中国中药杂志

2018, v.43(04) 645-650

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DNA条形码技术在动物类药材熊胆粉及其混伪品鉴定中的应用
Identification of bear bile powder and its adulterants by using DNA barcoding technology

许亚春;熊超;姜春丽;段永波;孙伟;刘绍勇;薛东升;薛建平;
XU Ya-chun;XIONG Chao;JIANG Chun-li;DUAN Yong-bo;SUN Wei;LIU Shao-yong;XUE Dong-sheng;XUE Jian-ping;School of Life Sciences,Huaibei Normal University;Institute of Chinese Materia Medica,China Academy of Chinese Medical Sciences;Shanghai Kaibao Pharmaceutical Co.,Ltd.;

摘要(Abstract):

应用DNA条形码技术对名贵中药材熊胆粉及混伪品进行DNA条形码鉴定研究,并建立其标准实验流程,以保障熊胆粉的安全有效利用。对收集到的12份熊胆粉样品进行DNA提取、PCR(聚合酶链式反应)扩增并双向测序、运用Codon Code Aligner V 7.0.1进行序列剪切去除引物区得到COI序列,同时与Gen Bank中得到棕熊及混伪品COI序列,运用MEGA 7.0对研究的5个物种50条序列比对分析,计算变异位点及种内和种间K2P(Kimura 2-Parameter)遗传距离并构建邻接(NJ)树。黑熊、棕熊的种内最大K2P遗传距离远小于其混伪品间最小K2P遗传距离;NJ树结果显示黑熊、棕熊各聚为一支,均可以与混伪品明显区分开。DNA条形码技术为一种安全便捷可靠的物种鉴定技术,建立动物类药材COI序列标准话流程,在中药研究领域具有重要的作用。
To identify the precious bile powder and its adulterants by DNA barcoding,and establish its standard experimental process to ensure the safe and effective utilization. Total twelve sequences from samples of bear bile powder which come from Ursus thibetanus for DNA extraction,PCR(polymerase chain reaction) and sequence,then using Codon Code Aligner V 7. 0. 1 shear primer region to obtain COI sequence. The COI sequences of U. arctos and their adulterants were obtained from Gen Bank. MEGA7. 0 software was applied for analyzing mutation,calculating intraspecific and interspecific K2 P(Kimura 2-Parameter) genetic distance and constructing the Neighbor-joining tree(NJ). The results showed that the maximum K2 P genetic distance of bear bile powder of U. thibetanus and U. arctos are far less than minimum K2 P genetic distance within its adulterants species,and the results of NJ tree demonstrated that each species could be distinguished from the counterfeits obviously. DNA barcoding is a safe,convenient and reliable technique for species identification,and it is important to establish the standard sequence of COI sequences for animal medicines.

关键词(KeyWords): 熊胆粉;COI序列;DNA条形码技术;分子鉴定
bear bile powder;COI sequence;DNA barcoding;adulterant molecular identification

Abstract:

Keywords:

基金项目(Foundation): 国家自然科学基金项目(81573518,81603248);; 安徽省自然科学基金项目(1408085MC58);; 中央级公益性科研院所基本科研业务费专项(GH2017-01-01,GH2017-01)

作者(Authors): 许亚春;熊超;姜春丽;段永波;孙伟;刘绍勇;薛东升;薛建平;
XU Ya-chun;XIONG Chao;JIANG Chun-li;DUAN Yong-bo;SUN Wei;LIU Shao-yong;XUE Dong-sheng;XUE Jian-ping;School of Life Sciences,Huaibei Normal University;Institute of Chinese Materia Medica,China Academy of Chinese Medical Sciences;Shanghai Kaibao Pharmaceutical Co.,Ltd.;

DOI: 10.19540/j.cnki.cjcmm.2018.0014

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