红豆杉属植物三种不同总DNA提取方法的分析比较

Comparative analysis of three different methods of total DNA extraction used in Taxus

关键词:  红豆杉属  DNA提取方法  PCR扩增

Key words:  Taxus  DNA extraction methods  PCR amplification

摘要:

红豆杉属植物均为濒危物种,也是国家一级保护植物。以红豆杉属植物叶片为材料,利用三种不同的DNA提取方法提取总DNA,用分光光度计和琼脂糖凝胶电泳方法检测所得总DNA的得率和纯度,用PCR扩增的方法检测所得总DNA的质量,并对三种不同提取方法的结果进行了比较分析。结果表明:CTAB法提取的DNA纯度和得率均较高,可直接用于下游分子生物学实验; 用试剂盒提取的总DNA质量也很好,纯度高,但得率较低且成本较高; SDS法提取的总DNA质量较差,很难直接用于下游分子生物学实验。同时对影响植物总DNA得率和纯度的限制因素进行了讨论,为植物总DNA提取方法的改进和选择提供借鉴作用。

Abstract:

Species of Taxus in China are endangered and were listed as the national first-class protected wild plant. In present study,three different methods of total DNA extraction,i.e.SDS,CTAB and DNA extraction Kit(DNeasy Plant Mini Kit),were used to isolate genomic DNA from dried leaf materials of Taxus. The total genomic DNA yielded by the three methods were quantified and analyzed by spectrophotometer,agarose gel electrophoresis and PCR reaction respectively. The results of the three methods of DNA extraction were compared and analyzed,which indicated that the CTAB method could yielded relatively pure and high amount total DNA and was highly suited for use directly in downstream applications. The method of DNA extraction using Kit(DNeasy Plant Mini Kit)also could yield high quality total DNA with the highest purity of the DNA comparing to other methods used in this study. However,the relatively lower DNA yield and higher costs of this method might be limited to its wide use. Whereas the SDS method could not yield high quantity of total DNA in most sampled accessions in the present study,which indicated low purity of the yielded DNA of SDS method could not be used in downstream application directly. We also discussed the factors influencing on the total genomic DNA yield and purity,which could help to improve and select the suitable protocol of DNA isolation of plants.