小鼠诺如病毒巢式RT-PCR检测方法的建立与应用

Establishment and application of nested RT-PCR for the detection of murine norovirus

本研究旨在利用巢式RT-PCR技术,建立高效检测小鼠诺如病毒(murine norovirus,MNV)的方法。通过Gen Bank上公布的一系列小鼠诺如病毒的基因序列,设计内、外各一对特异性引物,通过优化PCR反应条件,建立巢式RT-PCR检测方法,并对临床样品进行检测。结果显示,本研究建立的巢式PCR检测方法特异性好,最低检出量为1×102copies/L,高于使用参考引物RT-PCR的最低检出量1×105 copies/L,对临床样品检出率为73%,高于使用参考引物普通RT-PCR的检出率(60%),与RT-qPCR方法检出率一致。结果表明,本研究成功建立了一种检测小鼠诺如病毒的巢式RT-PCR方法,并能快速、高效地应用于临床样品的检测,为小鼠诺如病毒的诊断和检测提供了有力的手段。

Murine norovirus（MNV）is recognized as one of the most prevalent pathogens in laboratory mice,which causes significant effects on production of mice and scientific research.To improve and prevent this situation,establishment of a rapid and high sensitive detection method for MNV is necessary.According to a large number of genetically diverse MNV gene sequences,we designed two pairs of nested PCR primers,optimized PCR reaction conditions and detected the clinical samples.Our results suggested that the sensitivity of this method was high,the minimum detection level was 1×102copies/L and the detection rate of clinical samples was 73%,which was higher than the result of RT-PCR using one pair of referenced primers,1×105copies/L and 60%,respectively.Briefly,the established nested PCR detection method for MNV in this study showed a powerful mean for quick and efficient detection of MNV and for estimating the prevalence of MNV infection.