Taqman荧光实时PCR快速检测原料乳中EPEC

Taqman fluorescent quantitative PCR detection of EPEC in raw milk

建立可对原料乳中肠致病性大肠埃希氏菌(enteropathogenic Escherichia coli,EPEC)进行快速检测的Taqman荧光实时PCR方法。以eae致病基因为靶基因,用特异性引物和Taqman探针进行实时PCR扩增,研究反应的特异性和检测限,并用所建立的方法对16份市售原料乳进行EPEC检测。结果表明,所用引物和探针可高效扩增出目的片段,与原料乳中其他常见致病菌无交叉反应,经2h增菌后检测限为8.8mL-1。对16份市售原料乳样品,检出2份含有EPEC,血清型分别为O111:K58(B4)和O127a:K63(B18)。全部检测过程只需5h,可用于原料乳中EPEC的快速检测和污染调查。

To establish a fluorescent quantitative PCR. method for detecing enteropathogenic Escherichia coli （EPEC） in raw milk. Use the specific primers and Taqman probe to amplify the target pathogenic gene eae, and study on the specificity and sensitivity, and detect EPEC in 16 raw milk samples in Harbin with this method. The results showed that it can amplify the 102bp target fragment in eae gene efficiently and do not cross-react with other food-bom~ pathogens. The sensitivity of detection of artificially milk is 8.8 mL^-1 after being cultivated 2h. Among the 16 raw milk samples detectecd, 2 samples contained EPEC. The detection can be carried out within 5h.