食源性致病菌焦磷酸通用引物测序方法的建立

摘要

建立一种利用焦磷酸测序技术检测和鉴定沙门氏菌(Salmonellla)、单核细胞增生李斯特菌(Listeria monocytogenes)、金黄色葡萄球菌(Staphylococcus aureus)和志贺氏菌(Shigella)的方法。通过对4种菌16SrRNA的保守序列可变区的同源性分析，设计出一套适合焦磷酸测序的扩增引物和测序引物，通过可变区位点差异，利用一套通用引物同时鉴别4种食源性致病菌，测序结果与数据库比对判断细菌种属。结果表明：建立的检测方法可在4 h内完成对4种菌的鉴定，测序鉴定结果与国标的传统生化方法完全一致，焦磷酸测序技术可用于食源性致病菌的检测和鉴定。%The method of pyrosequencing technology for identification of Salmonellla spp, Listeria monocytogenes, Staphylococcus aureus and Shigella cereus was established. According to nucleotide homology analysis of 16S rRNA of 4 food-borne pathogenic bacteria,we designed one set of primer,including one universal PCR primers and one pyrosequencing sequencing primer. The 4 pathogenic bacteria were distinguished on the basis of pyrosequencing signal peaks. The result showed that the identification of four-kinds of food-borne pathogenic bacteria was completed in 4 h, the results of identification of bacteria were consistent with the result of GB/T method. The pyrosequencing method can use for detection and identification of food-borne pathogenic bacteria.