基于DNA条形码的藏药洪连鉴定研究

Identification of Tibetan Medicine Honglian Based on DNA Barcoding

目的:对藏区常使用的洪连药材进行DNA条形码鉴定,为准确鉴定藏区洪连药材提供方法。方法:从5个藏区收集12批洪连药材,提取其DNA,采用ITS以及psbA-trnH序列进行聚合酶链式反应(PCR)引物扩增并测序,所得序列从GenBank数据库进行相似性同源查询,采用DNAMAN、Editseq、Mega等软件,寻找其特异位点,计算样品Kimura2-parameter(K2-P)遗传距离,并构建N-J系统构建树。结果:12批样品来源于3个种,ITS序列长度为552 bp,变异位点为34个,在107 bp处能将3种兔耳草区分;psbA-trnH序列长度约为250 bp,存在12个变异位点,在136 bp处能将3种兔耳草区分。根据遗传距离构建N-J树,3种兔耳草明显分为3支。结论:ITS序列以及psbA-trnH序列可以作为兔耳草属鉴别的1种方法,为藏药的准确鉴别提供了方法。

Objective:To identify the DNA barcodes of Honglian medicinal materials commonly used in Tibetan areas,and to provide a method for accurately identifying Honglian medicinal materials in Tibetan areas.Methods:12 batches of Honglian medicinal materials were collected from 5 Tibetan areas,and the DNA was extracted.PCR primers were amplified and sequenced using ITS and psbA-trnH sequences.The sequence was similarly homologously searched from GenBank database,using DNAMAN,Editseq,Software such as Mega searches for its specific site,calculates the genetic distance of the sample Kimura2-parameter(k2-P),and constructs the NJ system building tree.Results:12 batches of samples were derived from three species.The length of ITS sequence was 552 bp and the number of mutation sites was 34.3 kinds of rabbit ear grasses could be distinguished at 107 bp.The length of psbA-trnH sequence was about 250 bp,and there were 12 mutation sites,3 kinds of Lagotis Gaertn can be distinguished at 136 bp.According to the genetic distance,N-J trees were constructed,and 3 kinds of Lagotis Gaertn were clearly divided into three branches.Conclusion:ITS sequence and psbA-trnH sequence can be used as a method to identify the ear genus of Aurora,providing a method for accurate identification of Tibetan medicine.