CN104046614A - Cell lysis buffer and preparation process thereof - Google Patents
Cell lysis buffer and preparation process thereof Download PDFInfo
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- CN104046614A CN104046614A CN201410274589.4A CN201410274589A CN104046614A CN 104046614 A CN104046614 A CN 104046614A CN 201410274589 A CN201410274589 A CN 201410274589A CN 104046614 A CN104046614 A CN 104046614A
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- proteinase
- hepes
- cell lysis
- proclin300
- dodecyl sulfate
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Abstract
The invention provides a cell lysis buffer and a preparation process thereof. The cell lysis buffer is prepared from Hepes sodium salt, Hepes free acid, lithium chloride, ethylene diamine tetra acetic acid, lithium dodecyl sulfate, Proclin300 and ProteinaseK. The cell lysis process adopting the cell lysis buffer provided by the invention is simple and convenient in operation and saves time and labor, and after cell lysis, cervical epithelial cast-off cell samples can be directly used in an nucleic acid test without need of DNA/RNApurification.
Description
Technical field
The present invention relates to biologic product technology field; Be specifically related to a kind of cell pyrolysis liquid and preparation technology thereof.
Background technology
At present, genome extracts and a committed step of protein research is exactly lysis, its lysis mode difference of different samples, also difference to some extent of the selection of lysate.Common lysis method CTAB method, physical method, SDS method etc. again at present.CTAB sends out and is applicable to plant tissue, the isocellular cracking of fungi.Physical method due to genomic destructive force compared with advising greatly and not adopting this method.SDS method is suitable for blood, cell, animal tissues, bacterium, yeast etc., is a kind of powerful lysis mode.In experimenter's actual mechanical process, need strictly will source and extraction per sample require to select suitable lysis method.Therefore the ubiquitous problem of this several method is exactly: 1, pyrolysis time is long; The problem such as 2, lytic effect is insufficient.
Therefore, the present invention develops on the basis that mainly solves more than currently available technology existence deficiency.
Summary of the invention
The object of invention is to solve above technical problem, and the special one that proposes reduces pyrolysis time, the cell pyrolysis liquid that cracking is abundant, simple in structure, with low cost and preparation technology thereof.
Based on above-mentioned purpose, technical problem solved by the invention realizes by the following technical solutions:
A kind of cell pyrolysis liquid and preparation technology thereof, it is made up of Hepes sodium salt, Hepes free acid, lithium chloride, ethylenediamine tetraacetic acid (EDTA), lithium dodecyl sulfate, Proclin300, Proteinase K, by weight percentage, wherein
A.Hepes sodium salt: 13-25%;
B.Hepes free acid: 1.92-12%;
C. lithium chloride: 40.88-62%;
D. ethylenediamine tetraacetic acid (EDTA): 2.5-5%;
E. lithium dodecyl sulfate: 56-65%;
f.Proclin300：0.2-2.5％；
G. Proteinase K: 1%.
Further, described following weight ratio is:
A.Hepes sodium salt: 13mg/ml;
B.Hepes free acid: 11.92mg/ml;
C. lithium chloride: 50.88mg/ml;
D. ethylenediamine tetraacetic acid (EDTA): 3mg/ml;
E. lithium dodecyl sulfate: 60mg/ml;
f.Proclin300：0.2ul/m；
G. Proteinase K: 1mg/ml.
Further, described the said products weight ratio is according to the following steps:
A. sample is transferred to centrifuge tube, centrifugal 5 minutes of 3000rpm, abandons supernatant;
B. add resuspended the mixing of 2ml cracking diluent, centrifugal 5 minutes of 3000rpm, abandons supernatant;
C. add 300ul lysate, then add 3ul Proteinase K, resuspended mixing;
D. place 65 DEG C of cracking of thermostat container, about 20 minutes concussion once, after 1 hour take out, sample is returned to room temperature, get supernatant liquor for detection of.
Further, the concentration of described Proteinase K is 20mg/ml.
The know-why of component of the present invention:
Hepes: be a kind of nonionic both sexes damping fluid.
Lithium chloride: as a kind of RNA precipitation agent.
Ethylenediamine tetraacetic acid (EDTA): denaturing agent, destroys cytolemma and carries out lysing cell.
Lithium dodecyl sulfate: anion surfactant.Can be used for RNA, DNA segregation, protein solubilizing agent.
Proclin300: sanitas.
Proteinase K: the activity of protease inhibition, for the degraded of protein.
The invention has the beneficial effects as follows: 1. this lysate lysing cell process, simple, convenient, time saving and energy saving.2. the epithelium of cervix uteri cast-off cells sample after cracking can directly carry out detection of nucleic acids, without DNA/RNA purifying.3. cracking process gentleness, effectively reduces the damage of cracking process amplifying nucleic acid.4. with low cost, the research that is suitable for exuviation cell in routine test.
Embodiment
Below by specific embodiment, further set forth the present invention.
A kind of cell pyrolysis liquid and preparation technology thereof, it is made up of Hepes sodium salt, Hepes free acid, lithium chloride, ethylenediamine tetraacetic acid (EDTA), lithium dodecyl sulfate, Proclin300, Proteinase K, by weight percentage, wherein
A.Hepes sodium salt: 13-25%;
B.Hepes free acid: 1.92-12%;
C. lithium chloride: 40.88-62%;
D. ethylenediamine tetraacetic acid (EDTA): 2.5-5%;
E. lithium dodecyl sulfate: 56-65%;
f.Proclin300：0.2-2.5％；
G. Proteinase K: 1%.
Embodiment 1, cell pyrolysis liquid preparation formula of the present invention is,
A.Hepes sodium salt: 13mg/ml;
B.Hepes free acid: 11.92mg/ml;
C. lithium chloride: 50.88mg/ml;
D. ethylenediamine tetraacetic acid (EDTA): 3mg/ml;
E. lithium dodecyl sulfate: 60mg/ml;
f.Proclin300：0.2ul/m；
G. Proteinase K: 1mg/ml.
Take from the said products weight ratio prepared according to the following steps:
A. sample is transferred to centrifuge tube, centrifugal 5 minutes of 3000rpm, abandons supernatant;
B. add resuspended the mixing of 2ml cracking diluent, centrifugal 5 minutes of 3000rpm, abandons supernatant;
C. add 300ul lysate, then add 3ul Proteinase K, resuspended mixing;
D. place 65 DEG C of cracking of thermostat container, about 20 minutes concussion once, after 1 hour take out, sample is returned to room temperature, get supernatant liquor for detection of.
Embodiment 2, cell pyrolysis liquid preparation formula of the present invention is,
A.Hepes sodium salt: 25mg/ml;
B.Hepes free acid: 1.92mg/ml;
C. lithium chloride: 62mg/ml;
D. ethylenediamine tetraacetic acid (EDTA): 5mg/ml;
E. lithium dodecyl sulfate: 56mg/ml;
f.Proclin300：2.5ul/m；
Preparation according to the method described above.
The foregoing is only preferred embodiment of the present invention, do not limit Application Areas of the present invention.All any amendments of making, be equal to replacement and improvement etc., within protection scope of the present invention all should be included within the present invention spirit and principle.
Claims (4)
1. cell pyrolysis liquid and a preparation technology thereof, is characterized in that, it is made up of Hepes sodium salt, Hepes free acid, lithium chloride, ethylenediamine tetraacetic acid (EDTA), lithium dodecyl sulfate, Proclin300, Proteinase K, by weight percentage, and wherein
A.Hepes sodium salt: 13-25%;
B.Hepes free acid: 1.92-12%;
C. lithium chloride: 40.88-62%;
D. ethylenediamine tetraacetic acid (EDTA): 2.5-5%;
E. lithium dodecyl sulfate: 56-65%;
f.Proclin300：0.2-2.5％；
G. Proteinase K: 1%.
2. cell pyrolysis liquid according to claim 1, is characterized in that, described following weight ratio is:
A.Hepes sodium salt: 13mg/ml;
B.Hepes free acid: 11.92mg/ml;
C. lithium chloride: 50.88mg/ml;
D. ethylenediamine tetraacetic acid (EDTA): 3mg/ml;
E. lithium dodecyl sulfate: 60mg/ml;
f.Proclin300：0.2ul/m；
G. Proteinase K: 1mg/ml.
3. lysis liquid preparing process according to claim 1, is characterized in that, described the said products weight ratio according to the following steps:
A. sample is transferred to centrifuge tube, centrifugal 5 minutes of 3000rpm, abandons supernatant;
B. add resuspended the mixing of 2ml cracking diluent, centrifugal 5 minutes of 3000rpm, abandons supernatant;
C. add 300ul lysate, then add 3ul Proteinase K, resuspended mixing;
D. place 65 DEG C of cracking of thermostat container, about 20 minutes concussion once, after 1 hour take out, sample is returned to room temperature, get supernatant liquor for detection of.
4. cell pyrolysis liquid according to claim 1 and preparation technology thereof, is characterized in that, the concentration of described Proteinase K is 20mg/ml.
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| CN201410274589.4A CN104046614A (en) | 2014-06-19 | 2014-06-19 | Cell lysis buffer and preparation process thereof |
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Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN108427000A (en) * | 2017-02-15 | 2018-08-21 | 广州市锐博生物科技有限公司 | A kind of method and kit of capture nucleic acid binding protein |
| CN109762869A (en) * | 2019-02-11 | 2019-05-17 | 张丽英 | A method of Salmonella in Food is detected using ATP bioluminescence reaction |
| CN114107440A (en) * | 2021-11-25 | 2022-03-01 | 和元生物技术（上海）股份有限公司 | Embryo lysate and application thereof |
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| CN101935704A (en) * | 2010-08-27 | 2011-01-05 | 中国科学院广州生物医药与健康研究院 | Liquid-phase chip detection system based on biological bar code |
| CN102007212A (en) * | 2008-03-07 | 2011-04-06 | 国立大学法人富山大学 | Homologous recombination method, cloning method, and kit |
| CN102234682A (en) * | 2010-04-20 | 2011-11-09 | 中国科学院广州生物医药与健康研究院 | Nucleic acid nano-gold biosensor based on bio-barcode |
| WO2013188872A1 (en) * | 2012-06-15 | 2013-12-19 | The Board Of Regents Of The University Of Texas System | High throughput sequencing of multiple transcripts of a single cell |
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2014
- 2014-06-19 CN CN201410274589.4A patent/CN104046614A/en active Pending
Patent Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102007212A (en) * | 2008-03-07 | 2011-04-06 | 国立大学法人富山大学 | Homologous recombination method, cloning method, and kit |
| CN102234682A (en) * | 2010-04-20 | 2011-11-09 | 中国科学院广州生物医药与健康研究院 | Nucleic acid nano-gold biosensor based on bio-barcode |
| CN101935704A (en) * | 2010-08-27 | 2011-01-05 | 中国科学院广州生物医药与健康研究院 | Liquid-phase chip detection system based on biological bar code |
| WO2013188872A1 (en) * | 2012-06-15 | 2013-12-19 | The Board Of Regents Of The University Of Texas System | High throughput sequencing of multiple transcripts of a single cell |
Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN108427000A (en) * | 2017-02-15 | 2018-08-21 | 广州市锐博生物科技有限公司 | A kind of method and kit of capture nucleic acid binding protein |
| CN108427000B (en) * | 2017-02-15 | 2021-06-08 | 广州市锐博生物科技有限公司 | Method and kit for capturing nucleic acid binding protein |
| CN109762869A (en) * | 2019-02-11 | 2019-05-17 | 张丽英 | A method of Salmonella in Food is detected using ATP bioluminescence reaction |
| CN114107440A (en) * | 2021-11-25 | 2022-03-01 | 和元生物技术（上海）股份有限公司 | Embryo lysate and application thereof |
| CN114107440B (en) * | 2021-11-25 | 2024-05-28 | 和元生物技术（上海）股份有限公司 | Embryo lysate and application thereof |
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