A kind of absolute quantitation digital nucleic acid analysis system based on efficient drop microreactorTechnical field
The present invention relates to the inspection and quarantine system such as biomedicine, agricultural, food hygiene, specifically a kind of based on efficientlyThe absolute quantitation digital nucleic acid analysis system of drop microreactor, mainly for detection of the molecular structure of nucleic acid, so that analyze lifeThe attribute of thing, belongs to biological, medical science.
Background technology
Nucleic acid is biological internal macromolecular compound.Including DNA (deoxyribonucleic acid,) and ribonucleic acid (ribonucleic acid, RNA) two big class DNA.Nucleic acid is not only basic inhereditary material, and in eggAlso critical positions are accounted in the biosynthesis of white matter, thus in a series of great biological phenomenas such as growth, heredity, variation play decisionThe effect of property.Nucleic acid analysis technique is advantageous in that its high sensitivity and specificity, by nucleic acid amplification technologies and miscellaneousFriendship technology is tested and analyzed to nucleic acid, so as to can achieve but being not limited to, the microorganism detection such as bacterium, virus, the disease such as tumourThe biomedical sectors such as diagnosis, antenatal lossless DNA diagnosis, the Risk-warning of genetic disease, and drug screening, accurate medical treatment shouldWith.
The premise of foranalysis of nucleic acids is nucleic acid amplification, i.e., PCR (Polymerase Chain Reaction,PCR), claim PCR amplifications, reaction is entered by specific enzymatic in vitro, one section of specific DNA or RNA fragment amplification is carried out, so as to obtainTake substantial amounts of target DNA fragments.Method for nucleic acid analysis experienced three developing stage.First stage is exactly common PCR amplificationsInstrument, the template DNA after amplification qualitatively detect the presence or absence of target fragment, and starting template can not be entered through gel electrophoresis analysisRow quantitative determination.Second stage, adds fluorophor in pcr amplification reaction system, is supervised using fluorescence signal accumulation in real timeWhole PCR processes are surveyed, and the cycle-index experienced when threshold value is reached finally by amplified production and calibration curve carry out quantitative startingThe concentration of template.Real-time fluorescence quantitative PCR using between calibration object and sample relatively carrying out relative quantification detection, calibration objectDeviation is introduced as background is different with sample room, for the target DNA of low copy number is difficult to detect.Simultaneously in DNA extraction processThe impurity of introducing may produce inhibitory action to amplification.It is the QX200 drop formulas of Bio-Rad companies of the U.S. that three phases are representedDigital pcr, it is characterised in that carry out microlayer model process to sample first, the reaction system containing nucleic acid molecules is separated intoThe drop of thousands of nanoliter levels, wherein each droplet are made without or with a nucleic acid target molecule to be checked, each dropletFor an independent PCR reactor, after expanding through PCR, the drop for having fluorescence signal is 1, the drop 0 without fluorescence signal, mostEventually according to Poisson distribution principle and the ratio of positive drop, analysis software can calculate the concentration or copy for providing target molecule to be checkedNumber.There is drop formula digital pcr high amplification efficiency and specificity, detection sensitivity can reach 0.001%.Divided by countingAnalysis testing result, it is not necessary to calibration curve, can achieve absolute quantitation.The sample detection of QX200 drop formula digital pcrs needs threeInstrument divides three steps to complete, and produces in instrument in drop first and produces drop, is transferred to PCR expansions with PCR pipe after collected dropIncrease and expanded in instrument, amplification is transferred to amplified production in digital independent analyzer after terminating again and carries out result reading, analyzeProcess is loaded down with trivial details, complicated, the cycle is long, is not suitable for carrying out quick foranalysis of nucleic acids and detection.
Content of the invention
For the above-mentioned weak point of prior art, it is an object of the invention to provide a kind of be based on the micro- reaction of efficient dropThe absolute quantitation digital nucleic acid analysis system of device.
The purpose of the present invention is achieved through the following technical solutions：
The present invention includes confocal fluorescent image capturing system, for carrying out fluorescence excitation to micro-fluidic chip, and gathersExcite the image of the sample in light area；
Micro-fluidic chip, is placed in temperature cycles heating system；
Temperature cycles heating system, on electric loading system platform, under the control of the controller, to micro-fluidic corePiece is circulated heating；
Electric loading system platform, for carrying micro-fluidic chip and temperature cycles heating system, and then drives micro-fluidic chipCarry out horizontal rectilinear motion or vertical linear motion or curvilinear motion；
Automatic sampling apparatus, inject for sample, realize that the drop on micro-fluidic chip produces and make the drop of generation to enterEnter in microreactor；
Controller, connects and controls confocal fluorescent image capturing system, temperature cycles heating system, electric loading system platformWork with automatic sampling apparatus；
Computer, connects controller, for being filtered the image of the PCR microreactors on the micro-fluidic chip for collectingRipple, rim detection, histogram calculation, determine detection limit, binary conversion treatment, calculate light field under all amount of droplets, calculate fluorescenceAmount of droplets, the absolute template number of analytical calculation and sample concentration off field；
Wherein:The confocal fluorescent image capturing system is located at the top of micro-fluidic chip, excites on micro-fluidic chipPCR microreactors region, and the image of light area is gathered, the confocal fluorescent image capturing system includes image sensingDevice, optics microscope group and optical filter, also include：
Excitation source, for sending light source under the control of the controller, is irradiated on dichroscope；
Dichroscope, for reflecting the light source that the excitation source sends, makes reflected light be irradiated on micro-fluidic chipAny one PCR microreactors region, excites positive template to send fluorescence, and the fluorescence for sending passes through after the dichroscope transmissionThe optical filter and optics microscope group are implemented as picture to imageing sensor；
The automatic sampling apparatus include that power source, executing agency and air-breather, wherein executing agency are connected to powerThe output end in source, the actuating station of the executing agency are connected with air-breather；The air-breather includes breather pipe, normally closed electromagnetismValve, threeway, negative-pressure vacuum pump and vacuum cup, the threeway are connected to the actuating station of the executing agency, and one end connection is described trueSuction disk, the other end are connected with the negative-pressure vacuum pump by breather pipe, and the 3rd end is connected with air by breather pipe, and withThe breather pipe that air is connected is provided with the normally closed solenoid valve；The power source, negative-pressure vacuum pump and normally closed solenoid valve respectively withThe controller electrical connection；
The micro-fluidic chip includes that multiple droplet generators and multiple PCR microreactors, each PCR microreactor haveEach independent droplet generator, respective injection port and oil phase injection port, multiple PCR microreactors share one and go outMouthful, described export for connecting automatic sampling apparatus, being generated by the automatic sampling apparatus vacuum suction drop enters whichEnter in each PCR microreactors；Each described PCR microreactor is connected with outlet by passage respectively, in each passageOn be equipped with to liquid flow backwards produce cushioning effect buffering area；The vacuum cup of the air-breather is with executing agency's movement coverIn the outlet of the micro-fluidic chip, the sample in injection port is pumped into by PCR microreactors by the negative-pressure vacuum pumpIn carry out reacting, detect；External diameter of the internal diameter of the vacuum cup more than the outlet of micro-fluidic chip, the vacuum cup is fallen micro-When in the outlet of fluidic chip by described outlet cover, and not with the outlet contact, and with micro-fluidic chip sealing absorption；
The executing agency is slide block guide rail executing agency, including vertical guide rail, slide block A, horizontal guide rail, position sensingDevice, gangbar, slide block B and cylinder B, the power source are cylinder A, and the output end of cylinder A is connected with slide block A, the slide block AIt is slidably connected with the vertical guide rail on cylinder A, the horizontal guide rail is set on slide block A on the horizontal guide railThe output end for having cylinder B, the cylinder B is connected with the slide block B being slidably connected with horizontal guide rail, and the vertical guide rail and level are ledThe position sensor being electrically connected with the controller is separately installed with rail；The threeway is connected in the slide block B by gangbar；
The executing agency is screw-nut executing agency, including leading screw A, screw A, motor B, screw B, leading screw B, positionSensor and gangbar, the power source are motor A, and the output end of motor A is connected with leading screw A, the screw A and leading screw AThreaded connection, on screw A, output end is connected with the leading screw B to the motor B, and the screw B is threadedly connected to silkOn thick stick B, it is separately installed with, on the leading screw A and leading screw B, the position sensor being electrically connected with the controller；The threeway is by linkageBar is connected on screw B；
One circulating-heating process of the temperature cycles heating system includes three temperature sections：
1) denaturation stage continues t1Second；
2) annealing stage continues t2Second；
3) extend stage lasting t3Second；
Or, the temperature cycles heating system only includes a thermostat temperature section；
Or, the temperature cycles heating system only includes the temperature section of two alternating temperatures；
Each phase temperature value and duration are arranged on computers；
The electric loading system platform is two-dimentional, three-dimensional, multidimensional or rotary motion platform；
The controller includes
Microprocessor, for controlling circulating-heating drive module, electric platforms drive module, light source driver module, vacuumPump drive module, sample introduction Locating driver module and communication module；
Communication module, connects microprocessor, for the interactive communication between controller and computer；
Light source driver module, is connected with microprocessor, for connecting confocal fluorescent image capturing system, and drives copolymerizationExcitation source in burnt fluoroscopic image acquisition system sends exciting light；
Circulating-heating drive module, is connected with microprocessor, hydronic each for controlling temperature cycles heating systemDuan Wendu, the heat time for controlling each temperature section and cycle-index；
Electric platforms drive module, is connected with microprocessor, for controlling electric loading system platform in level, vertical directionLinear motion, or rotate clockwise or counterclockwise；
Vavuum pump drive module, is connected with microprocessor, for controlling to drive vavuum pump action, is that automatic sampling apparatus are producedRaw negative pressure drives；
Sample introduction Locating driver module, is connected with microprocessor, for controlling the vacuum cup and miniflow of automatic sampling apparatusThe outlet docking of control chip, realizes sample introduction.
Advantages of the present invention with good effect is：
Relative to existing absolute quantitation foranalysis of nucleic acids system, advantage of the invention is that enormously simplify absolute quantitation coreThe complexity of acid analysis process, the detection sensitivity that improve absolute quantitation foranalysis of nucleic acids process and ageing, greatly reduceThe sample usage amount of absolute quantitation foranalysis of nucleic acids, realizes that high flux is detected.Advantageous characteristic of the present invention be microlayer model generation,Pcr amplification reaction is completed on a piece of micro-fluidic chip, and the micro-fluidic chip had not only completed drop and produced function but also complete PCR expansionsIncrease (microreactor) function；PCR amplification after optical sampling is carried out to drop by special image collecting device, afterwards again byEfficiently software for calculation carries out foranalysis of nucleic acids.Present system can achieve the micro-example detection of picoliters level, detectable limit as little asIndividual template (detection molecules).Article points are described below：
1. complete in microreactor of the reaction system on micro-fluidic chip, sample is extremely micro, realize that picoliters level is detected.
2. quick detection.Pre-treatment only slightly need to be carried, and without the need for culture or increasing bacterium process, all processes are completed for 2～5 hours.
3. high detection sensitivity, as little as one To Template of detectable limit.
4. micro- reaction is realized on chip, easily increase reaction member quantity, realize high flux.
5. the parallel detection of multiple samples can be realized on the same chip, detection efficiency is greatly improved.
6. drop is produced, PCR is expanded and Data Detection one-key operation, it is not necessary to manual intervention.
7. one-touch sample introduction, nucleic acid amplification and the fluoroscopic examination of Based Intelligent Control is can achieve, is calculated with efficient nucleic acid and is dividedAnalysis software and the man-machine interface of close friend.
8. relative to various sample injection methods such as existing malleation, negative pressure, electronic, gravity, the automatic sampling apparatus of the present inventionAdvantage no longer needs manual intervention after being to inject a sample into the injection port of micro-fluidic chip, and whole sample introduction process is by controllerControl is one-touch to be completed, full intellectualized.
9. automatic sampling apparatus adopt Ngatively pressurized sampling, as long as there is one outlet, no matter have several injection ports, all only need oneNegative-pressure vacuum pump can be completed.
10. automatic sampling apparatus wide adaptability, is suitable for the micro-fluidic chip sample introduction of various channel types.
Description of the drawings
Fig. 1 is that the system of the present invention constitutes block diagram；
Fig. 2 is one of micro-fluidic chip form of the present invention (8 PCR microreactors)；
Two (4 PCR microreactors) of the Fig. 3 for micro-fluidic chip form of the present invention；
Fig. 4 is system controller block diagram；
Fig. 5 is foranalysis of nucleic acids systematic functional structrue figure；
Fig. 6 is the structural representation of automatic sampling apparatus of the present invention；
Fig. 7 is the structural representation of automatic sampling apparatus another kind executing agency of the present invention；
Fig. 8 is the structural representation of air-breather in automatic sampling apparatus of the present invention；
Wherein：1 is imageing sensor, and 2 is optics microscope group, and 3 is optical filter, and 4 is dichroscope, and 5 is excitation source, and 6 areMicro-fluidic chip, 601 is PCR microreactors, and 602 is injection port, and 603 is droplet generator, and 604 are outlet, and 605 is passage,606 is buffering area, and 607 is oil phase injection port, and 7 is temperature cycles heating system, and 8 is electric loading system platform, and 9 fill for auto injectionPut, 10 is controller, and 11 is computer, and 12 is power source, and 13 is executing agency, and 1301 is vertical guide rail, and 1302 lead for levelRail, 1303 is slide block A, and 1304 is position sensor, and 1305 is gangbar, and 1306 is slide block B, and 1307 is leading screw A, and 1308 is silkFemale A, 1309 is motor B, and 1310 is screw B, and 1311 is leading screw B, and 1312 is cylinder B, and 14 is air-breather, and 1401 are ventilationPipe, 1402 is normally closed solenoid valve, and 1403 is threeway, and 1404 is negative-pressure vacuum pump, and 1405 is vacuum cup.
Specific embodiment
The invention will be further described below in conjunction with the accompanying drawings.
The present invention is such as schemed for can achieve the one-touch sample introduction of Based Intelligent Control, nucleic acid amplification, fluoroscopic examination foranalysis of nucleic acids systemShown in 1, including confocal fluorescent image capturing system, micro-fluidic chip 6, temperature cycles heating system 7, electric loading system platform 8,Automatic sampling apparatus 9, controller 10 and computer 11, wherein confocal fluorescent image capturing system include imageing sensor 1, lightMicroscope group 2 and excitation source 5 is learned, imageing sensor 1 includes but is not limited to the image collecting devices such as CCD camera, CMOS cameras, opticsMicroscope group 2 includes but is not limited to the parts such as optical lens, dichroscope 4, optical filter 3, and excitation source 5 includes but is not limited to LED lightSource, LASER Light Source, halogen light source etc..
As shown in figure 1, add the micro-fluidic chip after sample 6 being put on electric loading system platform 8, electric loading system platform is placed inAlso have temperature cycles heating system 7, the temperature cycles heating system 7 on 8 is located at below micro-fluidic chip 6.Auto injection is filledThe outlet on the control alignment micro-fluidic chip 6 of 9 via controller 10 is put, negative pressure vacuumizes generation drop and feeds them into micro- reactionIn device.Start-up temperature loop heating system 7 starts to enter PCR microreactors performing PCR amplification, after amplification terminates, opens exciting lightSource 5, exciting light are radiated at microreactor region after the reflection of dichroscope 4, excite To Template to send fluorescence, sent fluorescenceTransmitted through dichroscope 4, filtered 3 optical filtering, optics microscope group 2 are irradiated on imageing sensor 1 after focusing on, so as to gather quiltSurvey sample image.After gathered image uploads to computer, image is filtered through network analysis software, rim detection, straightSide figure calculate, determine detection limit, binary conversion treatment, calculate light field under all amount of droplets, calculate fluorescence off field amount of droplets, pointAnalysis calculates absolute template number and detection sample concentration etc..
Micro-fluidic chip 6 includes multiple droplet generators 603 and multiple equally distributed PCR microreactors 601, eachPCR microreactors 601 have each independent droplet generator 603, respective injection port 602 and oil phase injection port 607, eachDroplet generator 603 is connected with injection port 602 and oil phase injection port 607 respectively, can detect multiple different samples simultaneously, orMultiple different target templates in same sample；All of droplet generator can share one or more oil phases and surface-activeAgent import, if using multiple oil phases and surfactant import, these imports are also be interconnected.The micro- reactions of each PCRThere are columnar projections or the interception dam for lying across in device 601, PCR reactors 601 are divided into tens thousand of to hundreds thousand of cells, are used forFixed drop.Multiple PCR microreactors 601 share one outlet 604, and outlet 604 is used for connecting bearing in automatic sampling apparatus 9Pressure vavuum pump, generates drop by 9 vacuum suction of automatic sampling apparatus and makes it in each PCR microreactors 601；EachPCR microreactors 601 are connected with outlet 604 by passage 605 respectively, on each passage 605 are equipped with buffering area 606, are delayedArea 606 is rushed for snakelike (but be not limited to snakelike, including one section of straight channel or water storage type circular hole), when negative-pressure vacuum pump is withdrawn to liquidThe refluence of body produces a cushioning effect.The PCR microreactors of the present invention are also doubled as detection zone, and sample is full of the micro- reactions of PCRAfter device 601, detection sensor feeds back signal to controller 10, and controller 10 stops negative-pressure vacuum pump 1404 immediately and works, and stopsSample introduction.
The PCR microreactors of the present invention can be multiple, eight PCR microreactors as shown in Figure 2, or such as institute in Fig. 3The four PCR microreactors for showing, but it is not limited to eight or four.As shown in Fig. 2 there are eight circle PCR microreactors 601,After collection completes a PCR microreactor image, the electric loading system platform 8 in Fig. 1 rotates clockwise 45° angle, starts to gather theTwo PCR microreactor images；By that analogy, IMAQ and the process of whole eight PCR microreactors are completed.AnotherSituation, electric loading system platform 8 can rotate counterclockwise complete the collection of PCR microreactor images.Can be suitable with electric loading system platform 8Hour hands rotation completes a part of PCR microreactors IMAQ, and then rotate counterclockwise completes another part PCR microreactor figuresAs collection.As shown in figure 3, after there are four PCR microreactors 601, collection to complete a PCR microreactor image, in Fig. 1Electric loading system platform 8 rotates clockwise 90 ° of angles, starts to gather second PCR microreactor image；By that analogy, complete allThe IMAQ of four PCR microreactors and process.Another situation, electric loading system platform 8 can rotate counterclockwise complete PCRThe collection of microreactor image.Can be turned clockwise with electric loading system platform 8 and complete a part of PCR microreactors image and adoptCollection, then rotate counterclockwise completes another part PCR microreactor IMAQs.
Another situation, if the non-annular array as shown in Figure 2 or Figure 3 of microreactor, and using arranging in length and breadth, thenElectric loading system platform 8 is moved according to parallel or vertical direction, completes the collection of all images.
As shown in fig. 6, automatic sampling device 9 includes power source 12, executing agency 13 and air-breather 14, machine is wherein executedStructure 13 is connected to the output end of power source 12, is connected with air-breather 14 in the actuating station of executing agency.
As shown in figure 8, air-breather 14 includes breather pipe 1401, normally closed solenoid valve 1402, threeway 1403, negative-pressure vacuum pump1404 and vacuum cup 1405, the top of the threeway 1403 is connected to the actuating station of executing agency, and one end connects vacuum cup1405, the other end is connected with negative-pressure vacuum pump 1404 by breather pipe 1401, and the 3rd end is connected with air by breather pipe 1401,And normally closed solenoid valve 1402 is provided with the breather pipe 1401 being connected with air.The internal diameter of vacuum cup 1405 is more than micro-fluidic coreThe external diameter of the outlet 604 of piece 6, when vacuum cup 1405 is fallen in outlet 604, it is ensured that vacuum cup 1405 will outlet604 cover, and do not contact with outlet 604, so as to avoid cross pollution.The vacuum cup 1405 of the present invention is with elasticitySilica gel material, when vacuum cup 1405 is covered in outlet 604 with the sealing absorption of micro-fluidic chip 6, can play wellSealing function, it is ensured that enough vacuum realizes Ngatively pressurized sampling.
As shown in fig. 6, executing agency 13 is slide block guide rail executing agency, including vertical guide rail 1301, slide block A1303, waterLevel gauge 1302, slide block B 1306, position sensor 1304, gangbar 1305 and cylinder B1312, power source 12 are cylinder A, shouldThe output end of cylinder A is connected with slide block A1303, and slide block A1303 is slidably connected with the vertical guide rail 1301 on cylinder A；Horizontal guide rail 1302 is provided with cylinder B1312, the output of cylinder B1312 on the horizontal guide rail 1302 on slide block A1303End is connected with the slide block B 1306 being slidably connected with horizontal guide rail 1302, and the top of threeway 1403 is connected to by gangbar 1305In the slide block B 1306.The position electrically connected with controller 10 is separately installed with vertical guide rail 1301 and horizontal guide rail 1302Sensor 1304.
The executing agency 13 of the present invention is not limited to guide rail motion, also can be as shown in fig. 7, executing agency 13 is held for screw-nutRow mechanism, including leading screw A1307, screw A1308, motor B1309, screw B1310, position sensor 1304, gangbar 1305And leading screw B1311, power source 12 is motor A, and the output end of motor A is connected with leading screw A1307, screw A1308 and leading screwA1307 is threadedly coupled, and on screw A1308, output end is connected with leading screw B1311, screw B1310 spiral shells to motor B1309Line is connected on leading screw B1311, and the top of threeway 1403 is connected on screw B1310 by gangbar 1305.In leading screwThe position sensor 1304 electrically connected with controller 10 is separately installed with A1307 and leading screw B1311.
Or, executing agency can also be rotary motion, drive air-breather to reach the outlet 604 on micro-fluidic chip 6Place.
The power source 12 of the present invention, negative-pressure vacuum pump 1404, normally closed solenoid valve 1402 and position sensor 1304 respectively withController 10 is electrically connected.
Controller 10 so that executing agency 13 is slide block guide rail executing agency as an example, with embedded microprocessor as core(controller 10 of the present invention is prior art) controls whole automatic sampling apparatus 9.After controller 10 sends sample introduction to be instructed, gasCylinder A works, and band movable slider A1303 is moved on vertical guide rail 1301, and horizontal guide rail 1302 is with slide block A1303 synchronizing movings, waterThe slide block B 1306 that cylinder B1312 on level gauge 1302 drives moves on horizontal guide rail 1302 and (can first vertically move rear level to moveDynamic, or first move horizontally vertically move afterwards), be connected with air-breather 14 by gangbar 1305, by control executing agency13 horizontally or vertically motion makes air-breather 14 reach specified location, and the feedback stop motion signal of position sensor 1304 is to controlDevice processed 10, the control of controller 10 executing agency 13 stop movement, and make the vacuum cup 1405 on air-breather 14 just fallExport at 604, and outlet 604 is covered.The negative-pressure vacuum pump 1404 that controller 10 starts in air-breather 14 works, in negative pressureIn the presence of, vacuum cup 1405 tightly holds micro-fluidic chip 6, and starts to aspirate and make the sample in sample cell 603 from sample introductionMouth 602 enters into PCR microreactors 601, and is detected in the position of PCR microreactors 601；When sample is completely filled with PCRMicroreactor 601, the detection sensor in 601 region of PCR microreactors detect sample introduction end signal and feed back to controller 10 and carryShow that sample introduction is completed, controller 10 sends instruction stopping negative-pressure vacuum pump 1404 and works, and opens the normally closed electromagnetism of air-breather 14Valve 1402 admits air into release vacuum cup 1405 in threeway 1403, makes vacuum cup 1405 leave micro-fluidic chip 6.WithAfterwards, the control of controller 10 executing agency 13 drives air-breather 14 to leave outlet 604, returns to original position, completes single injected samplingProcess.
Temperature cycles heating system 7, attemperating unit is by certain heat exchange device, including but not limited to electric heating device(such as semiconductor chilling plate, resistance wire, electroplating film), pharoid (such as infrared heater), media heat exchanger (medium include butIt is not limited to the refrigerating mediums such as water, air, oil, ethylene glycol, ethanol) etc., circulating-heating is realized, a temperature cycles process includes threeTemperature section：1) denaturation stage continues t1Second, 2) annealing stage continues t2Second, 3) extend stage lasting t3Second, each phase temperature value andDuration can be arranged on control computer software or instrument control panel.Required according to the detection of actual sample, temperature controlSystem may also be configured to a thermostat temperature section, or the temperature section of only two alternating temperatures.
Electric loading system platform 8, for carrying micro-fluidic chip 6 and temperature cycles heater 7, can be two dimension, three-dimensional,Multidimensional or rotary motion platform, drive micro-fluidic chip 6 horizontally or vertically to be moved along a straight line, or move for arbitrary curve, such asThis can realize image collecting device timesharing multi-region IMAQ.
Controller 10 is the core component of foranalysis of nucleic acids system, and it controls foranalysis of nucleic acids system according to set program workMake.Controller 10 mainly includes that microprocessor, communication module, light source driver module, circulating-heating drive module, electric platforms driveDynamic model block, vavuum pump drive module, sample introduction Locating driver module, as shown in Figure 4.Microprocessor is the core of platform controllerPart, is responsible for peripheral interface circuit, circulating-heating drive module, motion platform drive module, light source driver module, vavuum pump and drivesModule, sample introduction Locating driver module, the control of communication module and communication work.Communication module is by certain media and calculatingMachine 11 interacts communication, and communication mode includes but is not limited to wire communication, radio communication, infrared communication etc., media bagInclude but be not limited to air, optical fiber, cable, electromagnetic wave, infrared ray etc..Light source driver module drives excitation source 5 to send light source, passes throughDichroscope 4 is irradiated to 601 region of PCR microreactors of micro-fluidic chip 6 after reflecting, excite positive template to send fluorescence.NegativePressure vavuum pump drive module, control drive vavuum pump action, are that automatic sampling apparatus produce negative pressure driving.Sample introduction Locating driver mouldBlock, the vacuum cup 1405 in precise control automatic sampling apparatus 9 are docked with outlet 604, realize sample introduction.Circulating-heating drives mouldBlock, the hydronic each section of temperature of precise control, the heat time for controlling each temperature section and cycle-index.Motion platform drives mouldBlock, linear motion of the precise control electric loading system platform 8 in level, vertical direction, or turn clockwise or counterclockwiseDynamic.
The Liquid particle image for collecting is filtered by computer 11, rim detection, histogram calculation, determine detection limit, twoValue processes, calculates all amount of droplets under light field, calculates fluorescence amount of droplets, the absolute template number of analytical calculation and detection off fieldSample concentration etc..Foranalysis of nucleic acids systematic functional structrue block diagram is as shown in Figure 5.