口蹄疫、水泡性口炎和猪水泡病多重荧光定量RT-PCR检测方法的建立

Establishment of simultaneous detection of foot-and-mouth disease virus, ve-sicular stomatitis virus and swine vesicular disease virus by multiplex real-time RT-PCR

中文关键词:  口蹄疫病毒  水泡型口炎病毒  猪水泡病病毒  多重荧光定量RT-PCR

英文关键词:foot and mouth disease virus  vesicular stomatitis virus  swine vesicular disease virus  multiplex real-time RT-PCR

中文摘要:

      目的 建立同时检测口蹄疫病毒、水泡性口炎病毒和猪水泡病病毒的多重荧光RT-PCR检测方法。方法 根据口蹄疫病毒3D蛋白编码基因、水泡性口炎病毒N蛋白编码基因和猪水泡病病毒VP1蛋白编码基因的高保守区设计特异性引物和探针, 对3种动物病毒进行多重荧光定量RT-PCR扩增。结果 经过扩增, 可以同时检测口蹄疫病毒、水泡性口炎病毒和猪水泡病病毒, 而其他参试病原均无扩增信号, 显示其良好的特异性。对口蹄疫病毒、水泡性口炎病毒、猪水泡病病毒的最低检测限分别达到101、102、102个质粒拷贝浓度。结论 本方法灵敏度高, 特异性良好, 可实现多种病毒混合感染的同时检测。

英文摘要:

      Objective To establish a multiplex real-time RT-PCR method for simultaneous detection of foot and mouth disease virus, vesicular stomatitis virus and swine vesicular disease virus. Methods According to the gene encoding 3D protein of foot-and-mouth disease virus, vesicular stomatitis virus N protein coding gene and swine vesicular disease virus VP1 protein encoding gene, specific primers and probe were designed, and 3 kinds of animal viruses were amplified by multiplex real-time RT-PCR. Results The FMDV, VSV and SVDV could be simultaneously detected after amplification, and other tested pathogens were no amplification signal, displaying the good specificity. The minimum detection limits of foot and mouth dis-ease virus, vesicular stomatitis virus and swine vesicular disease virus were 101, 102, and 102 plasmid copy concentration, respectively. Conclusion This method has a good specificity and sensitivity for simultaneous detection of FMDV, VSV and SVDV at the same time.