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目的:建立Taqman荧光PCR猪源性成分检测技术,并将其应用于地沟油检测。方法:利用直接沉淀DNA法提取油样DNA,以猪线粒体细胞色素b基因序列设计特异的引物和探针,在对引物和探针进行特异性和敏感性评价的基础上建立Taqman探针PCR方法,对地沟油、猪油、市售可疑油脂等油样进行检测。结果:该系统能够对目的基因进行特异性扩增,不与其他物种发生交叉反应,能够定量检出0.001ng的模板;可以检测出地沟油、猪油中的猪源性成分,并经动物源性基因PCR鉴定确证阳性结果的可靠性。结论:本研究所采用的Taqman荧光PCR技术可以扩增出地沟油中的猪线粒体细胞色素b(Cyt b)基因,得到阳性的结果,将可用于地沟油的检测。
OBJECTIVE: To establish a Taqman fluorescence PCR-based porcine source detection technology and apply it to the detection of waste oil. Methods: The direct DNA sample was used to extract the oil sample DNA. The primers and probes were designed based on the sequence of porcine mitochondrial cytochrome b gene. Based on the specificity and sensitivity of primers and probes, a Taqman PCR , On the waste oil, lard, commercially available oil and other suspicious oil testing. Results: The system can specifically amplify the target gene without cross-reaction with other species, and can quantitatively detect the template of 0.001 ng. It can detect the pig-derived components in waste oil and lard, Genotyping PCR confirms the reliability of the positive result. Conclusion: The Taqman fluorescent PCR technique used in this study can amplify the Cyt b gene of porcine mitochondria, which has positive result and will be used for the detection of genistein oil.