口蹄疫病毒RT-LAMP检测方法的建立

Establishment of RT-LAMP for Rapid Detection of Foot-and-Mouth Disease Virus

利用逆转录环介导等温核酸扩增技术(RT-LAMP),建立了口蹄疫病毒快速检测方法,同时评价了该方法的灵敏性和特异性。结果表明,根据口蹄疫病毒多聚蛋白基因保守区段设计的LAMP引物能够在65℃下,1 h内实现目标核酸区段的大量扩增,检测结果可直接用肉眼判断。该检测体系具有极高的特异性,只能检测到目标病毒,与其他类似病毒如猪水泡病病毒、猪瘟病毒、猪细小病毒等无交叉反应,可检测到10-5稀释度的目标病毒核酸量,比普通RT-PCR的灵敏性高100倍,比荧光PCR高10倍。

A rapid detection of foot-and-mouth disease virus （FMDV） was established by using reverse transcription loop-mediated isothermal amplification （RT-LAMP） method, meanwhile its specificity and sensitivity were assessed. The results showed that the FMDV RNA could be amplified by incubation at 65℃ for only lh using six primers designed based on FMDV polyprotein gene and the amplification products could be detected easily by naked-eye. There is no cross reaction with other virus such as SVDV.SFV and PPV by detecting their RNA samples. The detection limit of this method was found to be 10^-5 dilution of RNA sample which was 100-fold higher than that of PCR and 10-fold higher than real-time PCR.