应用通用引物PCR法对葡萄球菌B、C1、C2、C3型肠毒素基因的鉴定与分型研究

Detection and Differentiation of Type B,C1,C2 and C3 Enterotoxigenic Staphylococci by the Universal Primers PCR

本实验采用通用引物P1、P2聚合酶链反应(PCR)法对产肠毒素葡萄球菌的entB、entC1、entC2、entC3基因进行了扩增,结果表明,含有这四种肠毒素基因的葡萄球菌都可以产生595bp的特异性扩增片段,其余菌扩增均为阴性;扩增产物分别用在产物内部只有单一识别位点的限制酶HinfⅠ、BclⅠ、MboⅡ和MboⅡ酶切后产生大小不同的片段,因而可以达到酶切分型的目的。经敏感性试验表明,该方法可以检出10~0个细菌。

One pair of synthetic universal primers targeting to internal regions of the toxin genes were used in a polymerase chain reaction(PCR) protocal to detect and differentiate genes for staphylococcal enterotoxins B,C1,C2 and C3.amplification results showed that all type B,C1,C2 and C3 enterotoxigenic staphylococcal producted specific 595 bp fragments,31 reference strains including common food-born pathogens bacteria were negative by the PCR.Amplification fragments from ent B,ent C1,entC2 and entC4 which contained single restriction endonuclease recognize site were cleaved,respectively,by endonuclease Hinf I,Bcl I,Mbo I and Mbo I to differentiate toxin gene types.Study of the detection sensitivity showed that 10°enterotoxigenic staphyloccal could be detected.