摘要
采用TaqMan-MGB探针实时荧光PCR方法对太白贝母及其近缘种进行分子鉴定研究。提取太白贝母及其近缘种的DNA,依据ITS1区序列,利用MEGA7.0软件进行比较分析,找出太白贝母及其近缘种的变异位点,通过Primer Premier 6.0软件设计一对特异性引物和TaqMan-MGB探针。在Roche LightCycler 96系统中,该方法对太白贝母DNA模板的检测下限为0.002 39 ng·μL^(-1),最适Tm值区间为60与61℃。特异性鉴定研究表明,该方法对太白贝母有良好的特异性,能与暗紫贝母等其余13种不同贝母基原植物明显区分。TaqMan-MGB实时荧光PCR方法可实现对太白贝母及其近缘种的准确鉴定,为太白贝母资源的合理开发、中药材市场的管理和中药生产企业的原料监管提供技术支撑。
The molecular identification of Fritillaria taibaiensis and its relatives was studied by real-time PCR with a TaqMan-MGB probe. DNA was extracted from F. taibaiensis and its relatives. According to the sequence of ITS1 region, the mutation sites of F. taipaiensis and its related species were identified by MEGA7.0 software. The specific primers(a pair) and a TaqMan-MGB probe were designed by Primer Premier 6.0 software. In the Roche LightCycler 96 system, the lowest limit of detection for F. taipaiensis DNA template was 0.002 39 ng·μL^(-1), and the optimal Tm value range was 60 and 61 ℃. Specificity identification showed that the method had good specificity for F. taipaiensis, as it could be distinguished from other 13 different Fritillaria species including F. unibracteata.Since this method could accurately identify F. taipaiensis and its related species, it provides technical support for rational development of F. taipaiensis resources, management of Chinese medicinal market and supervision of raw materials in Chinese medicine manufacturing enterprises.
作者
张田
陈娇
蒋瑞平
邹萌
仰铁锤
付绍兵
周嘉裕
廖海
ZHANG Tian;CHEN Jiao;JIANG Rui-ping;ZOU Meng;YANG Tie-chui;FU Shao-bing;ZHOU Jia-yu;LIAO Hai(School of Life Science and Engineering,Southwest Jiaotong University,Chengdu 610031,China;Qinghai Lvkang Biological Development Co.,Ltd.,Xining 810003,China)
出处
《药学学报》
CAS
CSCD
北大核心
2021年第9期2577-2583,共7页
Acta Pharmaceutica Sinica
基金
国家自然科学基金青年科学基金项目(31500276)
四川省重点研发项目(2018SZ0061)
四川省科技计划项目(2021ZHFP0170)
四川省中医药管理局项目(2020JC0128,2021MS116)。
