Zhang Qingshan, Wan Chunhe, Li Chenxi, Bai Xiaofei, Liu Ming, Liu Siguo, Zhang Yun
State Key Laboratory of Veterinary Biotechnology, Harbin Veterinary Research Institute of Chinese Academy of Agricultural Sciences, Harbin 150001, PR China.
Institute of Animal Husbandry and Veterinary Medicine, Fujian Academy of Agricultural Sciences, Fuzhou 350013, China.
J Vet Med Sci. 2017 Dec 22;79(12):2057-2062. doi: 10.1292/jvms.17-0227. Epub 2017 Aug 5.
To establish an accurate, rapid, and a quantifiable method for the detection of Riemerella anatipestifer infection, a widespread infectious disease in birds, we developed a TaqMan-based real-time PCR assay by using DtxR gene-specific primers and a TaqMan probe. The standard curve established with a linear correlation (R) of 0.998 and efficiency of 99% between the Ct value and the logarithm of the plasmid copy number. The reproducibility and specificity of the real-time PCR assay were confirmed by using plasmids containing DtxR genes or DNAs extracted from well-known bacteria or viruses causing duck diseases. The real-time PCR assay was 100 times more sensitive than the conventional PCR. The results reveal that the established real-time PCR assay might be a useful method for diagnosis and quantitative detection of Riemerella anatipestifer in birds.
为建立一种准确、快速且可定量检测鸭疫里默氏菌感染（一种禽类广泛传播的传染病）的方法，我们使用DtxR基因特异性引物和TaqMan探针开发了一种基于TaqMan的实时荧光定量PCR检测方法。所建立的标准曲线在Ct值与质粒拷贝数的对数之间具有0.998的线性相关性（R）和99%的效率。通过使用含有DtxR基因的质粒或从引起鸭病的知名细菌或病毒中提取的DNA，证实了实时荧光定量PCR检测方法的重复性和特异性。实时荧光定量PCR检测方法比传统PCR灵敏100倍。结果表明，所建立的实时荧光定量PCR检测方法可能是诊断和定量检测禽类鸭疫里默氏菌的一种有用方法。
