CN102206628A - Extraction method of genome DNA of isolated soy proteins - Google Patents
Extraction method of genome DNA of isolated soy proteins Download PDFInfo
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- CN102206628A CN102206628A CN 201110081615 CN201110081615A CN102206628A CN 102206628 A CN102206628 A CN 102206628A CN 201110081615 CN201110081615 CN 201110081615 CN 201110081615 A CN201110081615 A CN 201110081615A CN 102206628 A CN102206628 A CN 102206628A
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Abstract
The invention discloses an extraction method of the genome DNA of isolated soy proteins, comprising the following steps of: firstly sufficiently mixing isolated soy proteins and a TE buffer solution to prepare a TE mixed solution; then adding a guanidinium isothiocyanate lysis solution with volume twice larger than that of the TE mixed solution, sufficiently uniformly mixing, and lysing at room temperature; adding isometric phenol and chloroform/isoamylol to extract proteins; centrifugalizing, and then adding isometric chloroform/isoamylol to a supernate to extract the proteins; centrifugalizing, and then adding isometric chloroform to the supernate to extract the proteins; centrifugalizing, and precipitating the supernate by using isopropanol; centrifugalizing, and washing by using ethanol for precipitation; airing at the room temperature; and dissolving in the TE buffer solution so as to obtain the TE solution of the genome DNA. According to the extraction method, the genome DNA which has high quality and is suitable for PCR (Polymerase Chain Reaction) detection is extracted from the isolated soy proteins, the concentration of the genome DNA is 1558 micrograms/ml, and the value of OD260/OD280 is 1.666; and in addition, the invention has the advantages of easy operation and short consuming time and is beneficial to fast detection.
Description
Technical field
The invention belongs to biological technical field, is a kind of extracting method of genomic dna of the soybean protein isolate that is applicable to pcr amplification.
Background technology
At present, about existing many researchs of the extraction of plant complete genome DNA and report, but because the component of the different tissues material of different plants or same kind of plant is different, the extracting method that is suited is also different.Soybean separation protein albumin content height is unfavorable for stripping and the extraction of DNA.General plant genome DNA extracting method can't extract genomic dna from soybean protein isolate, for next step the biological detection based on gene is brought difficulty.Existing soybean gene group DNA extraction method be to be that material extracts with soybean seedling blade, soybean dry seeds, a bright beanpod of drum grain phase, the said extracted method can't be extracted genomic dna from the soybean protein isolate of rich in proteins.
Summary of the invention
The objective of the invention is for what solve existing soybean gene group DNA extraction method is to be that material extracts with soybean seedling blade, soybean dry seeds, a bright beanpod of drum grain phase, the said extracted method can't be extracted the problem of genomic dna from the soybean protein isolate of rich in proteins, and then the extracting method of a kind of soybean protein isolate base because of group DNA is provided.
The present invention solves the scheme that its technical problem adopts: at first soybean protein isolate and TE damping fluid thorough mixing are made the TE mixed solution, add the long-pending guanidinium isothiocyanate lysate of mixed solution diploid then, abundant mixing, room temperature cracking.Add isopyknic phenol, chloroform/primary isoamyl alcohol extracting albumen, add isopyknic chloroform/primary isoamyl alcohol extracting albumen in the supernatant liquor of centrifugal back, centrifugal back supernatant liquor adds isopyknic chloroform extracting albumen again, centrifugal, the supernatant liquor isopropanol precipitating, centrifugal, the precipitation washing with alcohol, room temperature is dried, and is dissolved in the TE solution that gets final product genomic dna in the TE damping fluid.
The invention has the beneficial effects as follows: can from soybean protein isolate, extract the genomic dna that the high-quality PCR of being suitable for detects, weak point simple to operate, consuming time, be beneficial to rapid detection.The concentration of the DNA that obtains with method of the present invention is 1558 μ g/ml, the OD of DNA
260/ OD
280Value be 1.666.
Embodiment
Preferred implementation of the present invention is: at first soybean protein isolate and TE damping fluid thorough mixing are made the TE mixed solution, add the long-pending guanidinium isothiocyanate lysate of mixed solution diploid then, abundant mixing, room temperature cracking.Add isopyknic phenol, chloroform/primary isoamyl alcohol extracting albumen, add isopyknic chloroform/primary isoamyl alcohol extracting albumen in the supernatant liquor of centrifugal back, centrifugal back supernatant liquor adds isopyknic chloroform extracting albumen again, centrifugal, the supernatant liquor isopropanol precipitating, centrifugal, the precipitation washing with alcohol, room temperature is dried, and is dissolved in the TE solution that gets final product genomic dna in the TE damping fluid.
Soybean protein isolate is 30～80mg in the described TE mixed solution, and the TE damping fluid is 120～320 μ L.Preferable scope is soybean protein isolate 50mg, TE damping fluid 200 μ L.
Described guanidinium isothiocyanate lysate is 50～100mM pH, 6.0～8.0Tris-HCl wherein; 10～100mM pH 8.0EDTA; The 5M guanidinium isothiocyanate; 1.3%Triton X-100.Preferable scope 50mM pH 6.5Tris-HCl; 20mMpH 8.0EDTA; The 5M guanidinium isothiocyanate; 1.3%Triton X-100.
The described room temperature cracking time is 20～60min, and the preferable cracking time is 30min.
The ratio of described phenol, chloroform/primary isoamyl alcohol is 25: 24: 1.
The ratio of described chloroform/primary isoamyl alcohol is 24: 1.
The sedimentary alcohol concn of described washing DNA is 70～95%, and the alcohol concn of preferable washing DNA is 70%.
The precipitation temperature of described Virahol is-20～30 ℃, and preferable precipitation temperature is-20 ℃.
The sedimentation time of described Virahol is 20～60min.
The described room temperature air dried time is 10～15min.
Claims (10)
1. a soybean protein isolate base is because of organizing the extracting method of DNA, it is characterized in that, at first soybean protein isolate and TE damping fluid thorough mixing are made the TE mixed solution, add the long-pending guanidinium isothiocyanate lysate of mixed solution diploid then, abundant mixing, the room temperature cracking, add isopyknic phenol, chloroform/primary isoamyl alcohol extracting albumen adds isopyknic chloroform/primary isoamyl alcohol extracting albumen in the supernatant liquor of centrifugal back, and centrifugal back supernatant liquor adds isopyknic chloroform extracting albumen again, centrifugal, the supernatant liquor isopropanol precipitating, centrifugal, the precipitation washing with alcohol, room temperature is dried, and is dissolved in the TE solution that gets final product genomic dna in the TE damping fluid.
2. a kind of soybean protein isolate base according to claim 1 is characterized in that because of the extracting method of group DNA soybean protein isolate is 30～80mg in the described TE mixed solution, and the TE damping fluid is 120～320 μ L.
3. a kind of soybean protein isolate base according to claim 1 is characterized in that because of the extracting method of group DNA described guanidinium isothiocyanate lysate is 50～100mM pH, 6.0～8.0Tris-HCl wherein; 10～100mM pH 8.0EDTA; The 5M guanidinium isothiocyanate; 1.3%Triton X-100.
4. a kind of soybean protein isolate base according to claim 1 is characterized in that because of the extracting method of group DNA the described room temperature cracking time is 20～60min.
5. a kind of soybean protein isolate base according to claim 1 is characterized in that because of the extracting method of group DNA the ratio of described phenol, chloroform/primary isoamyl alcohol is 25: 24: 1.
6. a kind of soybean protein isolate base according to claim 1 is characterized in that because of the extracting method of group DNA the ratio of described chloroform/primary isoamyl alcohol is 24: 1.
7. a kind of soybean protein isolate base according to claim 1 is characterized in that because of the extracting method of group DNA the sedimentary alcohol concn of described washing DNA is 70～95%.
8. a kind of soybean protein isolate base according to claim 1 is characterized in that because of the extracting method of group DNA the precipitation temperature of described Virahol is-20～30 ℃.
9. a kind of soybean protein isolate base according to claim 1 is characterized in that because of the extracting method of group DNA the sedimentation time of described Virahol is 20～60min.
10. a kind of soybean protein isolate base according to claim 1 is characterized in that because of the extracting method of group DNA the described room temperature air dried time is 10～15min.
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| CN 201110081615 CN102206628B (en) | 2011-04-01 | 2011-04-01 | Extraction method of genome DNA of isolated soy proteins |
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| CN 201110081615 CN102206628B (en) | 2011-04-01 | 2011-04-01 | Extraction method of genome DNA of isolated soy proteins |
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| CN102206628A true CN102206628A (en) | 2011-10-05 |
| CN102206628B CN102206628B (en) | 2012-12-26 |
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| CN 201110081615 Expired - Fee Related CN102206628B (en) | 2011-04-01 | 2011-04-01 | Extraction method of genome DNA of isolated soy proteins |
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Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN107937391A (en) * | 2017-11-29 | 2018-04-20 | 上海透景生命科技股份有限公司 | Cast-off cells nucleic acid extraction lysate and its preparation method and application in human faecal mass |
Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101413018A (en) * | 2008-12-09 | 2009-04-22 | 中南大学 | Method for extracting genome DNA |
| CN101575597A (en) * | 2009-05-21 | 2009-11-11 | 中国农业大学 | Kit for quickly extracting plant genome and applications thereof |
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2011
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Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101413018A (en) * | 2008-12-09 | 2009-04-22 | 中南大学 | Method for extracting genome DNA |
| CN101575597A (en) * | 2009-05-21 | 2009-11-11 | 中国农业大学 | Kit for quickly extracting plant genome and applications thereof |
Non-Patent Citations (1)
| Title |
|---|
| 《Proceedings of 2010 First International Conference on Cellular,Molecular Biology, Biophysics and Bioengineering》 20101231 徐伟丽等 熟豆浆基因组DNA提取方法的优化 247-251 1-10 第6卷, * |
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN107937391A (en) * | 2017-11-29 | 2018-04-20 | 上海透景生命科技股份有限公司 | Cast-off cells nucleic acid extraction lysate and its preparation method and application in human faecal mass |
| CN107937391B (en) * | 2017-11-29 | 2021-07-09 | 上海透景生命科技股份有限公司 | Lysate for extracting abscisic acid from human excrement and preparation method and application thereof |
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| CN102206628B (en) | 2012-12-26 |
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Granted publication date: 20121226 Termination date: 20140401 |