单核细胞增生性李斯特氏菌荧光定量PCR快速检测方法的建立及初步应用

Establishment and preliminary application of real-time fluorogenic quantitative PCR assay to rapid detection of Listeria monocytogenes

摘要: 以单核细胞增生性李斯特氏菌的hly基因为靶基因,用Primer 5.0设计了特异性兼并引物,建立荧光定量PCR检测方法。并用阳性质粒标准品、不同菌株对该方法的特异性、灵敏度进行了验证,且在人工污染鲜切甜瓜上对该方法进行了初步应用。结果表明,建立的检测方法特异性强、灵敏度高,对阳性质粒标准品的灵敏度为7.69×10copies/μL,对人工污染的鲜切甜瓜中单核细胞增生性李斯特氏菌的最低检出限为3.11×10cfu/mL。

Abstract: Hly gene of Listeria monocytogenes was using as the target gene, a pair of degenerate primers were designed with Primer 5.0, and the method of real-time fluorogenic quantitative PCR assay to detection of Listeria monocytogenes was established. The specificity and sensitivity of plasmid standard and different strains were verified, its preliminary application in Artificial inoculation fresh-cut melon was realized. The results showed that the method had good specificity and sensitivity, and sensitivity for plasmid standard was 7.69 ×10copies/μL, the minimum detection limit of artificial inoculation fresh-cut melon was 3.11×10cfu/mL.