基于ITS2条形码及其二级结构的菊科药用植物的系统发育分析  

Phylogenetic Analysis of Compositae Medicinal Plants Based on ITS2 Barcode and ITS2 Secondary Structure

作　　者：韩芳平 谢才友 兰卓 任瑶瑶[1] 谭睿[1] Han Fangping;Xie Caiyou;Lan Zhuo;Ren Yaoyao;Tan Rui(School of Life Science and Engineering,Southwest Jiaotong University,Chengdu 610031,China)

机构地区：[1]西南交通大学生命科学与工程学院,成都610031

出　　处：《世界科学技术-中医药现代化》2020年第10期3760-3767,共8页Modernization of Traditional Chinese Medicine and Materia Medica-World Science and Technology

基　　金：四川省科学技术厅重点研发项目(2018SZ0061):快速鉴定冬虫夏草与川贝母分子技术的开发与应用,负责人:廖海;四川省中医药管理局重大专项(2016Z013):四川常用藏药化学基因双指纹信息建设,负责人:任瑶瑶;西南交通大学个性化实验项目(GX201812086):菊科药用植物ITS2条形码鉴定,负责人:任瑶瑶。

摘　　要：目的采用ITS2(Internal transcribed spacer2,内转录间隔区2)序列作为条形码技术对菊科药用植物进行鉴别。方法选取来自西藏自治区及四川康定、乐山和峨眉地区的菊科样品44种,提取DNA并对其进行PCR(Polymerase Chain Reaction,聚合酶链式反应)扩增,后测序,测序结果提交至GenBank;同时从GenBank上下载17条样本序列。使用MEGA7.0软件计算所提交和下载的61条序列的种内与种间遗传距离,采用邻接法(neighbor-joinging,NJ)构建系统聚类树直观反应鉴定结果。从ITS2数据库中预测并比较样本ITS2序列的二级结构差异。结果菊科植物种间最小遗传距离0.031大于种内最大遗传距离0.009,有较明显的barcoding gap;NJ树显示种各属样本分别聚为一大枝,种内之间各自聚成小支,表现出良好的单系性;各属及各种样品ITS2二级结构存在明显差异。结论以ITS2序列作为DNA条形码,能够对菊科药用植物进行准确、迅速的识别与鉴定,是物种间进化关系准确的分子鉴定依据,可为该科植物的分布、种群及种群数量的研究提供指导,为控制药品质量、合理开发利用及保护提供了新手段。Objective To identify the medicinal plants of Compositae using ITS2(Internal transcribed spacer2)sequence as a barcode technique. Methods Forty-four species of Compositae from Tibet Autonomous Region and Sichuan Kangding, Leshan and Emei regions were selected, their DNA was extracted and amplified by PCR(Polymerase Chain Reaction), then sequenced, and the sequencing results were submitted to GenBank. Seventeen sample sequences were downloaded from GenBank at the same time. Intraspecific and interspecific genetic distances were calculated for 61 sequences submitted and downloaded using MEGA7.0 software, and a systematic clustering tree was constructed using the neighbor-joining(NJ) method to visually respond to the identification results. Secondary structure differences of sample ITS2 sequences were predicted and compared from the ITS2 database. Results The minimum genetic distance between species is 0.031 which is greater than the maximum genetic distance within species is 0.009 in Asteraceae, and there is a more obvious barcoding gap. The NJ tree showed that species differed obviously from each other, gathering in two different main branches, and the intraspecities were clustered into small branches,which showed good monophyletic behavior. Same significant differences also appeared in the secondary structures, which could both be used to separate them. Conclusion Using ITS2 sequences as DNA barcodes can provide accurate and quick identification and characterization of medicinal plants in the Compositae family, which is an accurate molecular identification basis for evolutionary relationships among species, and can provide guidance for the study of distribution, population and population size of the family, and provide a new method for controlling medicinal plant quality, rational exploitation and conservation.

关 键 词：菊科 DNA条形码 ITS2序列 二级结构 分子鉴定 

分 类 号：R288[医药卫生—中药学]