李毅,张小蕾,蒋朝晖,吴晓黎,谢渊,单可人
LI Yi~1, ZHANG Xiao-lei~1, JIANG Zhao-hui~2, WU Xiao-Li~2, XIE Yuan~1, SHAN Ke-ren~1(1. The Key Laboratory of Molecular Biology of Guiyang Medical College, Guiyang 550004, China; 2. Guiyang College of Traditional Chinese Medicine, Guiyang 550001, China)

• 1:贵阳医学院分子生物学重点实验室
• 2:贵阳医学院分子生物学重点实验室
• 3:贵阳中医学院
• 4:贵阳中医学院

摘要(Abstract)：

目的:改良常用的植物DNA提取的CTAB,SDS法,以有效去除天麻多糖类杂质。方法:用CTAB法提取天麻鲜品不同部位(块茎、花序轴和胚芽)DNA;用生理盐水浸泡天麻干品之后,在应用CTAB,SDS法提取天麻干品的DNA;通过已知序列的天麻抗真菌蛋白基因片段的聚合酶链反应(PCR)扩增和PCR产物测序鉴定天麻DNA及其质量。结果:提取了44份天麻样本DNA,用CTAB法提取天麻鲜品不同部位的DNA,成功地用于PCR扩增,并通过DNA测序进一步鉴定;应用改良的CTAB,SDS法提取天麻干品DNA可用于PCR扩增,但DNA测序未成功。结论:用CTAB法提取的天麻鲜品各个部位DNA均可用于基因分析,而用天麻胚芽提取DNA的效果最理想;改良CTAB法能有效提取天麻干品DNA,提取的DNA可用于基因扩增。
Objective: To improve the common methods for extracting the plant DNA, so as to get rid of polyose impurity from Gastrodia elata frond. Methods: Using improved techniques of CTAB and SDS, we ideally extracted DNA from different parts of different samples of G. elata frond. Then, the extracted DNA was authenticated with PCR amplification and sequencing. Results: DNA extracted with CTAB technique from fresh samples of G.elata was successfully amplified with PCR technique and sequenced. DNA extracted with improved CTAB and SDS techniques from dried samples was also amplified with PCR, but its sequencing was not successful. Conclusions: DNA from all parts of G.elata fresh samples extracted with CTAB technique can be used for gene analysis, specially DNA from embryo bud. The improved CTAB technique can be used to extract DNA from dried samples of G.elata, but the DNA can be used only for gene amplification not for sequencing.

关键词(KeyWords)： 中草药;序列分析,DNA;聚合酶链反应;基因扩增;天麻抗真菌蛋白;天麻
drugs,Chinese herbal; sequence analysis,DNA; polymerase chain reaction; gene amplification; gene of antifungal protein of G.elata; G.elata

Abstract:

Keywords:

基金项目(Foundation): 贵州省中药现代化专项基金资助项目(200322号)

作者(Authors): 李毅,张小蕾,蒋朝晖,吴晓黎,谢渊,单可人
LI Yi~1, ZHANG Xiao-lei~1, JIANG Zhao-hui~2, WU Xiao-Li~2, XIE Yuan~1, SHAN Ke-ren~1(1. The Key Laboratory of Molecular Biology of Guiyang Medical College, Guiyang 550004, China; 2. Guiyang College of Traditional Chinese Medicine, Guiyang 550001, China)

DOI: 10.19367/j.cnki.1000-2707.2005.04.006

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