ITS2序列作为DNA条形码鉴定几种虎耳草科植物的有效性研究  

Validation studies of ITS2 sequence as DNA barcode to identify several Saxifragaceae

作　　者：任瑶瑶 郑颖 柯家伟 姚雨庭 谢浈 蒋合众[2] REN Yaoyao;ZHENG Ying;KE Jiawei;YAO Yuting;XIE Zhen;JIANG Hezhong(College of Chemistry,Southwest Jiaotong University,Chengdu 610031,China;School of Life Science and Engineering,Southwest Jiaotong University,Chengdu 610031,China)

机构地区：[1]西南交通大学化学学院,四川成都610031 [2]西南交通大学生命科学与工程学院,四川成都610031

出　　处：《实验技术与管理》2023年第4期20-24,32,共6页Experimental Technology and Management

基　　金：四川省科技厅重点研发计划(2023YFQ0001);四川省科技厅省院省校科技合作重点项目(2021YFSY0041);西南交通大学2022年校级实验教学研究与改革项目(20221460)。

摘　　要：利用DNA条形码分子诊断技术对几种虎耳草科药用植物进行鉴定并评价ITS2序列对虎耳草科植物鉴定的有效性,为虎耳草科植物鉴定提供一种新思路。于不同区域采集13份虎耳草科植物样本,提取基因组DNA后,利用ITS2序列扩增引物进行PCR扩增及测序,并从GenBank数据库中下载11条同属及同科序列。基于HHMer注释方法去掉两端区段获得ITS2序列,采用MEGA7.0软件进行序列长度、碱基含量对比,并基于K2P模型计算并分析种内、种间遗传距离,采用邻接法构建NJ系统进化树,采用靴带检验法(1000次重复)检验各分支的支持率,于ITS2数据库网站预测并比较ITS2序列的二级结构。结果显示,虎耳草科植物种间遗传距离大于种内遗传距离,能将各物种区分;系统进化树中各属分别聚为一支,同属内各物种大多分别聚为一小支,部分物种的鉴定需要进一步研究;各物种ITS2二级结构均存在不同差异,可对物种鉴定进行辅助分析。利用ITS2序列作为DNA条形码可有效地区分鉴别几种虎耳草科药用植物。Using DNA barcode molecular diagnostic technology to identify the medicinal plants of Saxifragaceae area and evaluate the effectiveness of ITS2 sequences in the identification,which providing a new idea for identifying plants of Saxifragaceae.Collecting 13 plant samples of Saxifragaceae in different areas to extract genomic DNA,PCR amplify and sequencing using ITS2 sequence amplification primers,then downloading 11 sequences of the same genus and family from the GenBank database.Removing the sequence at both ends to obtain the ITS2 sequences based on the HHMer annotation method,then using MEGA7.0 software to compare the sequence length and base content,and calculate and analyze the inter-and intra-specific genetic distance based on the K2P model.Constructing NJ phylogenetic tree using neighbor-joining way,checking the support rate of each branch using bootstrap inspection method(1000 repetitions).Predicting and comparing the secondary structures of the ITS2 sequences on the ITS2 database website.The results show that the intra-specific genetic distance is higher than the inter-specific genetic distance of Saxifragaceae, which can distinguish each species. Each genus is gathered into a branch in the phylogenetic tree, and most species in the same genus are gathered into a small branch, the identification of some species requires further study. The secondary structures of ITS2 sequences of each species are different, which can assist in the analysis of species identification. Using the ITS2 sequences as DNA barcode can identify the plants of several Saxifragaceae effectively.

关 键 词：虎耳草科 DNA条形码 ITS2序列 分子鉴定 遗传距离 

分 类 号：R282.5[医药卫生—中药学]