棘豆属蒙药材的DNA条形码鉴定研究

DNA Barcoding of Mongolian Oxytropis Medicinal Materials

目的:利用ITS2序列对棘豆属蒙药材进行分子鉴定。方法:对棘豆属正品药材硬毛棘豆Oxytropis fetissovii Bge.、多叶棘豆Oxytropis myriophylla(Pall.)DC.样品和伪品大花棘豆Oxytropis grandiflora(Pall.)DC.样品的ITS2序列进行了PCR扩增和测序。为扩大研究范围,又从Gen Bank中下载了棘豆属与黄芪属植物样本的ITS2序列。经ITS2 Database网站处理截取ITS2序列,并采用MEGA 5.10计算相关数据,基于K2P模型构建聚类树(NJ树)。并从ITS2网站上获得样本及下载序列的ITS2二级结构信息,分析各样本间ITS2序列二级结构的差异。结果:棘豆属药材的ITS2序列较短,序列长度为216~218 bp,有利于DNA的提取、PCR扩增和测序。多叶棘豆Oxytropis myriophylla(Pall.)DC.、硬毛棘豆Oxytropis fetissovii Bge.与大花棘豆Oxytropis grandiflora(Pall.)DC.的种内遗传距离大于种间遗传距离。在NJ树模型中,棘豆属正品药材与非正品样品可以区分开;与黄芪Astragalus membranaceus(Fisch.)Bge.比较也可以区分开。结论:以ITS2序列为标准的DNA条形码能够有效的鉴定棘豆属药材。

Objective：To identify Oxytropis medicinal materials using ITS2 sequence. Methods：The second internal transcribed spacer（ITS2） of Oxytropisfetissovii ,Oxytropis myriophylla and Oxytropis grandiflora medicinal material samples was amplified by PCR and sequenced. To expand scope of the research topic,ITS2 sequences of related species were downloaded fi＇om GenBank. Sequences as- sembly and consensus sequence generation were performed by ITS2 Database. The related data analysis and processing was performed u- sing software MEGA 5. 10 and the NJ tree was constructed. The ITS2 secondary structure was predicted using ITS2 web server,and the differences of the ITS2 secondary structures of the samples were analyzed. Results： Oxytropis medicinal materials ITS2 sequence was shorter,with sequence length of 216 ~ 218 bp,which was in favor of DNA extraction, PCR amplification and sequencing. Genetic dis- tances of Oxytropis myriophylla, Oxytropis fetissovii and Oxytropis grand^lora were much larger than the genetic distances of them- selves. In the NJ tree, Oxytropis medicinal materials and counterfeits could be distinguished, and Oxytropis medicinal materials could be distinguished from Astragalus membranaceus. Conclusion：The DNA barcode based on ITS2 sequence is a powerful and efficient tool for identification of Oxytropis medicinal materials.