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拉克罗斯病毒和雪靴野兔病毒双重RT-PCR方法的建立
2014-07-09 15:07:04 中国质量新闻网
    白红岩,孙肖红,林二妹,陈倩,甄维,徐宝梁

  中国检验检疫科学研究院,北京 100123

  摘要: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LAC 1.41×105copies/μl,SSH 5.6×104 copies/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。

  关键词:拉克罗斯病毒;雪靴野兔病毒;双重RT-PCR;检测;方法

  中图分类号:R521.3  文献标识码:B

  Development of duplex RT-PCR for La Crosse virus and

  Snowshoe hare virus

  BAI Hong-yan, SUN Xiao-hong, LIN Er-mei, CHEN Qian, ZHEN Wei, XUBao-liang

  Chinese Academy of Inspection and Quarantine, Beijing 100123,China

  Abstract: Objective  To develop a duplex RT-PCR method fordetecting of La Crosse virus(LAC) and snowshoe hare virus(SSH)simultaneously. Methods  Two sets of specific primers weredesigned for S gene of LAC and SSH, which were used in one PCRreaction. The PCR conditions were optimized. The specificity wasverified by using 8 kinds of viruses, including  KunjinVirus(KUNV)、 Yellow fever virus(YFV)、 Dengue virus(DENV)、 Japaneseencephalitis virus(JEV)、 Tahyna virus(TAHV)、 Barmah Forestvirus(BFV)、 Mayaro virus(MAYV) and Getah virus(GETV). Thesensitivity was detected by using different copy of viral RNA. Results  The specific PCR products of LAC and SSH wereamplified. The sensitivity was 1.41×105 copies/μl and 5.6×104copies/μl, respectively. None of other viruses was found positiveby duplex RT-PCR.  Conclusion  The duplex RT-PCR methodhad been developed, through which LAC and SSH could bedetected.

  Key words: LAC; SSH; Duplex RT-PCR; Detection; Method

  《中国国境卫生检疫杂志》2014年3期

  
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