中国中药杂志

2017, v.42(13) 2484-2491

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金银花配方颗粒的位点特异性PCR鉴别研究
Molecular authentication of Jinyinhua formula granule by using allele-specific PCR

蒋超;屠李婵;袁媛;黄璐琦;高伟;金艳;
JIANG Chao;TU Li-chan;YUAN Yuan;HUANG Lu-qi;GAO Wei;JIN Yan;State Key Laboratory of Dao-di Herbs Breeding Base,National Resource Center for Chinese Materia Medica,China Academy of Chinese Medical Sciences;School of Traditional Chinese Medicine,Capital Medical University;

摘要(Abstract):

中药配方颗粒已失去所有形态学辨识特征,传统鉴定方法无法有效鉴定其真伪。该文使用位点特异性PCR鉴别技术,根据金银花trnL-trnF的一个SNP位点设计特异性鉴别引物,对金银花基原植物及配方颗粒进行鉴定。经过优化DNA提取方法后,药材与配方颗粒均成功提取到DNA,金银花基原植物及配方颗粒均可扩增获得约110 bp的条带,混伪品无条带。且BLAST结果比对证明PCR产物序列为金银花trnL-trnF序列。该文研究结果表明,DNA分子鉴定方法可弥补性状、显微鉴别局限性,对中药配方颗粒真实性进行快速准确的鉴定,为其生产、流通、用药安全提供科学依据和保障。
Traditional authentication method is hard to identify herb's authenticity of traditional Chinese medicine(TCM) formula granules because they have lost all their morphological characteristics. In this study,a new allele-specific PCR method was established for identifying the authentication of Jinyinhua formula granule( made fromLonicerae Japonicae Flos) based on an SNP site in trnL-trnF fragment. Genomic DNA was successfully extracted fromLonicerae Japonicae Flos and its formula granules by using an improved spin column method and then PCR was performed with the designed primer. Approximately 110 bp specific bands was obtained only in the authentic Lonicerae Japonicae Flos and its formula granules,while no bands were found in fake mixed products. In addition,the PCR product sequence was proved fromLonicerae Japonicae Flos trnL-trnF sequence by using BLAST method. Therefore,DNA molecular authentication method could make up the limitations of character identification method and microscopic identification,and quickly identify herb's authenticity of TCM formula granules,with enormous potential for market supervision and quality control.

关键词(KeyWords): 金银花;配方颗粒;位点特异性PCR;分子鉴定
Lonicerae Japonicae Flos;formula granule;allele-specific PCR;molecular authentication

Abstract:

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基金项目(Foundation): 国家杰出青年科学基金项目(81325023);; 中央级公益性科研院所基本科研业务费专项(ZZ10-008);; 中医药行业科研专项(201407003)

作者(Author): 蒋超;屠李婵;袁媛;黄璐琦;高伟;金艳;
JIANG Chao;TU Li-chan;YUAN Yuan;HUANG Lu-qi;GAO Wei;JIN Yan;State Key Laboratory of Dao-di Herbs Breeding Base,National Resource Center for Chinese Materia Medica,China Academy of Chinese Medical Sciences;School of Traditional Chinese Medicine,Capital Medical University;

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