耐热型产肠毒素大肠杆菌检测方法的建立及应用

Establishment and Preliminary Application of Detection of Heat-stable Enterotoxigenic Genes in Enterotoxigenic Escherichia coli

为建立检测产肠毒素大肠杆菌(ETEC)耐热型肠毒素STa和STb的纳米金PCR方法,根据ETEC耐热型肠毒素STa和STb的基因序列,用Primer 5.0软件设计了2对特异性引物,优化得到最佳反应条件,并进行灵敏度和特异性的测试.实验结果表明:本文所建立的体系均能扩增到目的条带,灵敏度为47.5 cfu/mL,且特异性良好;所建立的纳米金PCR方法在人工染菌实验中也获得了较高的灵敏度和特异性.

The present study developed a nanogold-assisted PCR to detect two heat-stable enterotoxigenic genes STa and STb in enterotoxigenic Escherichia coli（ ETEC）. According to genome sequences of STa and STb published in GenBank,two pairs of primer were designed using the software of primer 5. 0. The reaction conditions,including sensitivity and specificity were optimized and assessed. The results indicated that using the method established,we could obtain expected target DNA fragments with high specificity,and the minimum detectable level was 47. 5 cfu /mL. Furthermore,the method also revealed high sensitivity and specificity in artificial contamination experiments.