垂体后叶粉中3种动物源性成分多重real-time PCR检测方法的建立及应用

Establishment and application of multiplex real-time PCR method for rapid detection of three animal-derived components in powdered posterior pituitary

目的基于三重实时聚合酶链式反应(real-time polymerase chain reaction,real-time PCR)技术同时快速检测垂体后叶粉中的猪、牛、羊性动物源性成分。方法采用CTAB法提取样品及对照品DNA,以线粒体基因为靶基因设计并合成三重real-time PCR的引物探针,并对建立方法进行验证,用于检测垂体后叶粉样品是否与标识一致。结果猪、牛、羊源性成分的最低检测浓度为10 pg/μL,采用建立的方法检测3批垂体后叶粉样品,均只检测出猪源性成分。结论建立的方法可用于同时检测垂体后叶粉中的猪、牛、羊源性成分,可为相关工作提供技术支持。

Objective To determine the porcine,bovine and sheep derived components in powdered posterior pituitary by a multiplex real-time polymerase chain reaction(PCR)method.Methods The DNA was extracted by CTAB method.The mitochondria genes were used as target genes to design specific primers and Taq Man probes.The developed method was validated and applied to detect whether the animal-derived components in the samples were consistent with those indicated.Results The minimum detection concentration was 10 pg/μL of template DNA.The 3 batches of powdered posterior pituitary samples were identified by the successfully established method,and only pig source components were detected.Conclusion The multiplex real-time PCR method can be used for the detection of porcine,bovine and sheep derived components in powdered posterior pituitary.