H4亚型和N2亚型禽流感病毒二重RT-PCR检测方法的建立

为建立一种简单、快速鉴别H4亚型和N2亚型禽流感病毒(AIV)的方法,针对H4亚型AIV HA基因和N2亚型AIV NA基因的保守区域,分别设计筛选出1对特异性引物,通过优化反应条件,建立了H4亚型和N2亚型AIV二重RT-PCR检测方法。该法可特异性扩增H4亚型和N2亚型AIV,对其他亚型AIV和常见禽病病原体无交叉反应;对H4亚型和N2亚型AIV的检测下限均为104拷贝/μL;对152份临床样品的检测结果与病毒分离鉴定结果一致。本研究建立的二重RT-PCR方法为H4亚型和N2亚型AIV的诊断提供了快速、特异、敏感和有效的检测方法。

Establishment of a Duplex RT-PCR Assay for Detection of H4 and N2 Subtype Avian Influenza Virus

In order to establish a simple and rapid method for detecting H4 and N2 subtype avian influenza virus(AIV),one pairs of specific primers were designed and screened respectively based on the conservative areas of HA and NA genes of the two subtypes of virus,then a duplex RT-PCR assay was established after optimization of reaction conditions,by which,H4 and N2 subtype AIV could be specifically amplified,while other subtypes AIV and the pathogens of common poultry diseases failed to be amplified;the detection limits for H4 and N2 subtype were all 104 copies/μL;the testing results of 152 clinical samples were consistent with that of virus isolation and identification.In short,the RT-PCR assay established was rapid,specific,sensitive and effective for detecting H4 and N2 subtype AIV.