CN107858402A - A kind of murine genes type authentication method - Google Patents
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- CN107858402A CN107858402A CN201711177866.XA CN201711177866A CN107858402A CN 107858402 A CN107858402 A CN 107858402A CN 201711177866 A CN201711177866 A CN 201711177866A CN 107858402 A CN107858402 A CN 107858402A
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Abstract
Include handling karyocyte using 0.2%~0.4% phosphate solutions of Triton X 100 the invention provides a kind of murine genes type authentication method so that the process that intracellular nucleic acid discharges.The present invention is it has been investigated that 0.2%~0.4% phosphate solutions of Triton X 100 handle karyocyte, intracellular nucleic acid can be made to be discharged into the aqueous solution, it is further purified without using isopropanol or ethanol, pyrolysis product directly can be used for PCR amplifications, greatly reduces the step of extracting DNA, shortens the time for obtaining DNA, the efficiency for improving experiment.
Description
Technical field
The present invention relates to the technical field of genetic engineering, in particular to a kind of murine genes type authentication method.
Background technology
The mouse (also known as transgenic mice) of foreign gene-carrying is the important tool of human diseases research, in medical science and life
The application for ordering field of scientific study is more and more extensive.Before application transgenic mice carries out scientific research, it usually needs to mouse
Genotype identified, distinguish transgenic mice and wild-type mice.Murine genes identify, be related to mouse DNA acquisition,
Target DNA fragments amplification, DNA electrophoresis.
DNA extraction method traditional at present includes phenol/chloroform extraction and high salt extraction method.Although these methods can obtain
Higher degree and the tissue DNA compared with high yield pulp1 are taken, but has some shortcomings part：First, operating procedure is comparatively laborious, for example, it is more
It is secondary to be related to centrifugation；Second, being related to excessive reagent, such as employ the reagents such as chloroform, ethanol, Proteinase K；Third, the time is inclined
Long, part Experiment step includes flow overnight.Method disclosed in pertinent literature even in recent years and available reagent box (such as
Vazyme, Promega and Takala company kit etc.) provide method, its DNA extraction part takes time also in 30min
To between a few hours, and contain consumptive material (such as collection post) and plurality of reagents (such as lysate, neutralizer, Proteinase K).
Above mentioned problem all adds experimental cost (including reagent cost, consumables cost and human cost) to a certain extent, so as to make
The about operating efficiency of transgenic mice Genotyping.
The content of the invention
In view of this, a kind of the defects of the technical problem to be solved in the present invention is to overcome prior art, there is provided mouse
Genotype identification method, including karyocyte is handled using 0.2%~0.4% Triton X-100 phosphate solutions so that
The process that intracellular nucleic acid discharges.
In some embodiments, the karyocyte picks up from mouse tail vein blood.
In some embodiments, the collocation method of described 0.2%~0.4%% Triton X-100 phosphate solutions
Including：
0.2~0.4mL Triton X-100 are dissolved in 100mL phosphate buffers, are made into 0.2%~0.4%
Triton X-100 phosphate solutions.
In some embodiments, it is further comprising the steps of：
S1：Take mouse tail vein blood；
S2：Erythrocyte cracked liquid is added into tail vein, treats erythroclasis, centrifugation removes red blood cell, obtains white thin
Born of the same parents are precipitated；
S3：Wash leukocyte cell pellet；
S4：The Triton X-100 phosphate solutions that content is 0.2%~0.4%, which are added in leukocyte cell pellet to handle, to be had
Nucleus so that intracellular nucleic acid discharges；
S5：PCR reacts；
S6：Aobvious band is observed in gel imager.
In some embodiments, the S1 is further comprising the steps of：
S11：Fixed mouse, mousetail is wiped with cotton ball soaked in alcohol, expands caudal vein；
S12：The tail vein of rat-tail distal dilation is poked with syringe needle, 5~10ul venous blood is drawn with siphon tubule；
S13：The venous blood of acquisition is moved in the sterile EP pipes of 1.5mL of the solution of ETDA containing 10uL, then with sterile dry cotton ball
By pressing mouth.
In some embodiments, the S2 is further comprising the steps of：
S21：The erythrocyte cracked liquid of 0.5mL precoolings is added into EP pipes, is mixed static 4~5 minutes on ice, is treated red thin
Born of the same parents are broken complete；
S22：1000r/min is centrifuged 5 minutes, is abandoned supernatant and is removed red blood cell, obtains leukocyte cell pellet.
In some embodiments, the S3 is further comprising the steps of：
S31：Washed 2 times with PBS；
S32：1000r/min is centrifuged 2 minutes, abandons supernatant.
In some embodiments, the S4 is further comprising the steps of：
S41：Configure 0.2%~0.4% phosphate solutions of X-100 containing Triton；
S42：0.2%~0.4%Triton X-100 phosphate solutions the 0.2mL configured is added to EP pipes to be mixed
Compound, concussion are resuspended, are stored at room temperature 10 minutes；
S43：Mixture after will be static centrifuges 5 minutes in 1000r/min, and supernatant is moved into new sterile EP manages, in-
80 degrees Celsius of preservations, or place on ice, operated in next step with preparing row.
In some embodiments, PCR reaction solutions composition is in the S5：
In some embodiments, PCR reaction conditions are in the S5：
Step1：Pre-degeneration：94℃、5min、1cycle；
Step2：PCR reacts：94 DEG C, 30s, 58 DEG C, 30s, 72 DEG C, 30s, 35cycle；
Step3：Thoroughly extension：72℃、7min.
In terms of existing technologies, beneficial effect is a kind of murine genes type authentication method provided by the invention：
Murine genes type authentication method provided by the invention, including use 0.2%~0.4% Triton X-100 phosphoric acid
Salting liquid handles karyocyte so that the process that intracellular nucleic acid discharges.
It is understood that the method currently used for whole blood DNA extraction, special agent used is more, during whole operation
Between it is longer.The present invention except with use erythrocyte cracked liquid splitting erythrocyte as other schemes in addition to, to the karyocyte of acquisition,
Use the auxiliary reagent Triton X-100 (Triton X-100) in laboratory.
Triton X-100 typical concentrations are 0.025%-0.1% in immunocytochemistry, for rinsing tissue specimen,
Slice surface tension force is reduced, makes reaction solution be easy to cover sample surface.Also have and think 0.025%-0.1%Triton X-100
Phosphate solution can dissolve the Fc acceptors of cell membrane surface and reduce nonspecific combination, increase the penetrating of cell membrane
Property.
It is to be appreciated that the present invention is it has been investigated that 0.2%~0.4% Triton X-100 phosphate solutions are handled
Karyocyte, intracellular nucleic acid can be made to be discharged into the aqueous solution, be further purified without using isopropanol or ethanol, split
Solution product directly can be used for PCR amplifications, greatly reduces the step of extracting DNA, shortens acquisition DNA time, improves
The efficiency of experiment.
Being studied by many experiments confirms, nonionic surface active agent Triton X-100 have under specific concentration
Above-mentioned special effect.Especially, 0.2%~0.4% Triton X-100 phosphate solutions.
Further, the karyocyte picks up from mouse tail vein blood.
It is understood that the mode majority currently for the kit extraction mouse DNA of transgenic mice identification is logical
Clip rat-tail, mouse ear or mouse toe is crossed to realize.
Using the One step Mouse Genotyping Kit products of Vazyme companies as representative, there is provided a whole set of DNA is thick
Extraction and PCR amplification system, by adding Tissue lysates and Proteinase K, (the 55 C water bath 2- by way of water-bath
4 hours or water-bath are stayed overnight, 95 degrees Celsius of fire extinguishing Proteinase Ks), mouse tissue is discharged DNA, then directly draw cracking
Liquid carries out follow-up target DNA fragments amplification and DNA electrophoretic procedures as DNA profiling.
The DNAiso Reagent that Takala companies provide also can extract DNA (traditional windings in animal tissue source
Method), sample can be fully cleaved in DNAiso Reagent, after ethanol is added, it may appear that the genome of white flock
DNA precipitate, collect and wash precipitation can obtain genomic DNA, operation can be completed in 30 minutes.
It is also useful on the market although rarely having the report that murine genes type is identified in a manner of whole blood extracts DNA on document
In the examination kit from whole blood extraction DNA.Using the Blood Genome DNA Extraction Kit of Takala companies as generation
Table, kit contain tri- kinds of GenTLE Solution I, II, III solution.Wherein GenTLE Solution I effect be
It is globulicidal simultaneously, with nucleic acid formed electroneutral complex.The complex is collected by centrifugation and uses GenTLE
After Solution II cleanings, GenTLE Solution III are added in precipitation, DNA is separated, then added different
DNA is precipitated and reclaimed by propyl alcohol, and whole operation needs 40 minutes or so.
The TaKaRa MiniBEST Whole Blood Genomic DNA Extraction Kit of Takala companies are adopted
Use column method.The kit, which is used to add in erythroplastid (such as mammal) whole blood of various anti-coagulants from 1uL~1mL, to be carried
Genomic DNA is taken, a large amount of erythroplastids in whole blood are cracked using special erythrocyte splitting method, then by cell pyrolysis liquid
Cracking leucocyte simultaneously discharges genomic DNA, and membrane technology purified genomic dna is prepared in conjunction with DNA.
It follows that currently for transgenic mice identification it is tissue-derived be typically the rat-tail of clip, mouse ear or
Mouse toe, larger wound is caused to experiment mice, and causes larger stress reaction, and be not easy after clip tissue hemostasis or
Bleeding stopping period is longer.The present invention from mouse rat-tail by gathering venous blood, it is only necessary to gathers 5~10 microlitres of venous blood, mitigates to small
The wound and stress reaction of mouse, and convenient hemostasis.
In summary, a kind of murine genes type authentication method provided by the invention has the advantages of above-mentioned many and value,
And similar method is there are no in like product and publishes or uses and really belong to innovation, generate handy and practical effect
Fruit, more existing technology have the multinomial effect promoted, and so as to more be suitable to practicality, and have extensive industrial value.
Brief description of the drawings
It should be appreciated that the following drawings illustrate only certain embodiments of the present invention, therefore it is not construed as to model
The restriction enclosed, for those of ordinary skill in the art, on the premise of not paying creative work, can also be according to these
Accompanying drawing obtains other related accompanying drawings.
Fig. 1 is method disclosed in embodiment of the present invention to 3 alpha-synapse nucleoprotein A53T mutation transgenes mouse and 3
The wild-type mice of littermate carries out the result of identified for genes；
Fig. 2 is the One step Mouse Genotyping Kit of Vazyme companies method to 3 α-cynapse core egg
The wild-type mice of white A53T mutation transgenes mouse and 3 littermates carries out the result of identified for genes.
Wherein, "+" represents transgenic mice, and "-" represents wild-type mice.Agargel electrophoresis qualification result：It is mutated bar
Band is located at 500bp, and wild type occurs without band.Both sides swimming lane is DNA marker： 100bp、200bp、300bp、400bp、
500bp、700bp、1000bp。
Embodiment
Hereinafter, the present invention will be described more fully with conjunction with the embodiments.The disclosure can have various embodiments, and can
Adjust and change wherein.It should be understood, however, that：It is disclosed herein specific in the absence of the various embodiments of the disclosure are limited to
The intention of embodiment, but the disclosure should be interpreted as to covering the institute in the spirit and scope for the various embodiments for falling into the disclosure
There are adjustment, equivalent and/or alternative.
Hereinafter, disclosed in the term " comprising " that can be used in the various embodiments of the disclosure or " may include " instruction
Function, operation or the presence of element, and do not limit the increase of one or more functions, operation or element.
In the various embodiments of the disclosure, stating "or" or " at least one in A or/and B " includes what is listed file names with
Any combinations of word or all combinations.For example, " A or B " or " at least one in A or/and B " may include A, Ke Bao for statement
Include B or may include A and B both.
The statement (" first ", " second " etc.) used in the various embodiments of the disclosure can be modified in various implementations
Various element in example, but respective sets can not be limited into element.For example, presented above be not intended to limit the suitable of the element
Sequence and/or importance.The purpose presented above for being only used for differentiating an element and other elements.For example, the first user fills
Put and indicate different user device with second user device, although the two is all user's set.For example, each of the disclosure is not being departed from
In the case of the scope of kind embodiment, the first element is referred to alternatively as the second element, and similarly, the second element is also referred to as first
Element.
It should be noted that：, can be by the first composition member if an element ' attach ' to another element by description
Part is directly connected to the second element, and " connection " the 3rd can be formed between the first element and the second element
Element.On the contrary, when an element " being directly connected to " is arrived into another element, it will be appreciated that be in the first element
And second be not present the 3rd element between element.
The term used in the various embodiments of the disclosure is only used for describing the purpose of specific embodiment and not anticipated
In the various embodiments of the limitation disclosure.As used herein, singulative is intended to also include plural form, unless context is clear
Chu it is indicated otherwise.Unless otherwise defined, all terms (including the technical term and scientific terminology) tool being otherwise used herein
There is the implication identical implication that the various embodiment one skilled in the art with the disclosure are generally understood that.The term
(term such as limited in the dictionary typically used) is to be interpreted as having and the situational meaning in correlative technology field
Identical implication and the implication with Utopian implication or overly formal will be not construed as, unless in the various of the disclosure
It is clearly defined in embodiment.
In some embodiments of the invention, a kind of murine genes type authentication method, including using 0.2%~0.4%
Triton X-100 phosphate solutions handle karyocyte so that the process that intracellular nucleic acid discharges.
It is above-mentioned, it is to be understood that currently used for the method for whole blood DNA extraction, special agent used is more, whole behaviour
It is longer to make the time.The present invention except with as other schemes use erythrocyte cracked liquid splitting erythrocyte in addition to, have core to acquisition
Cell, use the auxiliary reagent Triton X-100 (Triton X-100) in laboratory.
Triton X-100 typical concentrations are 0.025%-0.1% in immunocytochemistry, for rinsing tissue specimen,
Slice surface tension force is reduced, makes reaction solution be easy to cover sample surface.Also have and think 0.025%-0.1%Triton X-100
Phosphate solution can dissolve the Fc acceptors of cell membrane surface and reduce nonspecific combination, increase the penetrating of cell membrane
Property.
It is to be appreciated that the present invention is it has been investigated that 0.2%~0.4% Triton X-100 phosphate solutions are handled
Karyocyte, intracellular nucleic acid can be made to be discharged into the aqueous solution, be further purified without using isopropanol or ethanol, split
Solution product directly can be used for PCR amplifications, greatly reduces the step of extracting DNA, shortens acquisition DNA time, improves
The efficiency of experiment.
Being studied by many experiments confirms, nonionic surface active agent Triton X-100 have under specific concentration
Above-mentioned special effect.Especially, 0.2%~0.4% Triton X-100 phosphate solutions.
In some embodiments of the present invention, the karyocyte picks up from mouse tail vein blood.
In some embodiments of the present invention, the configuration of described 0.2%~0.4% Triton X-100 phosphate solutions
Method includes：
0.2~0.4mL Triton X-100 are dissolved in 100mL phosphate buffers, are made into 0.2%~0.4%
Triton X-100 phosphate solutions.
It is further comprising the steps of in some embodiments of the present invention：
S1：Take mouse tail vein blood；
S2：Erythrocyte cracked liquid is added into tail vein, treats erythroclasis, centrifugation removes red blood cell, obtains white thin
Born of the same parents are precipitated；
S3：Wash leukocyte cell pellet；
S4：The Triton X-100 phosphate solutions that content is 0.2%~0.4%, which are added in leukocyte cell pellet to handle, to be had
Nucleus so that intracellular nucleic acid discharges；
S5：PCR reacts；
S6：Aobvious band is observed in gel imager.
In some embodiments of the present invention, the S1 is further comprising the steps of：
S11：Fixed mouse, mousetail is wiped with cotton ball soaked in alcohol, expands caudal vein；
S12：The tail vein of rat-tail distal dilation is poked with syringe needle, 5~10ul venous blood is drawn with siphon tubule；
S13：The venous blood of acquisition is moved in the sterile EP pipes of 1.5mL of the solution of ETDA containing 10uL, then with sterile dry cotton ball
By pressing mouth.
In some embodiments of the present invention, the S2 is further comprising the steps of：
S21：The erythrocyte cracked liquid of 0.5mL precoolings is added into EP pipes, is mixed static 4~5 minutes on ice, is treated red thin
Born of the same parents are broken complete；
S22：1000r/min is centrifuged 5 minutes, is abandoned supernatant and is removed red blood cell, obtains leukocyte cell pellet.
In some embodiments of the present invention, the S3 is further comprising the steps of：
S31：Washed 2 times with PBS；
S32：1000r/min is centrifuged 2 minutes, abandons supernatant.
In some embodiments of the present invention, the S4 is further comprising the steps of：
S41：Configure 0.2%~0.4% phosphate solutions of X-100 containing Triton；
S42：0.2%~0.4%Triton X-100 phosphate solutions the 0.2mL configured is added to EP pipes to be mixed
Compound, concussion are resuspended, are stored at room temperature 10 minutes；
S43：Mixture after will be static centrifuges 5 minutes in 1000r/min, and supernatant is moved into new sterile EP manages, in-
80 degrees Celsius of preservations, or place on ice, operated in next step with preparing row.
In some embodiments of the present invention, PCR reaction solutions composition is in the S5：
In some embodiments of the present invention, PCR reaction conditions are in the S5：
Step1：Pre-degeneration：94℃、5min、1cycle；
Step2：PCR reacts：94 DEG C, 30s, 58 DEG C, 30s, 72 DEG C, 30s, 35cycle；
Step3：Thoroughly extension：72℃、7min.
Following embodiment facilitates a better understanding of the present invention, but does not limit the present invention.
Embodiment 1
(1) fixed mouse, mousetail is wiped with cotton ball soaked in alcohol, expands caudal vein, it is remote to poke rat-tail with No. 4 syringe needles
Hold the tail vein of expansion, 5uL venous blood drawn with siphon tubule, with i.e. by blood move to the 1.5mL of the solution of ETDA containing 10ul without
Bacterium EP is managed, and pressing mouth is pressed with sterile dry cotton ball；
(2) toward the erythrocyte cracked liquid of addition 0.5mL precoolings in EP pipes, mix static 4 minutes on ice, treat that red blood cell is complete
Broken, 1000r/min is centrifuged 5 minutes, is abandoned supernatant, is obtained leukocyte cell pellet, is washed 2 times with PBS, and 1000r/min centrifuges 2 points
Clock, abandon supernatant.
(3) 0.2% phosphate solutions of X-100 containing Triton are configured, add 0.2mL toward EP pipes, concussion is resuspended, and room temperature is quiet
Put 10 minutes, 1000r/min is centrifuged 5 minutes, and supernatant is moved into new sterile EP manages, in -80 degrees Celsius of preservations, or on ice
Place, operated in next step with preparing row.
(4) DNA profiling of extraction is used for condition when PCR reacts
1) PCR reaction solutions form
2) PCR reaction conditions
Step1：Pre-degeneration：94℃、5min、1cycle；
Step2：PCR reacts：94 DEG C, 30s, 58 DEG C, 30s, 72 DEG C, 30s, 35cycle；
Step3：Thoroughly extension：72℃、7min
(5) agargel electrophoresis, 150V, 25min, observed in gel imager.
Embodiment 2
(1) fixed mouse, mousetail is wiped with cotton ball soaked in alcohol, expands caudal vein, it is remote to poke rat-tail with No. 3 syringe needles
The tail vein of expansion is held, 10uL venous blood is drawn with siphon tubule, with the 1.5mL that blood is moved to the solution of ETDA containing 10uL
Sterile EP pipes, pressing mouth is pressed with sterile dry cotton ball.
(2) toward the erythrocyte cracked liquid of addition 0.5mL precoolings in EP pipes, mix static 5 minutes on ice, treat that red blood cell is complete
Broken, 1000r/min is centrifuged 5 minutes, is abandoned supernatant, is obtained leukocyte cell pellet, is washed 2 times with PBS, and 1000r/min centrifuges 2 points
Clock, abandon supernatant.
(3) 0.4% phosphate solutions of X-100 containing Triton are configured, add 0.2mL toward EP pipes, concussion is resuspended, and room temperature is quiet
Put 10 minutes, 1000r/min is centrifuged 5 minutes, and supernatant is moved into new sterile EP manages, in -80 degrees Celsius of preservations, or on ice
Place, operated in next step with preparing row.
(4) DNA profiling of extraction is used for condition when PCR reacts
1) PCR reaction solutions form
2) PCR reaction conditions
Step1：Pre-degeneration：94℃、5min、1cycle；
Step2：PCR reacts：94 DEG C, 30s, 58 DEG C, 30s, 72 DEG C, 30s, 35cycle；
Step3：Thoroughly extension：72℃、7min
(5) agargel electrophoresis, 150V, 25min, observed in gel imager.
It should be appreciated that although the present specification is described in terms of embodiments, not each embodiment only includes one
Individual independent technical scheme, this narrating mode of specification is only that those skilled in the art will should say for clarity
For bright book as an entirety, the technical scheme in each embodiment may also be suitably combined to form those skilled in the art can
With the other embodiment of understanding.
Inventor states that the present invention can only for of the invention by a series of describe in detail of those listed above
Row embodiment illustrates, but the invention is not limited in above-mentioned detailed process equipment and technological process.And i.e. not
Mean that the present invention should rely on above-mentioned detailed process equipment and technological process and could implement.Person of ordinary skill in the field should
This is clear, any improvement in the present invention, and the equivalence replacement and auxiliary element to each raw material of product of the present invention are added, are specific square
Selection of formula etc., within the scope of all falling within protection scope of the present invention and disclosing.
Claims (10)
- A kind of 1. murine genes type authentication method, it is characterised in that：Including the Triton X-100 phosphorus using 0.2%~0.4% Acid salt solution handles karyocyte so that the process that intracellular nucleic acid discharges.
- A kind of 2. murine genes type authentication method as claimed in claim 1, it is characterised in that：The karyocyte picks up from mouse Tail vein.
- A kind of 3. murine genes type authentication method as claimed in claim 2, it is characterised in that：Described 0.2%~0.4% The collocation method of Triton X-100 phosphate solutions includes：0.2~0.4mL Triton X-100 are dissolved in 100mL phosphate buffers, are made into 0.2%~0.4% Triton X-100 phosphate solutions.
- A kind of 4. murine genes type authentication method as claimed in claim 1, it is characterised in that：It is further comprising the steps of：S1：Take mouse tail vein blood；S2：Erythrocyte cracked liquid is added into tail vein, treats erythroclasis, centrifugation removes red blood cell, obtains leucocyte and sink Form sediment；S3：Wash leukocyte cell pellet；S4：The Triton X-100 phosphate solutions that content is 0.2%~0.4% are added into processing in leukocyte cell pellet has core thin Born of the same parents so that intracellular nucleic acid discharges；S5：PCR reacts；S6：Aobvious band is observed in gel imager.
- A kind of 5. murine genes type authentication method as claimed in claim 4, it is characterised in that：The S1 also includes following step Suddenly：S11：Fixed mouse, mousetail is wiped with cotton ball soaked in alcohol, expands caudal vein；S12：The tail vein of rat-tail distal dilation is poked with syringe needle, 5~10ul venous blood is drawn with siphon tubule；S13：The venous blood of acquisition is moved in the sterile EP pipes of 1.5mL of the solution of ETDA containing 10uL, then pressed with sterile dry cotton ball Pin mouth.
- A kind of 6. murine genes type authentication method as claimed in claim 4, it is characterised in that：The S2 also includes following step Suddenly：S21：The erythrocyte cracked liquid of 0.5mL precoolings is added into EP pipes, mixes static 4~5 minutes on ice, treats erythroclasis Completely；S22：1000r/min is centrifuged 5 minutes, is abandoned supernatant and is removed red blood cell, obtains leukocyte cell pellet.
- A kind of 7. murine genes type authentication method as claimed in claim 4, it is characterised in that：The S3 also includes following step Suddenly：S31：Washed 2 times with PBS；S32：1000r/min is centrifuged 2 minutes, abandons supernatant.
- A kind of 8. murine genes type authentication method as claimed in claim 4, it is characterised in that：The S4 also includes following step Suddenly：S41：Configure 0.2%~0.4% phosphate solutions of X-100 containing Triton；S42：0.2%~0.4%Triton X-100 phosphate solutions the 0.2mL configured is added to EP pipes and obtains mixture, Concussion is resuspended, and is stored at room temperature 10 minutes；S43：Mixture after will be static centrifuges 5 minutes in 1000r/min, and supernatant is moved into new sterile EP manages, and is taken the photograph in -80 Family name's degree preserves, or places on ice, is operated in next step with preparing row.
- A kind of 9. murine genes type authentication method as claimed in claim 4, it is characterised in that：PCR reaction solutions group in the S5 Turn into：
- A kind of 10. murine genes type authentication method as claimed in claim 4, it is characterised in that：PCR reaction conditions in the S5 For：Step1：Pre-degeneration：94℃、5min、1cycle；Step2：PCR reacts：94 DEG C, 30s, 58 DEG C, 30s, 72 DEG C, 30s, 35cycle；Step3：Thoroughly extension：72℃、7min.
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| CN110904241A (en) * | 2018-09-14 | 2020-03-24 | 广州铂晋生物科技有限公司 | Rapid identification method for transgenic mouse genotype |
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| US20060240449A1 (en) * | 2005-01-19 | 2006-10-26 | Mcglennen Ronald C | Methods and compositions for preparation of biological samples |
| CN101122601A (en) * | 2007-09-24 | 2008-02-13 | 孙艳萍 | Method for separating and authenticating erythroblast of blood |
| CN102914657A (en) * | 2012-10-10 | 2013-02-06 | 中国人民解放军军事医学科学院放射与辐射医学研究所 | Method for detecting protein in peripheral blood karyocyte cells |
| CN105734051A (en) * | 2016-03-18 | 2016-07-06 | 中国人民解放军第三军医大学第三附属医院 | Genotyping-based kit and method for extracting DNA of transgenic mice |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN110904241A (en) * | 2018-09-14 | 2020-03-24 | 广州铂晋生物科技有限公司 | Rapid identification method for transgenic mouse genotype |
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