新型葡萄球菌肠毒素多重PCR检测方法的建立

Establishment of Multiplex PCR for Simultaneous Detection of New Entertoxin Genotypes of Staphylococcus

针对新发现的SEO、SEQ、SER、SEU等4个葡萄球菌肠毒素基因型,分析比对GenBank中报道的序列,设计特异性引物,进一步通过调节退火温度、模板浓度和多重PCR引物浓度等,优化了多重PCR反应体系,建立了同时检测4种金黄色葡萄球菌肠毒素的多重PCR检测方法。该方法可同时扩增出4种新的肠毒素基因,与其他基因型无交叉反应,对SEO、SEQ、SER、SEU检测的敏感性分别达到1.45×102、2.34×103、1.89×102、2.09×102 pg。可对新型葡萄球菌肠毒素进行快速、准确的检测与分型。

Our study designed 4 pairs of primers to detect 16 S rDNA and SE genes（SEO, SEQ, SER, SEU） based on bioinformaties analyzing, optimized conditions of PCR amplification and staphylococcal ge- nome extraction to establish a systematic approach to rapid and simultaneous detection of different geno- types of SEs by multiplex PCR method. The sensitivity and specificity of the method were evaluated with different entertoxin genotypes of Staphylococcus and other standard bacterial isolates. The results showed that the method was very specific and sensitive. The sensitivities for detecting SEO, SEQ, SER and SEU reached 1.45 ×102 , 2.34 × 103 , 1.89×102 and 2.09× 102 pg.The method can detect and type novel staphylo- coccal enterotoxins rapidly and accurately