多重位点特异性PCR法同时鉴别牛膝和川牛膝配方颗粒  

Simultaneous identification of formula granules of Achyranthes bidentata and Cyathula officinalis by multiplex allele-specific polymerase chain reaction

作　　者：黄上书 罗宇琴 宋叶 范耀耀 雷欣荷 李国卫 陈向东 HUANG Shangshu;LUO Yuqin;SONG Ye;FAN Yaoyao;LEI Xinhe;LI Guowei;CHEN Xiangdong(Guangdong Yifang Pharmaceutical Co.,Ltd./Guangdong Provincial Key Laboratory of Traditional Chinese Medicine Formula Granules Enterprises,Foshan 528244,China;School of Chinese Materia Medica,Guangdong Pharmaceutical University/Key Laboratory of Digital Quality Evaluation of Traditional Chinese Medicine,National Administration of Traditional Chinese Medicin/Guangdong Provincial Traditional Chinese Medicine Quality Engineering and Technology Research Center,Guangzhou 510006,China)

机构地区：[1]广东一方制药有限公司/广东省中药配方颗粒企业重点实验室,广东佛山528244 [2]广东药科大学中药学院/国家中医药管理局中药数字化质量评价技术重点研究室/广东(省)教育厅高校中药质量工程技术研究中心,广东广州510006

出　　处：《广东药科大学学报》2023年第3期73-79,共7页Journal of Guangdong Pharmaceutical University

基　　金：佛山市核心技术攻关项目(1920001000378);中华人民共和国工业和信息化部2022年产业技术基础公共服务平台项目(2022-230-221)。

摘　　要：目的建立一种同时鉴别牛膝和川牛膝药材及配方颗粒的多重位点特异性PCR鉴别方法。方法通过比较牛膝属相近物种核基因ITS序列,设计牛膝和川牛膝的特异性引物,经过优化PCR反应体系及PCR反应条件,建立了牛膝和川牛膝配方颗粒的多重位点特异性PCR鉴别方法。结果在退火温度为58℃、循环次数为31时,牛膝和川牛膝配方颗粒分别可扩增出187 bp和164 bp的特异性条带。该方法对牛膝和川牛膝药材的掺伪检出限分别为1%、5%,对两者配方颗粒的掺伪检出限分别为10%、50%。结论所建立的多重位点特异性PCR鉴别可有效鉴别牛膝和川牛膝配方颗粒及其掺伪体系。Objective To establish a multiplex allele-specific polymerase chain reaction(PCR)identification method for the simultaneous identification of materials and formula granules of Achyranthes bidentata and Cyathula officinalis.Methods By comparing the ITS sequences of the nuclear genes of Achyranthes sp.and similar species,the specific primers were designed.After optimizing the PCR reaction system and PCR reaction conditions,the multiplex allele-specific PCR identification method of materials formula granules of Achyranthes bidentata and Cyathula officinalis was established.Results When the annealing temperature was 58℃and the number of cycles was 31,the 187 bp and 164 bp specific bands of formula granules of Achyranthes bidentata and Cyathula officinalis could be amplified,respectively.The adulteration detection limits of this method were 1%and 5%for Achyranthes bidentata and Cyathula officinalis medicinal materials,and the adulteration detection limits for the two formula granules were 10%and 50%,respectively.Conclusion The established multiplex allelespecific PCR method can effectively identify formula granules of Achyranthes bidentata and Cyathula officinalis and their adulteration systems.

关 键 词：牛膝 川牛膝 配方颗粒 多重位点特异性PCR 掺伪 鉴定 

分 类 号：R932[医药卫生—生药学]