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首页> 外文期刊>Archiv fur Geflugelkunde >Development of duplex and triplex conventional RT-PCR for the detection of H5N1 avian influenza virus
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Development of duplex and triplex conventional RT-PCR for the detection of H5N1 avian influenza virus

机译：双和三层传统的发展rt - pcr检测H5N1禽流感病毒

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摘要

Molecular diagnostic tests are commonly used to diagnose avian influenza virus because they are sensitive, rapid and cost effective. However, continuous shift and drift of avian influenza viruses and H5N1 in particular, is a real challenge for development of specific genetic based detection techniques. Furthermore, most of current molecular diagnostic commercial kits rely on the detection of H5 and/or Nl genes without the presence of the Ml gene which represents the most conservative gene in the typeA avian influenza viruses (AIV). This enable spread of undetected mutants. In the present investigation we have optimized a primer set for polymerase chain reaction (PCR)-based detection of influenza A viruses H5, Nl and Ml genes that was validated witha panel of influenza A virus reference strains representing H5, H7, H9 and H13. Specificity test was carried out by the use of eight type A AIV subtypes (H5N1, H5N2, H5N9, H7N1, H7N3, H7N7, H9N2 and H13N6). Results showed that this protocol is capable of produce two distinguished bands represents the H5 and Nl genes in the duplex format. While, it generated three distinguished bands represents the H5, Nl and Ml genes (in the triplex format). Moreover, the specificity test showed that the used primers do not have cross-reactivity with the AIV subtypes other than H5N1. In addition, Sensitivity test revealed that the duplex format has a similar sensitivity limit as the triplex one.
机译：分子诊断测试常用的因为他们是诊断禽流感病毒敏感、快速、成本效益。连续转变和禽流感的漂移尤其是病毒和H5N1,是真实的挑战的发展特定的基因基于检测技术。当前分子诊断商业工具的依赖基因的检测H5和/或问代表了Ml基因的存在最保守的基因typeA禽流感禽流感病毒(AIV)。未被发现的突变体。我们已经优化引物聚合酶流感的连锁反应(PCR)的检测H5病毒,问和Ml基因进行验证与甲型流感病毒的小组参考代表H5毒株,H7, H9和H13。特异性测试是由使用八个类型AIV亚型(H5N1, H5N2 H5N9,H7N1 H7N3、H7N7 H9N2和H13N6)。这个协议能够产生两个著名的乐队代表了H5和问基因双格式。三个杰出的乐队代表了H5,问和Ml基因(三层格式)。引物特异性测试表明,使用没有大的AIV吗除了H5N1亚型。敏感性试验表明,双格式也有类似的灵敏度极限的三层一个。

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