Objective To establish a multiplex PCR method for the detection of minute virus of mice

(MVM) , mouse hepatitis virus (MHV) and murine norovirus (MNV) . Method Specific primers were

designed according to the application software of MVM, MHV and MNV gene conserved regions. By

optimizing PCR amplification conditions, a multiplex PCR assay was established to detect MVM, MHV

and MNV simultaneously. The specificity, sensitivity and repeatability of the assay were verified, and the

preliminary clinical application of the assay was carried out. Result The multi-PCR method was

successfully established by optimizing the PCR amplification conditions. The specificity test result showed

that the method could simultaneously amplify the target fragments of MVM, MHV and MNV, and the

Sensitivity test result showed that the lower limits of detection of MVM, MHV and MNV were 10  , 10 

and 10  copies/ μL, respectively. Repeatability test shows that the proposed method has good

repeatability. The method was used to detect 287 clinical samples, and the coincidence rate of detection

result with single PCR result reached 100%. Conclusion The established multiplex PCR method for detection of MVM, MHV and MNV viruses has the characteristics of high specificity, high sensitivity and excellent repeatability, which is suitable for clinical sample detection of the infection of the three pathogens and epidemiological investigation of a large number of samples.

minute virus of mice, mouse hepatitis virus, mouse norovirus, multiplex PCR

目的 建立一种同时检测小鼠细小病毒(MVM) 、肝炎病毒(MHV)和诺如病毒(MNV)的多重 PCR 检测方法。

方法 分别根据 MVM、MHV 和 MNV 基因保守区应用软件设计特异性引物,构建在同一反应体系中可同时检测

MVM、MHV 和 MNV 的多重 PCR 检测方法,验证其特异性、敏感性及重复性,并进行初步检测应用。 结果 通过优

化 PCR 扩增条件成功建立多重 PCR 检测方法,特异性实验结果表明,该方法可同时扩增 MVM、MHV 和 MNV 的目

的片段,片段大小与预期相符,未出现非特异性片段;敏感性测试显示,MVM、MHV 和 MNV 三种病毒的最低检测下

限为 10 2 、10 4 、10 2 copies/ μL;重复性实验显示该方法在不同时间、不同场所应用的重复性稳定。 应用该方法对 287

份小鼠结肠匀浆样品进行检测,与单重 PCR 检测结果完全一致。 结论 所建立的 MVM、MHV 和 MNV 三种病毒多

重 PCR 检测方法具有特异性高、敏感性强和重复性优的特点,适用于三种病原体感染的临床样品检测和大量样品

的流行病学调查。

小鼠细小病毒, 小鼠肝炎病毒, 小鼠诺如病毒, 多重 PCR 检测方法