沙门氏菌通用PCR快速检测试剂盒的研制与应用

Research and application on rapid detection kit of salmonella by universal PCR

根据沙门氏菌fimY基因建立了用于检测沙门氏菌的通用PCR方法。对收集的A-F群标准菌株和临床分离的60株沙门氏菌分离株和11种非沙门氏菌进行PCR检测,采用2.0%琼脂糖电泳进行检测,结果所有沙门氏菌均扩增出526bp的特异性条带,而非沙门氏菌均未扩增出任何条带。通过电泳判定结果,该法可检出扩增体系中93cfu的沙门氏菌,还可检出含3cfu的样品用BP预增菌或MM增菌后的菌液。说明该方法敏感性高、特异性强。采用直接菌体加入法、热裂解、反复冻融、CTAB碱裂解法和柱式试剂盒提取法分别提取核酸模板,结果CTAB法效果最好,但操作烦琐,而直接菌体加入法和热裂解法可以达到和柱式试剂盒同样的效果,且操作简便快速、经济、效果好,值得推广应用。

This study evaluated universal PCR to detect Salmonella species according to fimY gene.The specific primers to the fimY gene were used to amplify standard bacteria of A-F salmonella,60 salmonella separate strains and 11 non-salmonella species by PCR. Experiment results indicated that only Salmonella species generated the specific 526 bp DNA fragments on the 2.0% agarose gel and non- salmonella species controls gave negative results,demonstrating that the primers should be specific for Salmonella genus. The ...