CN110914424A - 一种优化的大片段dna提取方法 - Google Patents
一种优化的大片段dna提取方法 Download PDFInfo
- Publication number
- CN110914424A CN110914424A CN201780090792.8A CN201780090792A CN110914424A CN 110914424 A CN110914424 A CN 110914424A CN 201780090792 A CN201780090792 A CN 201780090792A CN 110914424 A CN110914424 A CN 110914424A
- Authority
- CN
- China
- Prior art keywords
- solution
- supernatant
- fragment dna
- large fragment
- centrifuging
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Pending
Links
- 238000000034 method Methods 0.000 title claims abstract description 25
- 239000012634 fragment Substances 0.000 title claims abstract description 19
- 238000007400 DNA extraction Methods 0.000 title claims description 13
- 239000000243 solution Substances 0.000 claims abstract description 46
- HEDRZPFGACZZDS-UHFFFAOYSA-N Chloroform Chemical compound ClC(Cl)Cl HEDRZPFGACZZDS-UHFFFAOYSA-N 0.000 claims abstract description 18
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 claims abstract description 16
- 238000000605 extraction Methods 0.000 claims abstract description 14
- PHTQWCKDNZKARW-UHFFFAOYSA-N isoamylol Chemical compound CC(C)CCO PHTQWCKDNZKARW-UHFFFAOYSA-N 0.000 claims abstract description 13
- 238000000227 grinding Methods 0.000 claims abstract description 10
- 238000005336 cracking Methods 0.000 claims abstract description 8
- 239000011780 sodium chloride Substances 0.000 claims abstract description 8
- 108020002230 Pancreatic Ribonuclease Proteins 0.000 claims abstract description 6
- 102000005891 Pancreatic ribonuclease Human genes 0.000 claims abstract description 6
- KCXVZYZYPLLWCC-UHFFFAOYSA-N EDTA Chemical compound OC(=O)CN(CC(O)=O)CCN(CC(O)=O)CC(O)=O KCXVZYZYPLLWCC-UHFFFAOYSA-N 0.000 claims abstract description 4
- WHNWPMSKXPGLAX-UHFFFAOYSA-N N-Vinyl-2-pyrrolidone Chemical compound C=CN1CCCC1=O WHNWPMSKXPGLAX-UHFFFAOYSA-N 0.000 claims abstract description 4
- 229920002594 Polyethylene Glycol 8000 Polymers 0.000 claims abstract description 4
- 239000007983 Tris buffer Substances 0.000 claims abstract description 4
- 238000004140 cleaning Methods 0.000 claims abstract description 4
- HRZFUMHJMZEROT-UHFFFAOYSA-L sodium disulfite Chemical compound [Na+].[Na+].[O-]S(=O)S([O-])(=O)=O HRZFUMHJMZEROT-UHFFFAOYSA-L 0.000 claims abstract description 4
- 235000010262 sodium metabisulphite Nutrition 0.000 claims abstract description 4
- LENZDBCJOHFCAS-UHFFFAOYSA-N tris Chemical compound OCC(N)(CO)CO LENZDBCJOHFCAS-UHFFFAOYSA-N 0.000 claims abstract description 4
- 239000006228 supernatant Substances 0.000 claims description 31
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 claims description 22
- KFZMGEQAYNKOFK-UHFFFAOYSA-N Isopropanol Chemical compound CC(C)O KFZMGEQAYNKOFK-UHFFFAOYSA-N 0.000 claims description 15
- 238000002156 mixing Methods 0.000 claims description 10
- 239000000203 mixture Substances 0.000 claims description 10
- IJGRMHOSHXDMSA-UHFFFAOYSA-N Atomic nitrogen Chemical compound N#N IJGRMHOSHXDMSA-UHFFFAOYSA-N 0.000 claims description 8
- 238000011534 incubation Methods 0.000 claims description 8
- 239000000843 powder Substances 0.000 claims description 7
- 239000002244 precipitate Substances 0.000 claims description 7
- SCVFZCLFOSHCOH-UHFFFAOYSA-M potassium acetate Chemical compound [K+].CC([O-])=O SCVFZCLFOSHCOH-UHFFFAOYSA-M 0.000 claims description 6
- 238000001556 precipitation Methods 0.000 claims description 6
- 239000007788 liquid Substances 0.000 claims description 5
- 238000007605 air drying Methods 0.000 claims description 4
- 238000005119 centrifugation Methods 0.000 claims description 4
- 229910052757 nitrogen Inorganic materials 0.000 claims description 4
- 230000008021 deposition Effects 0.000 claims description 3
- 239000003480 eluent Substances 0.000 claims description 3
- 235000011056 potassium acetate Nutrition 0.000 claims description 3
- 229940001584 sodium metabisulfite Drugs 0.000 claims description 3
- 239000000725 suspension Substances 0.000 claims description 3
- 235000013824 polyphenols Nutrition 0.000 abstract description 5
- 229920001282 polysaccharide Polymers 0.000 abstract description 5
- 239000005017 polysaccharide Substances 0.000 abstract description 5
- -1 polysaccharide polyphenol Chemical class 0.000 abstract description 3
- VMHLLURERBWHNL-UHFFFAOYSA-M Sodium acetate Chemical compound [Na+].CC([O-])=O VMHLLURERBWHNL-UHFFFAOYSA-M 0.000 abstract 1
- DBMJMQXJHONAFJ-UHFFFAOYSA-M Sodium laurylsulphate Chemical compound [Na+].CCCCCCCCCCCCOS([O-])(=O)=O DBMJMQXJHONAFJ-UHFFFAOYSA-M 0.000 abstract 1
- 230000001376 precipitating effect Effects 0.000 abstract 1
- 239000001632 sodium acetate Substances 0.000 abstract 1
- 235000017281 sodium acetate Nutrition 0.000 abstract 1
- 239000004296 sodium metabisulphite Substances 0.000 abstract 1
- 241000894007 species Species 0.000 description 10
- 241001523681 Dendrobium Species 0.000 description 5
- 241000167854 Bourreria succulenta Species 0.000 description 4
- 235000019693 cherries Nutrition 0.000 description 4
- 238000007671 third-generation sequencing Methods 0.000 description 4
- 241000251468 Actinopterygii Species 0.000 description 3
- LZZYPRNAOMGNLH-UHFFFAOYSA-M Cetrimonium bromide Chemical compound [Br-].CCCCCCCCCCCCCCCC[N+](C)(C)C LZZYPRNAOMGNLH-UHFFFAOYSA-M 0.000 description 3
- 241000227653 Lycopersicon Species 0.000 description 3
- 235000007688 Lycopersicon esculentum Nutrition 0.000 description 2
- 230000009089 cytolysis Effects 0.000 description 2
- 238000001962 electrophoresis Methods 0.000 description 2
- 150000004676 glycans Chemical class 0.000 description 2
- 239000004570 mortar (masonry) Substances 0.000 description 2
- 150000008442 polyphenolic compounds Chemical class 0.000 description 2
- 238000012360 testing method Methods 0.000 description 2
- 238000005406 washing Methods 0.000 description 2
- 235000002568 Capsicum frutescens Nutrition 0.000 description 1
- 240000008574 Capsicum frutescens Species 0.000 description 1
- 238000006243 chemical reaction Methods 0.000 description 1
- 238000001816 cooling Methods 0.000 description 1
- 238000001514 detection method Methods 0.000 description 1
- 238000010586 diagram Methods 0.000 description 1
- 230000000694 effects Effects 0.000 description 1
- 239000012149 elution buffer Substances 0.000 description 1
- 238000005516 engineering process Methods 0.000 description 1
- 238000002474 experimental method Methods 0.000 description 1
- 239000003550 marker Substances 0.000 description 1
- 239000000463 material Substances 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 238000005185 salting out Methods 0.000 description 1
- 238000005070 sampling Methods 0.000 description 1
- 238000006467 substitution reaction Methods 0.000 description 1
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 1
- 238000012070 whole genome sequencing analysis Methods 0.000 description 1
Images
Classifications
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N15/00—Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
- C12N15/09—Recombinant DNA-technology
- C12N15/10—Processes for the isolation, preparation or purification of DNA or RNA
Landscapes
- Genetics & Genomics (AREA)
- Engineering & Computer Science (AREA)
- Chemical & Material Sciences (AREA)
- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Biomedical Technology (AREA)
- Wood Science & Technology (AREA)
- Organic Chemistry (AREA)
- Biotechnology (AREA)
- General Engineering & Computer Science (AREA)
- Zoology (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Plant Pathology (AREA)
- Molecular Biology (AREA)
- Microbiology (AREA)
- Crystallography & Structural Chemistry (AREA)
- Physics & Mathematics (AREA)
- Biochemistry (AREA)
- General Health & Medical Sciences (AREA)
- Biophysics (AREA)
- Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)
Abstract
提供了一种大片段DNA的提取方法，包括三个步骤：（1）样品研磨与裂解：其中使用了包含PVP、焦亚硫酸钠、氯化钠、Tris、EDTA和SDS的第一溶液和核糖核酸酶A；（2）抽提：分别使用了5M醋酸钠的第二溶液，包含PEG8000和氯化钠的第三溶液后，再用氯仿/异戊醇抽提；（3）沉淀与清洗。该方法可用于解决多糖多酚物种的提取难题，提取样品完整性大于40kb。
Description
PCT国内申请，说明书已公开。
Claims (10)
- PCT国内申请，权利要求书已公开。
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| PCT/CN2017/098455 WO2019036899A1 (zh) | 2017-08-22 | 2017-08-22 | 一种优化的大片段dna提取方法 |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN110914424A true CN110914424A (zh) | 2020-03-24 |
Family
ID=65439956
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201780090792.8A Pending CN110914424A (zh) | 2017-08-22 | 2017-08-22 | 一种优化的大片段dna提取方法 |
Country Status (2)
| Country | Link |
|---|---|
| CN (1) | CN110914424A (zh) |
| WO (1) | WO2019036899A1 (zh) |
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN111440789A (zh) * | 2020-04-09 | 2020-07-24 | 嘉兴菲沙基因信息有限公司 | 一种金银莲花基因组dna的提取方法 |
| CN113278610A (zh) * | 2021-06-17 | 2021-08-20 | 浙江省亚热带作物研究所 | 一种秋茄rna的提取方法 |
Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102643794A (zh) * | 2012-04-05 | 2012-08-22 | 湖北省农业科学院经济作物研究所 | 一种采用多手段提取桑树根围土壤微生物总dna的方法 |
| CN105368815A (zh) * | 2015-11-25 | 2016-03-02 | 上海派森诺生物科技股份有限公司 | 多糖多酚植物基因组的提取方法 |
| CN105368818A (zh) * | 2015-12-10 | 2016-03-02 | 上海交通大学 | 一种适于茄果类主要蔬菜的高效、快速dna提取方法 |
Family Cites Families (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN100398647C (zh) * | 2006-03-02 | 2008-07-02 | 上海交通大学 | 大片段海绵宏基因组dna快速提取方法 |
| CN106894094A (zh) * | 2015-12-17 | 2017-06-27 | 中国海洋大学 | 构建的一个南极乔治王岛土壤宏基因组文库 |
| CN105505920A (zh) * | 2016-02-23 | 2016-04-20 | 南京农业大学 | 一种提取顽拗植物组织dna的新方法 |
-
2017
- 2017-08-22 CN CN201780090792.8A patent/CN110914424A/zh active Pending
- 2017-08-22 WO PCT/CN2017/098455 patent/WO2019036899A1/zh active Application Filing
Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102643794A (zh) * | 2012-04-05 | 2012-08-22 | 湖北省农业科学院经济作物研究所 | 一种采用多手段提取桑树根围土壤微生物总dna的方法 |
| CN105368815A (zh) * | 2015-11-25 | 2016-03-02 | 上海派森诺生物科技股份有限公司 | 多糖多酚植物基因组的提取方法 |
| CN105368818A (zh) * | 2015-12-10 | 2016-03-02 | 上海交通大学 | 一种适于茄果类主要蔬菜的高效、快速dna提取方法 |
Non-Patent Citations (1)
| Title |
|---|
| BAPTISTE MAYJONADE等: "Extraction of high-molecular-weight genomic DNA for long-read sequencing of single molecules" * |
Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN111440789A (zh) * | 2020-04-09 | 2020-07-24 | 嘉兴菲沙基因信息有限公司 | 一种金银莲花基因组dna的提取方法 |
| CN113278610A (zh) * | 2021-06-17 | 2021-08-20 | 浙江省亚热带作物研究所 | 一种秋茄rna的提取方法 |
| CN113278610B (zh) * | 2021-06-17 | 2022-05-20 | 浙江省亚热带作物研究所 | 一种秋茄rna的提取方法 |
Also Published As
| Publication number | Publication date |
|---|---|
| WO2019036899A1 (zh) | 2019-02-28 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| Meisel et al. | A rapid and efficient method for purifying high quality total RNA from peaches (Prunus persica) for functional genomics analyses | |
| CN107868834B (zh) | 一种达氏鲟性腺差异表达基因的实时定量pcr检测引物组及方法 | |
| CN103146683B (zh) | 一种提取哺乳动物和鸟类粪便dna的方法 | |
| CN110914424A (zh) | 一种优化的大片段dna提取方法 | |
| CN108949751B (zh) | 一种提取富含果胶类多糖植物dna的试剂盒及方法 | |
| CN102115743A (zh) | 一种从石蜡包埋组织中提取高质量rna的新方法和试剂盒 | |
| Yu et al. | An easily-performed high-throughput method for plant genomic DNA extraction | |
| CN105713901A (zh) | 一种改良的多糖多酚植物高山杜鹃总dna提取方法 | |
| CN104531679A (zh) | 从干燥杏叶片中提取dna的方法 | |
| Gautam | Isolation of DNA from blood samples by salting method | |
| CN114621950A (zh) | 无内毒素质粒快速提取试剂盒及质粒提取方法 | |
| CN116218840A (zh) | 莲叶桐核酸提取改良版ctab法 | |
| CN113502286A (zh) | 富含次生代谢物的植物组织中dna的提取方法 | |
| CN113136384A (zh) | 濒危半红树植物莲叶桐核酸提取的方法 | |
| CN108018345A (zh) | 应用于组织样本中染色体开放结合区域定位的ATAC-seq方法 | |
| CN116064838A (zh) | 检测食品中牡蛎成分的引物组、试剂盒和方法 | |
| CN110295164B (zh) | 一种提取鲍肝胰腺rna的方法 | |
| CN103966201B (zh) | 基于样品高效保鲜的提取水生植物dna的方法 | |
| CN114703173A (zh) | 一种λ噬菌体DNA提取试剂盒及提取方法 | |
| CN106011130B (zh) | 一种土壤dna提取试剂盒及提取土壤dna的方法 | |
| CN110923304A (zh) | 一种用于鉴别银杏性别的分子标记、引物对和方法 | |
| Shyu et al. | Comparison of Six DNA Extraction Procedures and the Application of Plastid DNA Enrichment Methods in Selected Non-photosynthetic Plants. | |
| CN115478073A (zh) | 一种利用改良ctab法提取铁皮石斛茎基因组dna的方法 | |
| Buragohain et al. | An Efficient and Reliable Method of DNA Extraction from Meyna spinosa—A Traditional Medicinal Plant from North-East India | |
| Sasikala et al. | Analysis of Cytogenetic Alterations in Patients Affected by Hepatocellular Carcinoma in Tamilnadu Population |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication | ||
| PB01 | Publication | ||
| SE01 | Entry into force of request for substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| RJ01 | Rejection of invention patent application after publication |
Application publication date: 20200324 |