摘要
目的通过DNA条形码技术和建立特异性聚合酶链式反应(PCR)方法,有效鉴别九味羌活丸中的水稻源性成分掺伪问题。方法通过建立特异性PCR技术对九味羌活丸中掺伪的水稻源性成分样品的GOS9序列进行扩增,并对PCR反应体系与程序、方法学和系统适用性进行考察,从而对掺伪水稻源性成分进行鉴别。结果该引物体系只针对水稻源性成分DNA进行扩增,与中成药中其他药味的DNA均无交叉扩增,最低能检测到1/1000掺伪的水稻源性成分DNA。将其应用于3厂家8批次样品中检测,以2%琼脂糖凝胶电泳检测PCR产物,掺伪水稻源性成分的批次在68 bp附近处有一清晰条带,而阴性样品无条带。结论通过特异性引物PCR扩增、电泳检测和克隆测序的方法,可以有效鉴别九味羌活丸中的水稻源性成分掺伪问题。该方法具有快速、特异、灵敏的优点,为水稻源性成分掺伪的分子生物学鉴定方法提供了参考。
OBJECTIVE To effectively identify the adulteration of rice-derived ingredients in Jiuwei Qianghuo pills through DNA barcoding method and specific PCR.METHODS The GOS9 sequence of rice-derived ingredients adulterated in Jiuwei Qianghuo pills was amplified by specific PCR,The PCR reaction system,procedure,methodology and applicability were investigated,so as to identify the adulterated rice-derived ingredients.RESULTS The primer system could only amplify rice-derived DNA,and there was no cross amplification with DNA of other traditional Chinese medicine ingredients in Chinese patent medicines.This method can detect at least 1/1000 adulterated rice-derived DNA.The method was applied to 8 batches of samples from 3 manufacturers.The PCR product was detected by 2%agarose gel electrophoresis.The batches of adulterated rice-derived ingredients had a clear band near 68BP,while negative samples had no bands.CONCLUSION The adulteration of rice-derived ingredients in Jiuwei Qianghuo pills can be effectively identified by PCR amplification through specific PCR,electrophoresis detection and cloning and sequencing.This method is rapid,specific and sensitive,which provides a reference for molecular biological identification of adulteration of ricederived ingredients.
作者
刘晶晶
王菲菲
任秀
崔生辉
戴忠
马双成
LIU Jing-jing;WANG Fei-fei;REN Xiu;CUI Sheng-hui;DAI Zhong;MA Shuang-cheng(National Institutes for Food and Drug Control,Beijing 100050,China)
出处
《中国药学杂志》
CAS
CSCD
北大核心
2021年第23期1894-1898,共5页
Chinese Pharmaceutical Journal
关键词
九味羌活丸
聚合酶链式反应
分子生物学鉴定
特异性扩增
水稻源性成分掺伪
Jiuwei Qianghuo pill
polymerase chain reaction
molecular biological identification
specific amplification
rice-derived ingredients adulteration
