEQ ID NO:19 or SEQ ID NO:20 and wherein the tracrRNA sequence comprises the nucleotide sequence of SEQ ID NO:68. 663. The system of embodiment 654, wherein the reference protein sequence is SEQ ID 12^^^^RU^6(4^,'^12^^^^DQG^ZKHUHLQ^WKH^FU51$^VHTXHQFH^FRPSULVHV^WKH^VSDFHU^VHTXHQFH^^¶^WR^WKH^ nucleotide sequence of SEQ ID NO:69. 664. The system of embodiment 654 or embodiment 663, wherein the reference protein sequence is SEQ ID NO:25 or SEQ ID NO:26 and wherein the tracrRNA sequence comprises the nucleotide sequence of SEQ ID NO:70. 665. The system of embodiment 654, wherein the reference protein sequence is SEQ ID NO:31 or SEQ ID NO:32 and wherein the crRNA sequenFH^FRPSULVHV^WKH^VSDFHU^VHTXHQFH^^¶^WR^WKH^ nucleotide sequence of SEQ ID NO:71. 666. The system of embodiment 654 or embodiment 665, wherein the reference protein sequence is SEQ ID NO:31 or SEQ ID NO:32 and wherein the tracrRNA sequence comprises the nucleotide sequence of SEQ ID NO:72. 667. The system of embodiment 654, wherein the reference protein sequence is SEQ ID 12^^^^RU^6(4^,'^12^^^^DQG^ZKHUHLQ^WKH^FU51$^VHTXHQFH^FRPSULVHV^WKH^VSDFHU^VHTXHQFH^^¶^WR^WKH^ nucleotide sequence of SEQ ID NO:73. 668. The system of embodiment 654 or embodiment 667, wherein the reference protein sequence is SEQ ID NO:37 or SEQ ID NO:38 and wherein the tracrRNA sequence comprises the nucleotide sequence of SEQ ID NO:74. 669. The system of embodiment 654, wherein the reference protein sequence is SEQ ID 12^^^^RU^6(4^,'^12^^^^DQG^ZKHUHLQ^WKH^FU51$^VHTXHQFH^FRPSULVHV^WKH^VSDFHU^VHTXHQFH^^¶^WR^WKH^ nucleotide sequence of SEQ ID NO:75. 670. The system of embodiment 654 or embodiment 669, wherein the reference protein sequence is SEQ ID NO:43 or SEQ ID NO:44 and wherein the tracrRNA sequence comprises the nucleotide sequence of SEQ ID NO:76. 671. The system of embodiment 654, wherein the reference protein sequence is SEQ ID NO:49 or SEQ ID NO:50 and wherein the crRNA sequence comprises the spacer sequence ^¶^WR^WKH^ nucleotide sequence of SEQ ID NO:77. 672. The system of embodiment 654 or embodiment 671, wherein the reference protein sequence is SEQ ID NO:49 or SEQ ID NO:50 and wherein the tracrRNA sequence comprises the nucleotide sequence of SEQ ID NO:78. 673. The system of embodiment 654, wherein the reference protein sequence is SEQ ID 12^^^^RU^6(4^,'^12^^^^DQG^ZKHUHLQ^WKH^FU51$^VHTXHQFH^FRPSULVHV^WKH^VSDFHU^VHTXHQFH^^¶^WR^WKH^ nucleotide sequence of SEQ ID NO:79. 674. The system of embodiment 654 or embodiment 673, wherein the reference protein sequence is SEQ ID NO:55 or SEQ ID NO:56 and wherein the tracrRNA sequence comprises the nucleotide sequence of SEQ ID NO:80. 675. The system of any one of embodiments 609 to 674, wherein the gRNA comprises separate crRNA and tracrRNA molecules. 676. The system of any one of embodiments 609 to 674, wherein the gRNA is a single guide 51$^^VJ51$^^FRPSULVLQJ^WKH^VSDFHU^DQG^D^VJ51$^VFDIIROG^^ZKHUHLQ^WKH^VSDFHU^LV^SRVLWLRQHG^^¶^WR^WKH^ sgRNA scaffold. 677. The system of embodiment 676, wherein the sgRNA scaffold is a sgRNA scaffold described in any one of embodiments 418 to 608. 678. The system of embodiment 676 or embodiment 677, wherein the sgRNA scaffold comprises the nucleotide sequence of any one SEQ ID NOS:81-115. 679. The system of any one of embodiments 609 to 678, wherein the spacer is a spacer described in any one of embodiments 303 to 608. 680. The system of any one of embodiments 609 to 679, which is a ribonucleoprotein (RNP) comprising the Type II Cas protein complexed to the gRNA or means for targeting the Type II Cas protein to a target genomic sequence. 681. A nucleic acid encoding the Type II Cas protein of any one of embodiments 1 to 287, optionally wherein the nucleotide sequence encoding the Type II Cas protein is operably linked to a promoter that is heterologous to the Type II Cas protein. 682. The nucleic acid of embodiment 681, wherein the nucleotide sequence encoding the Type II Cas protein is codon optimized for expression in human cells. 683. The nucleic acid of embodiment 682, wherein when the reference protein sequence is SEQ ID NO:1 or SEQ ID NO:2, the nucleotide sequence encoding the Type II Cas protein comprises a nucleotide sequence that is at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% identical to the nucleotide sequence of SEQ ID NO:5 or SEQ ID NO:6. 684. The nucleic acid of embodiment 682, wherein when the reference protein sequence is SEQ ID NO:7 or SEQ ID NO:8, the nucleotide sequence encoding the Type II Cas protein comprises a nucleotide sequence that is at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% identical to the nucleotide sequence of SEQ ID NO:11 or SEQ ID NO:12. 685. The nucleic acid of embodiment 682, wherein when the reference protein sequence is SEQ ID NO:13 or SEQ ID NO:14, the nucleotide sequence encoding the Type II Cas protein comprises a nucleotide sequence that is at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% identical to the nucleotide sequence of SEQ ID NO:17 or SEQ ID NO:18. 686. The nucleic acid of embodiment 682, wherein when the reference protein sequence is SEQ ID NO:19 or SEQ ID NO:20, the nucleotide sequence encoding the Type II Cas protein comprises a nucleotide sequence that is at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% identical to the nucleotide sequence of SEQ ID NO:23 or SEQ ID NO:24. 687. The nucleic acid of embodiment 682, wherein when the reference protein sequence is SEQ ID NO:25 or SEQ ID NO:26, the nucleotide sequence encoding the Type II Cas protein comprises a nucleotide sequence that is at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% identical to the nucleotide sequence of SEQ ID NO:29 or SEQ ID NO:30. 688. The nucleic acid of embodiment 682, wherein when the reference protein sequence is SEQ ID NO:31 or SEQ ID NO:32, the nucleotide sequence encoding the Type II Cas protein comprises a nucleotide sequence that is at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% identical to the nucleotide sequence of SEQ ID NO:35 or SEQ ID NO:36. 689. The nucleic acid of embodiment 682, wherein when the reference protein sequence is SEQ ID NO:37 or SEQ ID NO:38, the nucleotide sequence encoding the Type II Cas protein comprises a nucleotide sequence that is at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% identical to the nucleotide sequence of SEQ ID NO:41 or SEQ ID NO:42. 690. The nucleic acid of embodiment 682, wherein when the reference protein sequence is SEQ ID NO:43 or SEQ ID NO:44, the nucleotide sequence encoding the Type II Cas protein comprises a nucleotide sequence that is at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% identical to the nucleotide sequence of SEQ ID NO:47 or SEQ ID NO:48. 691. The nucleic acid of embodiment 682, wherein when the reference protein sequence is SEQ ID NO:49 or SEQ ID NO:50, the nucleotide sequence encoding the Type II Cas protein comprises a nucleotide sequence that is at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% identical to the nucleotide sequence of SEQ ID NO:53 or SEQ ID NO:54. 692. The nucleic acid of embodiment 682, wherein when the reference protein sequence is SEQ ID NO:55 or SEQ ID NO:56, the nucleotide sequence encoding the Type II Cas protein comprises a nucleotide sequence that is at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% identical to the nucleotide sequence of SEQ ID NO:59 or SEQ ID NO:60. 693. The nucleic acid of any one of embodiments embodiment 681 to 692, which is a plasmid. 694. The nucleic acid of any one of embodiments embodiment 681 to 692, which is a viral genome. 695. The nucleic acid of embodiment 694, wherein the viral genome is an adeno-associated virus (AAV) genome. 696. The nucleic acid of embodiment 695, wherein the AAV genome is an AAV2, AAV5, AAV7m8, AAV8, AAV9, AAVrh8r, or AAVrh10 genome. 697. The nucleic acid of embodiment 696, wherein the AAV genome is an AAV2 genome. 698. The nucleic acid of embodiment 696, wherein the AAV genome is an AAV5 genome. 699. The nucleic acid of embodiment 696, wherein the AAV genome is an AAV7m8 genome. 700. The nucleic acid of embodiment 696, wherein the AAV genome is an AAV8 genome. 701. The nucleic acid of embodiment 696, wherein the AAV genome is an AAV9 genome. 702. The nucleic acid of embodiment 696, wherein the AAV genome is an AAVrh8r genome. 703. The nucleic acid of embodiment 696, wherein the AAV genome is an AAVrh10 genome. 704. The nucleic acid of any one of embodiments 681 to 703, further encoding a gRNA, optionally wherein the gRNA is a gRNA according to any one of embodiments 288 to 608. 705. A nucleic acid encoding the gRNA of any one of embodiments 288 to 608. 706. The nucleic acid of embodiment 705, which is a plasmid. 707. The nucleic acid of embodiment 705, which is a viral genome. 708. The nucleic acid of embodiment 707, wherein the viral genome is an adeno-associated virus (AAV) genome. 709. The nucleic acid of embodiment 708, wherein the AAV genome is a AAV2, AAV5, AAV7m8, AAV8, AAV9, AAVrh8r, or AAVrh10 genome. 710. The nucleic acid of embodiment 709, wherein the AAV genome is an AAV2 genome. 711. The nucleic acid of embodiment 709, wherein the AAV genome is an AAV5 genome. 712. The nucleic acid of embodiment 709, wherein the AAV genome is an AAV7m8 genome. 713. The nucleic acid of embodiment 709, wherein the AAV genome is an AAV8 genome. 714. The nucleic acid of embodiment 709, wherein the AAV genome is an AAV9 genome. 715. The nucleic acid of embodiment 709, wherein the AAV genome is an AAVrh8r genome. 716. The nucleic acid of embodiment 709, wherein the AAV genome is an AAVrh10 genome. 717. The nucleic acid of any one of embodiments 705 to 716, further encoding a Type II Cas protein, optionally wherein the Type II Cas protein is a Type II Cas protein according to any one of embodiments 1 to 287. 718. A nucleic acid encoding the Type II Cas protein and gRNA of the system of any one of embodiments 609 to 680. 719. The nucleic acid of embodiment 718, wherein the nucleotide sequence encoding the Type II Cas protein is codon optimized for expression in human cells. 720. The nucleic acid of embodiment 718 or embodiment 719, which is a plasmid. 721. The nucleic acid of embodiment 718 or embodiment 719, which is a viral genome. 722. The nucleic acid of embodiment 721, wherein the viral genome is an adeno-associated virus (AAV) genome. 723. The nucleic acid of embodiment 722, wherein the AAV genome is an AAV2, AAV5, AAV7m8, AAV8, AAV9, AAVrh8r, or AAVrh10 genome. 724. The nucleic acid of embodiment 723, wherein the AAV genome is an AAV2 genome. 725. The nucleic acid of embodiment 723, wherein the AAV genome is an AAV5 genome. 726. The nucleic acid of embodiment 723, wherein the AAV genome is an AAV7m8 genome. 727. The nucleic acid of embodiment 723, wherein the AAV genome is an AAV8 genome. 728. The nucleic acid of embodiment 723, wherein the AAV genome is an AAV9 genome. 729. The nucleic acid of embodiment 723, wherein the AAV genome is an AAVrh8r genome. 730. The nucleic acid of embodiment 723, wherein the AAV genome is an AAVrh10 genome. 731. A plurality of nucleic acids comprising separate nucleic acids encoding the Type II Cas protein and gRNA of the system of any one of embodiments 609 to 680. 732. The plurality of nucleic acid of embodiment 731, wherein the separate nucleic acids encoding the Type II Cas protein and gRNA are plasmids. 733. The plurality of nucleic acids of embodiment 731, wherein the separate nucleic acids encoding the Type II Cas protein and gRNA are viral genomes. 734. The plurality of nucleic acids of embodiment 733, wherein the viral genomes are adeno- associated virus (AAV) genomes. 735. The plurality of nucleic acids of embodiment 734, wherein the AAV genomes the encoding the Type II Cas protein and gRNA are independently an AAV2, AAV5, AAV7m8, AAV8, AAV9, AAVrh8r, or AAVrh10 genome. 736. A Type II Cas protein according to any one of embodiments 1 to 287, a gRNA according to any one of embodiments 288 to 608, a system according to of any one of embodiments 609 to 680, a nucleic acid according to any one of embodiments 681 to 730, or a plurality of nucleic acids according to of any one of embodiments 731 to 735 for use in a method of editing a human genomic sequence. 737. The Type II Cas protein, gRNA, system, nucleic acid, or a plurality of nucleic acids for use according to embodiment 736, wherein the human genomic sequence is a CCR5, EMX1, Fas, FANCF, HBB, ZSCAN2, Chr6, ADAMTSL1, B2M, CXCR4, PD1, DNMT1, Match8, TRAC, TRBC, VEGFAsite2, VEGFAsite3, CACNA, HEKsite3, HEKsite4, Chr8, BCR, ATM, HBG1, HPRT, IL2RG, NF1, USH2A, RHO, BcLenh, or CTFR genomic sequence. 738. The Type II Cas protein, gRNA, system, nucleic acid, or a plurality of nucleic acids for use according to embodiment 736, wherein the human genomic sequence is a B2M genomic sequence. 739. The Type II Cas protein, gRNA, system, nucleic acid, or a plurality of nucleic acids for use according to embodiment 736, wherein the human genomic sequence is a TRAC genomic sequence. 740. The Type II Cas protein, gRNA, system, nucleic acid, or a plurality of nucleic acids for use according to embodiment 736, wherein the human genomic sequence is a PD1 genomic sequence. 741. A particle comprising a Type II Cas protein according to any one of embodiments 1 to 287, a gRNA according to any one of embodiments 288 to 608, a system according to of any one of embodiments 609 to 680, a nucleic acid according to any one of embodiments 681 to 730, or a plurality of nucleic acids according to of any one of embodiments 731 to 735. 742. The particle of embodiment 738, which is a lipid nanoparticle, a vesicle, a gold nanoparticle, a viral-like particle (VLP) or a viral particle. 743. The particle of embodiment 742, which is a lipid nanoparticle. 744. The particle of embodiment 742, which is a vesicle. 745. The particle of embodiment 742, which is a gold nanoparticle. 746. The particle of embodiment 742, which is a viral-like particle (VLP). 747. The particle of embodiment 742, which is a viral particle. 748. The particle of embodiment 746, which is an adeno-associated virus (AAV) particle. 749. The particle of embodiment 748, wherein the AAV particle is an AAV2, AAV5, AAV7m8, AAV8, AAV9, AAVrh8r, or AAVrh10 particle. 750. The particle of embodiment 749, wherein the AAV particle is an AAV2 particle. 751. The particle of embodiment 749, wherein the AAV particle is an AAV5 particle. 752. The particle of embodiment 749, wherein the AAV particle is an AAV7m8 particle. 753. The particle of embodiment 749, wherein the AAV particle is an AAV8 particle. 754. The particle of embodiment 749, wherein the AAV particle is an AAV9 particle. 755. The particle of embodiment 749, wherein the AAV particle is an AAVrh8r particle. 756. The particle of embodiment 749, wherein the AAV particle is an AAVrh10 particle. 757. A pharmaceutical composition comprising a Type II Cas protein according to any one of embodiments 1 to 287, a gRNA according to any one of embodiments 288 to 608, a system according to of any one of embodiments 609 to 680, a nucleic acid according to any one of embodiments 681 to 730, or a plurality of nucleic acids according to of any one of embodiments 731 to 735, or a particle according to any one of embodiments 738 to 756 and at least one pharmaceutically acceptable excipient. 758. A cell comprising a Type II Cas protein according to any one of embodiments 1 to 287, a gRNA according to any one of embodiments 288 to 608, a system according to any one of embodiments 609 to 680, a nucleic acid according to any one of embodiments 681 to 730, or a plurality of nucleic acids according to of any one of embodiments 731 to 735, or a particle according to any one of embodiments 738 to 756. 759. The cell of embodiment 758, which is a human cell. 760. The cell of embodiment 758 or embodiment 759, wherein the cell is a hematopoietic progenitor cell. 761. The cell of any one of embodiments 758 to 760, which is a stem cell. 762. The cell of embodiment 761, wherein the stem cell is a hematopoietic stem cell (HSC), a pluripotent stem cell, or an induced pluripotent stem cell (iPS). 763. The cell of embodiment 762, wherein the stem cell is an embryonic stem cell. 764. The cell of embodiment 758 or embodiment 759, wherein the cell is a T cell. 765. The cell of any one of embodiments 758 to 764, which is an ex vivo cell. 766. A population of cells according to any one of embodiments 758 to 765. 767. A method for altering a cell, the method comprising contacting the cell with a Type II Cas protein according to any one of embodiments 1 to 287, a gRNA according to any one of embodiments 288 to 608, a system according to of any one of embodiments 609 to 680, a nucleic acid according to any one of embodiments 681 to 730, or a plurality of nucleic acids according to of any one of embodiments 731 to 735, a particle according to any one of embodiments 738 to 756, or a pharmaceutical composition according to embodiment 757. 768. The method of embodiment 767, which comprises contacting the cell with the Type II Cas protein of any one of embodiments 1 to 287. 769. The method of embodiment 767, which comprises contacting the cell with the gRNA of any one of embodiments 288 to 608. 770. The method of embodiment 767, which comprises contacting the cell with the system of any one of embodiments 609 to 680. 771. The method of embodiment 770, which comprises electroporation of the cell prior to contacting the cell with the system. 772. The method of embodiment 770, which comprises lipid-mediated delivery of the system to the cell, optionally wherein the lipid-mediated delivery is cationic lipid-mediated delivery. 773. The method of embodiment 770, which comprises polymer-mediated delivery of the system to the cell. 774. The method of embodiment 770, which comprises delivery of the system to the cell by lipofection. 775. The method of embodiment 770, which comprises delivery of the system to the cell by nucleofection. 776. The method of embodiment 767, which comprises contacting the cell with the nucleic acid of any one of embodiments 681 to 730. 777. The method of embodiment 767, which comprises contacting the cell with the plurality of nucleic acids of any one of embodiments 731 to 735. 778. The method of embodiment 767, which comprises contacting the cell with the particle of any one of embodiments 738 to 756. 779. The method of embodiment 767, which comprises contacting the cell with the pharmaceutical composition of embodiment 757. 780. The method of any one of embodiments 767 to 779, wherein the contacting alters a CCR5, EMX1, Fas, FANCF, HBB, ZSCAN2, Chr6, ADAMTSL1, B2M, CXCR4, PD1, DNMT1, Match8, TRAC, TRBC, VEGFAsite2, VEGFAsite3, CACNA, HEKsite3, HEKsite4, Chr8, BCR, ATM, HBG1, HPRT, IL2RG, NF1, USH2A, RHO, BcLenh, or CTFR genomic sequence 781. The method of any one of embodiments 767 to 779, wherein the contacting alters a B2M gene. 782. The method of any one of embodiments 767 to 779, wherein the contacting alters a TRAC gene. 783. The method of any one of embodiments 767 to 779, wherein the contacting alters a PD1 gene. 784. The method of any one of embodiments 767 to 779, wherein the contacting alters a CCR5, EMX1, Fas, FANCF, HBB, ZSCAN, Chr6, ADAMTSL1, B2M, CXCR4, PD1, DNMT1, Match8, TRAC, TRBC, VEGFAsite2, VEGFAsite3, CACNA, HEKsite3, HEKsite4, Chr8, or BCR genomic sequence. 785. The method of any one of embodiments 767 to 784, wherein the cell is a human cell. 786. The method of any one of embodiments 767 to 785, wherein the cell is a hematopoietic progenitor cell. 787. The method of any one of embodiments 767 to 786, wherein the cell is a stem cell. 788. The method of embodiment 787, wherein the stem cell is a hematopoietic stem cell (HSC), a pluripotent stem cell, or an induced pluripotent stem cell (iPS). 789. The method of embodiment 788, wherein the stem cell is an embryonic stem cell. 790. The method of any one of embodiments 767 to 785, wherein the cell is a T cell. 791. The method of any one of embodiments 767 to 790, wherein the contacting is in vitro. 792. The method of embodiment 791, further comprising transplanting the cell to a subject. 793. The method of any one of embodiments 767 to 790, wherein the contacting is in vivo in a subject. 794. A cell or population of cells produced by the method of any one of embodiments 767 to 791. 9. CITATION OF REFERENCES [0273] All publications, patents, patent applications and other documents cited in this application are hereby incorporated by reference in their entireties for all purposes to the same extent as if each individual publication, patent, patent application or other document were individually indicated to be incorporated by reference for all purposes. In the event that there is an inconsistency between the teachings of one or more of the references incorporated herein and the present disclosure, the teachings of the present specification are intended.

Claims

WHAT IS CLAIMED IS: 1. A Type II Cas protein comprising an amino acid sequence having at least 50% sequence identity to: (a) the amino acid sequence of a RuvC-I domain of a reference protein sequence; (b) the amino acid sequence of a RuvC-II domain of a reference protein sequence; (c) the amino acid sequence of a RuvC-III domain of a reference protein sequence; (d) the amino acid sequence of a BH domain of a reference protein sequence; (e) the amino acid sequence of a REC1_1 domain of a reference protein sequence; (f) the amino acid sequence of a REC2 domain of a reference protein sequence; (g) the amino acid sequence of a REC1_2 domain of a reference protein sequence; (h) the amino acid sequence of a HNH domain of a reference protein sequence; (i) the amino acid sequence of a Topo domain of a reference protein sequence; (j) the amino acid sequence of a CTD domain of a reference protein sequence; or (k) the amino acid sequence of the full length of a reference protein sequence; wherein the reference protein sequence is SEQ ID NO:1, SEQ ID NO:2, SEQ ID NO:7, SEQ ID NO:8, SEQ ID NO:13, SEQ ID NO:14, SEQ ID NO:19, SEQ ID NO:20, SEQ ID NO:25, SEQ ID NO:26, SEQ ID NO:31, SEQ ID NO:32, SEQ ID NO:37, SEQ ID NO:38, SEQ ID NO:43, SEQ ID NO:44, SEQ ID NO:49, SEQ ID NO:50, SEQ ID NO:55, or SEQ ID NO:56. 2. The Type II Cas protein of claim 1, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 55% identical, at least 60% identical, at least 65% identical, at least 70% identical, at least 75% identical, at least 80% identical, at least 85% identical, at least 90% identical, at least 95% identical, at least 96% identical, at least 97% identical, at least 98% identical, at least 99% identical, or is identical to the full length of the reference protein sequence. 3. The Type II Cas protein of claim 1, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is at least 95% identical, at least 96% identical, at least 97% identical, at least 98% identical, at least 99% identical, or is identical to the full length of the reference protein sequence. 4. The Type II Cas protein of claim 1, wherein the amino acid sequence of the Type II Cas protein comprises an amino acid sequence that is identical to the full length of the reference protein sequence 5. The Type II Cas protein of any one of claims 1 to 4, which is a fusion protein. 6. The Type II Cas protein of claim 5, which comprises one or more nuclear localization signals, such as two or more nuclear localization signals, and which optionally comprises an N-terminal nuclear localization signal and/or a C-terminal nuclear localization signal. 7. The Type II Cas protein of claim 6, wherein the amino acid sequence of one or more of the nuclear localization signals comprises the amino acid sequence KRTADGSEFESPKKKRKV (SEQ ID NO:116), PKKKRKV (SEQ ID NO:117), PKKKRRV (SEQ ID NO:118), KRPAATKKAGQAKKKK (SEQ ID NO:119), YGRKKRRQRRR (SEQ ID NO:120), RKKRRQRRR (SEQ ID NO:121), PAAKRVKLD (SEQ ID NO:122), RQRRNELKRSP (SEQ ID NO:123), VSRKRPRP (SEQ ID NO:124), PPKKARED (SEQ ID NO:125), PQPKKKPL (SEQ ID NO:126), SALIKKKKKMAP (SEQ ID NO:127), PKQKKRK (SEQ ID NO:128), RKLKKKIKKL (SEQ ID NO:129), REKKKFLKRR (SEQ ID NO:130), KRKGDEVDGVDEVAKKKSKK (SEQ ID NO:131), RKCLQAGMNLEARKTKK (SEQ ID NO:132), NQSSNFGPMKGGNFGGRSSGPYGGGGQYFAKPRNQGGY (SEQ ID NO:133), RMRIZFKNKGKDTAELRRRRVEVSVELRKAKKDEQILKRRNV (SEQ ID NO:134). 8. The Type II Cas protein of claim 6, wherein the amino acid sequence of one or more of the nuclear localization signals comprises the amino acid sequence PAAKKKKLD (SEQ ID NO:582) or KRPAATKKAGQAKKKKLD (SEQ ID NO:583) 9. The Type II Cas protein of any one of claims 6 to 8, wherein the amino acid sequence of each nuclear localization signal is the same. 10. The Type II Cas protein of any one of claims 5 to 9, which comprises a fusion partner which is a DNA, RNA or protein modification enzyme, optionally wherein the DNA, RNA or protein modification enzyme is an adenosine deaminase, a cytidine deaminase, a reverse transcriptase, a guanosyl transferase, a DNA methyltransferase, a RNA methyltransferase, a DNA demethylase, a RNA demethylase, a dioxygenase, a polyadenylate polymerase, a pseudouridine synthase, an acetyltransferase, a deacetylase, a ubiquitin-ligase, a deubiquitinase, a kinase, a phosphatase, a NEDD8-ligase, a de-NEDDylase, a SUMO-ligase, a deSUMOylase, a histone deacetylase, a histone acetyltransferase, a histone methyltransferase, or a histone demethylase. 11. The Type II Cas protein of any one of claims 5 to 10, which comprises (a) a fusion partner which is an adenosine deaminase, optionally wherein the amino acid sequence of the adenosine deaminase comprises an amino acid sequence having at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, at least 95%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with SEQ ID NO:137, optionally wherein the adenosine deaminase is the adenosine deaminase moiety contained in the adenine base editor ABE8e; (b) a fusion partner which is a cytidine deaminase; or (c) a fusion partner which is a reverse transcriptase. 12. The Type II Cas protein of any one of claims 5 to 11, which comprises a tag, e.g., a SV5 tag, optionally wherein the SV5 tag comprises the amino acid sequence GKPIPNPLLGLDST (SEQ ID NO:135). 13. The Type II Cas protein of any one of claims 1 to 12, wherein (a) the reference protein sequence is SEQ ID NO:1 or SEQ ID NO:2, optionally wherein the amino acid sequence of the Type II Cas protein comprises the amino acid sequence of SEQ ID NO:1, SEQ ID NO:2, or SEQ ID NO:3; (b) the reference protein sequence is SEQ ID NO:7 or SEQ ID NO:8, optionally wherein the amino acid sequence of the Type II Cas protein comprises the amino acid sequence of SEQ ID NO:7, SEQ ID NO:8, or SEQ ID NO:9; (c) the reference protein sequence is SEQ ID NO:13 or SEQ ID NO:14, optionally wherein the amino acid sequence of the Type II Cas protein comprises the amino acid sequence of SEQ ID NO:13, SEQ ID NO:14 or SEQ ID NO:15; (d) the reference protein sequence is SEQ ID NO:19 or SEQ ID NO:20, optionally wherein the amino acid sequence of the Type II Cas protein comprises the amino acid sequence of SEQ ID NO:19, SEQ ID NO:20 or SEQ ID NO:21; (e) the reference protein sequence is SEQ ID NO:25 or SEQ ID NO:26, optionally wherein the amino acid sequence of the Type II Cas protein comprises the amino acid sequence of SEQ ID NO:25, SEQ ID NO:26 or SEQ ID NO:27; (f) the reference protein sequence is SEQ ID NO:31 or SEQ ID NO:32, optionally wherein the amino acid sequence of the Type II Cas protein comprises the amino acid sequence of SEQ ID NO:31, SEQ ID NO:32 or SEQ ID NO:33; (g) the reference protein sequence is SEQ ID NO:37 or SEQ ID NO:38, optionally wherein the amino acid sequence of the Type II Cas protein comprises the amino acid sequence of SEQ ID NO:37, SEQ ID NO:38, or SEQ ID NO:39; (h) the reference protein sequence is SEQ ID NO:43 or SEQ ID NO:44, optionally wherein the amino acid sequence of the Type II Cas protein comprises the amino acid sequence of SEQ ID NO:43, SEQ ID NO:44, SEQ ID NO:45; (i) the reference protein sequence is SEQ ID NO:49 or SEQ ID NO:50, optionally wherein the amino acid sequence of the Type II Cas protein comprises the amino acid sequence of SEQ ID NO:49, SEQ ID NO:50, SEQ ID NO:51; (j) the reference protein sequence is SEQ ID NO:55 or SEQ ID NO:56, optionally wherein the amino acid sequence of the Type II Cas protein comprises the amino acid sequence of SEQ ID NO:55, SEQ ID NO:56, SEQ ID NO:57. 14. A Type II Cas protein whose amino acid sequence is identical to a Type II Cas protein of any one of claims 1 to 13 except for one or more amino acid substitutions relative to the reference sequence that provide nickase activity, optionally wherein the one or more amino acid substitutions comprise a substitution (e.g., alanine substitution) at a position corresponding to position D10 of SpCas9 or H840 of SpCas9 (e.g., as shown in Table 3). 15. A Type II Cas protein whose amino acid sequence is identical to a Type II Cas protein of any one of claims 1 to 13 except for one or more amino acid substitutions relative to the reference sequence that render the Type II Cas protein catalytically inactive, optionally wherein the one or more amino acid substitutions comprise a substitution (e.g., alanine substitution) at a position corresponding to position D10 of SpCas9 or H840 of SpCas9 (e.g., as shown in Table 3), or a combination thereof. 16. A guide RNA (gRNA) molecule for editing a human B2M gene comprising a spacer whose nucleotide sequence comprises 15 or more consecutive nucleotides of a reference sequence or comprises a nucleotide sequence that is at least 85% identical to the reference sequence, wherein the reference sequence is selected from SEQ ID NOs:138-162. 17. A guide RNA (gRNA) molecule for editing a human TRAC gene comprising a spacer whose nucleotide sequence comprises 15 or more consecutive nucleotides of a reference sequence or comprises a nucleotide sequence that is at least 85% identical to the reference sequence, wherein the reference sequence is selected from SEQ ID NOs:163-194. 18. A guide RNA (gRNA) molecule for editing a human PD1 gene comprising a spacer whose nucleotide sequence comprises 15 or more consecutive nucleotides of a reference sequence or comprises a nucleotide sequence that is at least 85% identical to the reference sequence, wherein the reference sequence is selected from SEQ ID NOs:195-217. 19. The gRNA of any one of claims 16 to 18, which comprises a spacer that is 15 to 30 nucleotides in length, 18 to 30 nucleotides in length, 20 to 28 nucleotides in length, 22 to 26 nucleotides in length, 23 to 25 nucleotides in length, 22 to 25 nucleotides in length, 15 to 25 nucleotides in length, 16 to 24 nucleotides in length, 17 to 23 nucleotides in length, 18 to 22 nucleotides in length, 19 to 21 nucleotides in length, 25 nucleotides in length, 24 nucleotides in length, 23 nucleotides in length, 22 nucleotides in length, 21 nucleotides in length, or 20 nucleotides in length. 20. The gRNA of any one of claims 16 to 19, wherein the spacer comprises the reference sequence. 21. The gRNA of any one of claims 16 to 20, which is a single guide RNA (sgRNA). 22. A gRNA comprising a spacer and a sgRNA scaffold, wherein: (a) WKH^VSDFHU^LV^SRVLWLRQHG^^¶^WR^WKH^VJ51$^VFDIIROG^^DQG (b) the nucleotide sequence of the sgRNA scaffold comprises a nucleotide sequence that is at least 50% identical to a reference scaffold sequence, wherein the reference scaffold sequence is any one of SEQ ID NOS:81-102. 23. The gRNA of claim 22, wherein the sgRNA scaffold comprises a nucleotide sequence that is at least 55% identical, at least 60% identical, at least 65% identical, at least 70% identical, at least 75% identical, at least 80% identical, at least 85% identical, at least 90% identical, at least 95% identical, at least 96% identical, at least 97% identical, at least 98% identical, or at least 99% identical to the reference scaffold sequence; comprises a nucleotide sequence that has no more than 5 nucleotide mismatches, no more than 4 nucleotide mismatches, no more than 3 nucleotide mismatches, no more than 2 nucleotide mismatches, or no more than 1 nucleotide mismatch with the reference scaffold sequence; or comprises a nucleotide sequence that is 100% identical to the reference scaffold sequence. 24. The gRNA of claim 22 or claim 23, wherein the sgRNA scaffold comprises 1 to 8 uracils DW^LWV^^¶^HQG^ 25. The gRNA of any one of claims 22 to 24, wherein the nucleotide sequence of the spacer is partially or fully complementary to a target mammalian genomic sequence. 26. A gRNA comprising (i) a crRNA comprising a spacer and a crRNA scaffold, wherein the VSDFHU^LV^^¶^WR^WKH^FU51$^VFDIIROG^^DQG^^LL^^D^WracrRNA, wherein the nucleotide sequence of the spacer is partially or fully complementary to a target mammalian genomic sequence and the nucleotide sequence of the crRNA scaffold comprises the nucleotide sequence of SEQ ID NO:61, SEQ ID NO:63, SEQ ID NO:65, SEQ ID NO:67, SEQ ID NO:69, SEQ ID NO:71, SEQ ID NO:73, SEQ ID NO:75, SEQ ID NO:77, or SEQ ID NO:79. 27. The gRNA of claim 26, wherein the gRNA comprises separate crRNA and tracrRNA molecules. 28. The gRNA of claim 26, wherein the gRNA is a single guide RNA (sgRNA). 29. The gRNA of any one of claims 22 to 28, wherein the target mammalian genomic sequence is a human genomic sequence. 30. The gRNA of any one of claims 22 to 29, wherein the target mammalian genomic sequence is upstream of a protospacer adjacent motif (PAM) sequence in the non-target strand recognized by a Type II Cas protein, optionally wherein the Type II Cas protein is a Type II Cas protein according to any one of claims 1 to 15. 31. The gRNA of claim 30, wherein the PAM sequence is NGG, NNGNC, NNGNCNY, NNRC, NNAC, NNACNT, NNRTA, NNNNCC, NNANCC, NNACCC, NNAT, NNATM, NNATA, NVAT, NVATM, NNNAC, NRNAC, NRHAC, NRNACD, NRHACD, NRNACA, NRHACA, NNNRNY, NRNRNY,NRNRNC, NANANC, NNNRDC, NNART, or NRART. 32. The gRNA of claim 30, wherein the PAM sequence is NVRTA, NRRTA, NVATA, or NRATA. 33. The gRNA of any one of claims 22 to 32, wherein the spacer is 15 to 30 nucleotides in length.

34. A system comprising the Type II Cas protein of any one of claims 1 to 15 and a guide RNA (gRNA) comprising a spacer sequence, optionally wherein the gRNA is a gRNA according to any one of claims 16 to 33. 35. A nucleic acid encoding the Type II Cas protein of any one of claims 1 to 15, optionally wherein the nucleotide sequence encoding the Type II Cas protein is operably linked to a promoter that is heterologous to the Type II Cas protein. 36. A nucleic acid encoding the gRNA of any one of claims 16 to 33. 37. A nucleic acid encoding the Type II Cas protein and gRNA of the system of claim 34. 38. A plurality of nucleic acids comprising separate nucleic acids encoding the Type II Cas protein and gRNA of the system of claim 34. 39. A Type II Cas protein according to any one of claims 1 to 15, a gRNA according to any one of claims 16 to 33, a system according to claim 34, a nucleic acid according to any one of claims 35 to 37, or a plurality of nucleic acids according to claim 38 for use in a method of editing a human genomic sequence. 40. A particle comprising a Type II Cas protein according to any one of claims 1 to 15, a gRNA according to any one of claims 16 to 33, a system according to claim 34, a nucleic acid according to any one of claims 35 to 37, or a plurality of nucleic acids according to claim 38. 41. A pharmaceutical composition comprising a Type II Cas protein according to any one of claims 1 to 15, a gRNA according to any one of claims 16 to 33, a system according to claim 34, a nucleic acid according to any one of claims 35 to 37, or a plurality of nucleic acids according to claim 38, or a particle according to claim 40 and at least one pharmaceutically acceptable excipient. 42. An ex vivo cell comprising a Type II Cas protein according to any one of claims 1 to 15, a gRNA according to any one of claims 16 to 33, a system according to claim 34, a nucleic acid according to any one of claims 35 to 37, or a plurality of nucleic acids according to claim 38, or a particle according to claim 40. 43. A method for altering an ex vivo cell, the method comprising contacting the cell with a Type II Cas protein according to any one of claims 1 to 15, a gRNA according to any one of claims 16 to 33, a system according to claim 34, a nucleic acid according to any one of claims 35 to 37, a plurality of nucleic acids according to claim 38, a particle according to claim 40, or a pharmaceutical composition according to claim 41. 44. An ex vivo cell or population of cells produced by the method of claim 43.