Protoplast (from Ancient Greek πρωτόπλαστος (prōtóplastos) 'first-formed'), is abiological term coined byHanstein in 1880 to refer to the entire cell, excluding the cell wall.[1][2] Protoplasts can be generated by stripping thecell wall fromplant,[3]bacterial,[4][5] orfungal cells[5][6] by mechanical, chemical or enzymatic means.
Protoplasts differ fromspheroplasts in that their cell wall has been completely removed.[4][5] Spheroplasts retain part of their cell wall.[7] In the case ofGram-negative bacterial spheroplasts, for example, thepeptidoglycan component of the cell wall has been removed but theouter membrane component has not.[4][5]
Cell walls are made of a variety ofpolysaccharides. Protoplasts can be made by degrading cell walls with a mixture of the appropriate polysaccharide-degradingenzymes:
| Type of cell | Enzyme |
|---|---|
| Plant cells | Cellulase,pectinase,xylanase[3] |
| Gram-positive bacteria | Lysozyme,N,O-diacetylmuramidase,lysostaphin[4] |
| Fungal cells | Chitinase[6] |
During and subsequent to digestion of the cell wall, the protoplast becomes very sensitive toosmotic stress. This means cell wall digestion and protoplast storage must be done in anisotonic solution to prevent rupture of theplasma membrane.[citation needed]
Protoplasts can be used to studymembrane biology, including the uptake ofmacromolecules andviruses . These are also used insomaclonal variation.
Protoplasts are widely used forDNAtransformation (for makinggenetically modified organisms), since the cell wall would otherwise block the passage of DNA into the cell.[3] In the case of plant cells, protoplasts may be regenerated into whole plants first by growing into a group of plant cells that develops into acallus and then by regeneration of shoots (caulogenesis) from the callus usingplant tissue culture methods.[8] Growth of protoplasts into callus and regeneration of shoots requires the proper balance of plant growth regulators in the tissue culture medium that must be customized for each species of plant.[9][10] Unlike protoplasts fromvascular plants, protoplasts frommosses, such asPhyscomitrella patens, do not needphytohormones for regeneration, nor do they form a callus duringregeneration. Instead, they regenerate directly into the filamentousprotonema, mimicking a germinating moss spore.[11]
Protoplasts may also be used forplant breeding, using a technique calledprotoplast fusion. Protoplasts from different species are induced to fuse by using anelectric field or a solution ofpolyethylene glycol.[12] This technique may be used to generatesomatic hybrids in tissue culture.[citation needed]
Additionally, protoplasts of plants expressing fluorescent proteins in certain cells may be used for Fluorescence Activated Cell Sorting (FACS), where only cells fluorescing a selected wavelength are retained. Among other things, this technique is used to isolate specific cell types (e.g.,guard cells from leaves,pericycle cells from roots) for further investigations, such as transcriptomics.[citation needed]
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