菟丝子及混伪品的DNA条形码鉴定  

Identification of Cuscutae Semen and adulterants using DNA barcode

作　　者：秦新月 金钺 高志晖 魏建和 QIN Xinyue;JIN Yue;GAO Zhihui;WEI Jianhe(National Engineering Laboratory for Breeding of Endangered Medicinal Materials,Key Laboratory of Bioactive Substances and Resources Utilization of Chinese Herbal Medicine of Ministry of Education,Institute of Medicinal Plant Development,Chinese Academy of Medical Sciences and Peking Union Medical College,Beijing 100193,China;Key Laboratory of State Administration of Traditional Chinese Medicine for Agarwood Sustainable Utilization,Hainan Provincial Key Laboratory of Resources Conservation and Development of Southern Medicine,Hainan Branch of the Institute of Medicinal Plant Development,Chinese Academy of Medical Sciences and Peking Union Medical College,Haikou 570311,China)

机构地区：[1]中国医学科学院北京协和医学院药用植物研究所中草药物质基础与资源利用教育部重点实验室濒危药材繁育国家工程实验室,北京100193 [2]中国医学科学院北京协和医学院药用植物研究所海南分所海南省南药资源保护与开发重点实验室国家中医药管理局沉香可持续利用重点研究室,海南海口570311

出　　处：《中草药》2024年第17期5982-5990,共9页Chinese Traditional and Herbal Drugs

基　　金：国家重点研发计划项目“高品质中药材生态调控栽培技术研究”(2022YFC3501504);医科院重大协同创新项目“药用植物种质资源库建设”(2021-I2M-1-032)。

摘　　要：目的调研市售菟丝子Cuscutae Semen混杂情况,为菟丝子药材质量的提升和安全应用提供依据。方法收集市售菟丝子25份,对样品进行ITS2基因扩增并双向测序,利用HMMer注释方法获得ITS2序列,使用MEGA6.0进行遗传距离计算并构建邻接法(neighbor-joining,NJ)系统发育树;运用DNAsp5.0进行功能域检测和单倍型分析;比较菟丝子及杂质的二级结构差异;用R语言将所得序列转换成彩色条形码图片及二维码。结果通过序列分析、遗传距离分析、构建系统发育树、二级结构分析、功能域分析及单倍型分析可将菟丝子药材中不同科属及同一属的杂质准确区分;成功检测出25份菟丝子中分别含有藜种子、绿穗苋种子、虫实种子、铁苋菜种子、稗子、酸模叶蓼种子、大豆碎粒等1种或多种杂质。结论首次调研了市售菟丝子的混杂情况,表明其混杂种类多,且部分混杂种子有毒,为菟丝子相关标准制定提供了依据。此外,ITS2序列可准确鉴定菟丝子及其混伪品,为微小种子类药材鉴别提供了理论依据;与二维码技术结合可实现菟丝子监管的标准化和信息化。Objective To investigate the purity and species of adulterants in commercially available Tusizi(Cuscutae Semen)in order to provide a basis for the improvement of the quality of Cuscuta and its safe application..Methods Genomic DNA was extracted from a total of 25 Cuscutae Semen samples collected from markets.ITS2 sequences were amplified,sequenced and then annotated by HMMer.MEGA6.0 was used to calculate genetic distance and construct a NJ phylogenetic tree.The functional domain,haplotype and secondary structures of ITS2 sequences were analyzed using DNAsp5.0.The obtained sequences were transformed into chromatic barcode images and two-dimensional DNA barcodes using the R Program.Results The adulteration of commercially available Cuscutae Semen was serious.All Cuscuta samples contained at least one adulterant.Seeds of Chenopodium album,Amaranthus hybridus,Corispermum hyssopifolium,Acalypha australis,Echinochloa crus-galli,Persicaria lapathifolia and cracked soybean(Glycine max(Linn.)Merr.)were successfully identified in Cuscutae Semen as adulterants.Conclusion This study confirmed the serious adulteration of Cuscutae Semen and clarified the species of these adulterants.ITS2 sequence barcode is able to identify Cuscuta and other common adulterants,providing a method for identifying tiny seeds.In addition,a combination of ITS2 sequences and two-dimensional code technology enables standardization and informationization in the management of Cuscuta market.

关 键 词：菟丝子 混伪品 ITS2 彩色条形码 二维码 DNA条形码 

分 类 号：R286.12[医药卫生—中药学]