多重RT-PCR检测猪口蹄疫病毒、猪水泡病病毒和猪水疱性口炎病毒的研究

建立了一种同时检测猪口蹄疫病毒(FMDV)、猪水泡病病毒(SVDV)和猪水疱性口炎病毒(VSV)三种病原体的多重RT-PCR方法。参照文献报道的基因序列,设计合成了三对特异性引物;PCR扩增条件进行优化后,用这三对引物对同一样品中的FMDV、SVDV、VSVRNA模板进行扩增,结果同时得到了三条特异性条带,大小与试验设计相符:FMDV(208bp)、SVDV(862bp)、VSV(638bp),且对猪瘟病毒(CSFV)、猪繁殖与呼吸综合症病毒(PRRSV)和猪传染性胃肠炎病毒(TGEV)核酸扩增结果为阴性;三种病毒RNA模板检出的最小量均为10fg。试验证明,此方法经济、快速、敏感、特异,可用于FMDV、SVDV和VSV这三种猪水泡性疾病的鉴别诊断及流行病学调查。

Simultaneous Detection of Foot-and-mouth Disease Virus, Swine Vesicular Disease Virus and Vesicular Stomatitis Virus by Multiplex RT-PCR

A multiplex RT-PCR was optimized to simultaneously detect foot-and-mouth disease virus (FMDV), swine vesicular disease virus (SVDV) and vesicular stomatitis virus (VSV). Three specific primer sets were selected according to FMDV, SVDV, and VSV gene sequences reported. The results showed that all the samples which contained FMDV, SVDV and VSV could be amplified by this method with optimized factor of reaction. Three specific bands of FMDV 208 bp, and SVDV 862 bp, and VSV 638 bp, as low as 10fg of FMDV-RNA, SVDV-RNA, and VSV-RNA could be detected and visualized under UV light, but not the gene fragments of PRRSV,TGEV and CSFV. The multiplex PCR appears to be rapid, sensitive and specific method for detecting FMDV, SVDV and VSV simultaneously, and may be applied to clinical diagnosis and epidemiological investigation.