植物类药材基因组DNA提取与纯化的方法学研究

植物类药材基因组DNA提取与纯化的方法学研究

曹晖, 毕培曦, 邵鹏柱

1. 中国中医研究院中药研究所, 北京 100700;
2. 香港中文大学生物系, 香港;
3. 香港中文大学生物化学系, 香港;
4. 香港中文大学中药研究中心, 香港

收稿日期:1997-05-16 修回日期:1998-04-07 出版日期:1998-09-15 发布日期:1998-09-15

Methodological Studies on Genomic DNA Extraction and Purification from Plant Drug Materials

Hui Cao, Paul Pui-Hay But, Pang-Chui Shaw

1.I nstitute of Chinese Materia Medica, China Academy of Traditional Chinese Medicine, Beijing 100700;
2. Department of Biology, The Chinese University of HongKong, HongKong;
3. Department of Biochemistry, The Chinese University of HongKong, HongKong;
4. Chinese Medicinal Material Research Center, The Chinese University of HongKong, HongKong

Received:1997-05-16 Revised:1998-04-07 Online:1998-09-15 Published:1998-09-15

摘要/Abstract

摘要： 本文探讨了菊科9种植物和地胆草的4种对口商品药材基因组DNA提取与纯化的原理、方法通过对3种常用植物基因组DNA提取方法(CsCl梯度超速离心法、CTABCsCl梯度超速离心法和CTAB微量提取法)的条件摸索在DNA产率、纯度以及提取纯化过程中影响PCR扩增因子方面进行比较,认为CTAB微量提取法是植物类药材基因组DNA一种比较省时、有效、经济的提取方法。

关键词: DNA提取, DNA纯化, CTAB微量提取法, 基因组指纹图谱法, 任意引物聚合酶链反应

Abstract: This paper reports the rationale and methods of DNA extraction and purification from nine species of Compositae and four commercial drugs of corresponding plant Elephantopus scaber. The comparison of three methods: CsCl gradient, CTAB/CsCl gradient and CTAB miniprep extraction by yield, purity and factors affecting PCR was carried out. In conclusion, CTAB miniprep method provides a rapid, effective, economic approach for isolating genomic DNA for Chinese drug identification by genomic fingerprints.

Key words: DNA extraction, DNA purification, CTAB miniprep, Genomic fingerprinting, AP-PCR

Supporting:

曹晖, 毕培曦, 邵鹏柱. 植物类药材基因组DNA提取与纯化的方法学研究[J]. .

Hui Cao, Paul Pui-Hay But, Pang-Chui Shaw. Methodological Studies on Genomic DNA Extraction and Purification from Plant Drug Materials[J]. .

参考文献

[1] Saiki RK, Scharf SJ, Faloona F, Mullis KB, Horn GT, Erlich HA, Arnheim N. Science, 1985, 230: 1350.
[2] Mullis KB, Faloona F, Scharf S, Saiki RK, Horn GT, Erlich HA. Cold Spring Harbor Symp Quant Biol, 1986, 51: 263.
[3] Welsh J, McClelland M. Nucl Acids Res, 1990, 18: 7213.
[4] Williams J GK, Kubelik AR, Livak KJ, Rafalski JA, Tingey SV. Nucl Acids Res, 1990, 18: 6531.
[5] Pääbo S. Amplifying Ancient DNA. In: Innis MA, Gelfend DH, Sninsky JJ, White TJ. eds. PCR Protocols. A Guide to Methods and Application. New York: Academic Press, 1990: 159~166.
[6] Cao H, Liu YP. Trad Chin Drug Res & Clin Pharmacol, 1997, 8: 112.
[7] Saghai-Maroof MA, Soliman KM, Jorgensen RA, Allard RW. Proc Natl Acad Sci USA, 1984, 81: 8014.
[8] Doyle JJ, Doyle JL. Phytochem Bull, 1987, 19: 11.
[9] Doyle JJ . Doyle JL. Focus (BRL), 1990, 12: 13.
[10] Deragon JM, Landry BS. Methods and Applications (Cold Spring Harbor Laboratory Press), 1992, 1: 175.
[11] Edwards K, Johnstone C, Thompson C. Nucl Acids Res, 1991, 19: 1349.
[12] Michaels SD, John MC, Amasino RM. BioTechniques, 1994, 17: 274.
[13] Richards E. Preparation of Genomic DNA from Plant Tissue. In: Ausulbel FM, Brent R, Kingston RE, Moore DD, Smith JA, Seidman JG, Struhl K. eds. Current Protocols in Molecular Biology. Vol 1, New York: John Wiley & Sons, 1987: 2.1.1~2.3.3.
[14] Treco DA. Purification and Concentration of DNA from Aqueous Solutions. In: Ausulbel FM, Brent R, Kingston RE, Moore DD, Smith JA, Seidman J Gand Struhl K, eds. Current Protocols in Molecular Biology, Vol 1, New York: John Wiley & Sons, 1987: 2.1.1~2.1.3.
[15] Draper J, Scott R. The Isolation of Plant Nucleic Acids. In: Draper J, Scott R, Armitage P, Walden R, eds. Plant Genetic Transformation and Gene Expression. A Laboratory Manual. London: Blockwell Scientific Publishing Co., 1988: 199~236.
[16] Doyle JJ . DNA Protocols for Plants. In: Hewitt GM, Johnston AWB, Young J PW. eds. Molecular Techniques in Taxonomy. Berlin: Springer-Verlag, 1991: 283~285.
[17] Doyle JJ, Dickson EE. Taxon, 1987, 36: 715.
[18] Rogers SO, Bendich AJ. Plant Mol Biol, 1985, 5: 69.
[19] Sambrook J, Fritsch EF, Maniatis T. Molecular Cloning. A L aboratory Manual . 2nd ed. New York: Cold Spring Harbor Laboratory Press, 1989: 150~161.
[20] Bassam BJ, Caetano-Anoll (sG. BioTechniques, 1993, 14: 31.
[21] Zhang ML, Zuo J, Xu XS, Fang FD, Chen L Y. Acta Acad Med Sin, 1994, 16: 348.
[22] Murray HG, Thompson WF. Nucl Acids Res, 1980, 8: 4321.
[23] Taylor BH, Powell A. Focus (BRL), 1982, 4: 4.
[24] Taylor BH, Manhart JR, Amasino RM. Isolation and Characterization of Plant DNAs. In: Glick BR, Thompson JE. eds. Methods in Plant Molecular Biology and Biotechnology . Boca Raton: CRC Press, 1993: 37~47.
[25] Fang GW, Hammer S, Grumet R. BioTechniques, 1992, 13: 52.
[26] John ME. Nucl Acids Res, 1992, 20: 2381.