锦鲤疱疹病毒TaqMan荧光定量PCR快速检测方法的建立及应用

摘要

To establish a rapid and effective quarantine method of the Kio Hepesvinis disease, a set of primers and TaqMan probe for Fluorescent Quantitation PCR were designed to detect the conservative sequence of Kio Hepesvirus Sph genes. The cell cultures were detected by using the established quantitative PCR assay, and the results were compared with those of the routine PCR. The quantitative PCR sensitivity was higher than that of the routine PCR. The quantity of KHV DNA was 1.6×102 copies/μL. A reliable diagnostic result can be obtained just within 4 h. The assay proved to be a rapid, sensitive, specific and repetitive method for rapid detection of Kio Hepesvirus from fish in quarantine.%为建立快速有效的锦鲤疱疹病毒病检疫方法,根据锦鲤疱疹病毒(KHV)聚合酶基因(Sph)的保守序列设计1对引物和相应的TaqMan探针,建立了一种快速检测锦鲤疱疹病毒病荧光PCR方法.用建立的检测方法对细胞培养物进行检测,并与常规PCR对比,结果荧光PCR灵敏度高于普通PCR,能检出的最低拷贝数为1.6×102/μL.应用该方法对样品进行检测,结果表明:所建立的荧光PCR检测方法4h内可报告检测结果.该方法具有快速、灵敏、特异及重复性好等特点,适用于锦鲤疱疹病毒的快速检疫.