ITS2序列鉴定“功劳叶”药材及其近缘混伪品  

Identification of “Gonglao Leaf ” and Its Sibling Adulterants Based on ITS2 Sequence

作　　者：夏叶[1] 周红[2] 向丽[3] 张秀桥[1] 胡志刚[1,3] 

机构地区：[1]湖北中医药大学药学院,武汉430065 [2]中国医学科学院北京协和医学院药用植物研究所,北京100193 [3]中国中医科学院中药研究所,北京100700

出　　处：《世界科学技术-中医药现代化》2014年第11期2361-2365,共5页Modernization of Traditional Chinese Medicine and Materia Medica-World Science and Technology

基　　金：科学技术部国家重大新药创制专项(2014ZX09304307001-021):中药新药安全性检测技术与标准研究-中药原料药混伪品分子基因鉴定技术与标准;负责人:胡志刚;国家自然科学基金委重点项目(81130069):道地药材形成的生物学实质;负责人:陈士林

摘　　要：目的:中药材同名异物现象对临床安全用药造成了潜在的威胁。本研究对两种"功劳叶"药材及其近缘混伪品进行分子鉴定,以确保临床准确用药。方法:本实验收集枸骨、阔叶十大功劳和细叶十大功劳及其近缘混伪品共9个物种45份实验样品,用改良的CTAB法提取总DNA,扩增ITS2序列并双向测序,采用Condon Code Aligner V 4.2.4对测序峰图进行拼接,应用MEGA 6.0进行序列变异分析,计算种内种间Kimura 2-Parameter(K2P)遗传距离,构建系统邻接(NJ)树。结果:基于ITS2序列的序列变异、最近距离法和系统邻接树分析结果均表明,冬青属枸骨以及十大功劳属阔叶十大功劳和细叶十大功劳均能与其混伪品很好地区分开。结论:ITS2序列可有效区分两种"功劳叶"药材及其近缘混伪品,为临床准确用药提供依据。Objective： The Phenomenon of different species with the same name has created potential threats to cinical safety of medication. Because of that, two kinds of ＂gonglao leaf＂ and their sibling adulterants have been identified by molecular methods in this study, to secure its safety in medication. Methods： Ilexcornuta, Mahoniabealei, Mahoniafortuneiand their sibling adulterants total of 9 species 45 samples were collected in this experiment,Total genomic DNA was extracted from them by the method of improved CTAB, ITS2 sequences were amplified, and the PCR products were sequenced. Sequence assembly and consensus sequence generation were performed by the CodonCode Aligner V 4.2.4.. The genetic distances were computed by MEGA 6.0 in accordance with the Kimura 2-Parameter （K2P） model and the phylogenetic tree constructed by the neighbor-joining （NJ） method. Results： The analysis results of the genetic distances, variable sites and the NJ phylogenetic tree showed that Ilexcornuta, Mahoniabealei, Mahoniafortunei were seperated from their sibling adulterants obviously. Conclusion： ITS2 sequence was able to identify two kinds of ＂gonglao leaf＂ and their sibling adulterants which can provide a basis for clinical accurate medication.

关 键 词：枸骨叶 功劳叶 ITS2序列 分子鉴定 近缘混伪品 

分 类 号：R284.1[医药卫生—中药学]