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目的建立针对拉克罗斯病毒(LAC)和雪靴野兔病毒(SSH)的双重RT-PCR检测方法。方法分别针对LAC和SSH病毒的S基因设计2对特异性引物,建立同时检测LAC和SSH的双重RT-PCR方法。以黄病毒属的黄热病毒(YFV)、库京病毒(KUNV)、登革病毒(DENV)、乙型脑炎病毒(JEV),布尼亚病毒属的塔希纳病毒(TAHV)和甲病毒属的巴马哈森林病毒(BFV)、玛雅罗病毒(MAYV)、盖塔病毒(GETV)为模板验证方法的特异性,以不同拷贝数的病毒RNA检测该方法的敏感性。结果建立的双重RT-PCR方法能同时扩增到LAC和SSH的目的片段;敏感度分别达到LAC1.41×105copies/μl,SSH 5.6×104copies/μl;与YFV、KUNV、BFV、MAYV无交叉反应。结论建立了双重RT-PCR方法,可用于LAC和SSH病毒的快速检测。
Objective To establish a dual RT-PCR assay for lacrosavirus (LAC) and snowshoe rabbit virus (SSH). Methods Two pairs of specific primers were designed for the S gene of LAC and SSH viruses, respectively, and a dual RT-PCR method for simultaneous detection of LAC and SSH was established. In the case of flavivirus-containing yellow fever virus (YFV), Kurume virus (KUNV), dengue virus (DENV), Japanese encephalitis virus (JEV), Tahenavirus (Bunny’s virus) Virulence of the Bamaha forest virus (BFV), Mayarrow virus (MAYV), and Geta virus (GETV) as template-specific methods of detection, with different copy number of viral RNA to detect the sensitivity of the method. Results The double RT-PCR method could amplify the target fragments of LAC and SSH simultaneously. The sensitivity was up to LAC1.41 × 105copies / μl and SSH 5.6 × 104copies / μl respectively. There was no cross reaction with YFV, KUNV, BFV and MAYV . Conclusion A dual RT-PCR method was established for the rapid detection of LAC and SSH viruses.