基于DNA条形码技术的制草乌疑似伪品基原鉴定  

Original identification of doubtful counterfeit of Aconiti Kusnezoffi Radix Cocta based on DNA barcoding technology

作　　者：刘杰[1,2] 过立农[2] 马双成[2] 高妍[2] 郑健 王俊丽[1] LIU Jie;GUO Li-nong;MA Shuang-cheng;GAO Yan;ZHENG Jian;WANG Jun-li(Minzu University of China,Beijing 100081,China;National Institutes for Food and Drug Control,Beijing 102629,China)

机构地区：[1]中央民族大学,北京100081 [2]中国食品药品检定研究院,北京102629

出　　处：《药物分析杂志》2020年第2期246-252,共7页Chinese Journal of Pharmaceutical Analysis

摘　　要：目的:基于DNA条形码技术鉴定制草乌疑似伪品的基原。方法:通过聚合酶链式反应(PCR)对建库样品及制草乌药材的ITS2序列进行扩增;对建库样品进行N-J聚类和变异位点分析,通过与GenBank数据库进行Blast比对,以及通过与建库样品进行比对推断制草乌药材疑似伪品的基原。结果:制草乌药材疑似伪品经Blast比对,结果显示能比对到多个种,且一致率均达到100%;建库样品ITS2序列进行N-J聚类和变异位点分析,结果均显示乌头与北乌头不能区分,其余建库样品均可区分;通过将5.8S与28S区间位点准确的56条制草乌药材疑似伪品的ITS2序列与建库样品的ITS2序列进行变异位点分析,其中2条序列与康定乌头完全一致,18条序列与准噶尔乌头完全一致,6条序列与甘青乌头完全一致,剩余30条序列与乌头、北乌头序列一致。结论:鉴定结果显示,市场制草乌药材存在掺伪及全伪现象,混伪品有康定乌头、甘青乌头、准噶尔乌头,其中准噶尔乌头为主要混伪品,此研究为DNA条形码技术鉴别制草乌正伪品开辟了一条新途径。Objective:To identify origin of doubtful counterfeit of Aconiti Kusnezoffi Radix Cocta samples based on DNA barcoding technology. Methods:ITS2 sequences of doubtful fake Aconiti Kusnezoffi Radix Cocta samples and samples to create a database for reference were amplified with polymerase chain reaction(PCR). ITS2 sequences of samples to create a database for reference were analyzed by the methods of N-J phylogenetic and variation sites comparison. And ITS2 sequences of doubtful fake Aconiti Kusnezoffi Radix Cocta samples were blasted with GenBank database and ITS2 sequences of samples to create a database for reference to determine species. Results:The Blast result of doubtful fake Aconiti Kusnezoffi Radix Cocta were multiple species with consistency of 100%. The analysis result of N-J phylogenetic and variation sites comparison indicated that Aconitum carmichaelii and Aconitum kusnezoffii couldn’t be differentiated while the other samples to create a database for reference could be differentiated. By comparing variation sites of 56 ITS2 sequences of doubtful fake Aconiti Kusnezoffi Radix Cocta samples and those of samples creating reference database,2,18,6 and 30 sequences consisted with Aconitum tatsienense,Aconitum soongaricum,Aconitum tanguticum,Aconitum carmichaelii and Aconitum kusnezoffii,respectively. Conclusion:The identification result indicated that adulteration or all false counterfeit of Aconiti Kusnezoffi Radix Cocta from exist in market. The origin of counterfeits were Aconitum tatsienense,Aconitum soongaricum and Aconitum tanguticum. And Aconitum soongaricum was the main counterfeit. DNA barcoding technology explores a new strategy for authentication of Aconiti Kusnezoffi Radix Cocta.

关 键 词：制草乌 聚合酶链式反应(PCR) 内部转录间隔区2(ITS2) 基原鉴定 植物鉴别 DNA条形码技术 

分 类 号：R917[医药卫生—药物分析学]