摘要
根据GenBank中H6、N1亚型禽流感病毒(AIV)HA、NA基因,分别设计并筛选出2对特异性引物,用于H6亚型AIV和N1亚型AIV的检测,优化反应条件,建立了H6N1亚型AIV的二重RT-PCR检测方法。对该法进行特异性、敏感性检验,并用该法对临床样品进行检测。所建立的方法对H6N1亚型AIV可特异性扩增出447 bp(H6亚型)和325 bp(N1亚型)目的条带,对H6Ny亚型AIV仅扩增出447 bp目的条带,对HXN1亚型AIV仅扩增出325 bp目的条带,对常见禽病病原体均未扩增出任何条带;该法对H6N1亚型AIV检测下限为5×102拷贝/μL;341份临床样品检测结果与病毒分离鉴定一致。本研究所建立的H6N1亚型AIV RT-PCR特异性强、灵敏度高,一管可同时检测H6和N1亚型AIV,为H6N1亚型AIV的检测提供一种简便、快速和有效的检测方法。
According to the sequences of HA and NA genes of H6N1 avian influenza virus （AIV） in GenBank, two pairs of specific prim- ers were designed and used for detection of H6 and N1 subtype AIV, respectively, and the concentrations of primers and reaction conditions were optimized to develop a duplex RT-PCR assay for detection H6N1. Specificity analysis showed that two specific bands could be amplified from H6N1 AIV by this RT-PCR, and the lengths of bands were 447 bp （H6-AIV） and 325 bp （ N1 -AIV）, respectively. No specific band was amplified from other subtypes of AIV or avian pathogenic virus. The limit of detection for H6N1 AIV was 5xl02copies/μL. In the detection of 341 clinical samples, the results of this assay were accorded with the viral isolation completely. It suggests that this duplex RT-PCR assay is a specific and sensitive method for the detection of H6 and N1 subtypes of AIV, and can be applied for rapid differential diagnosis for clinical samples.
出处
《畜牧与兽医》
北大核心
2015年第8期24-27,共4页
Animal Husbandry & Veterinary Medicine
基金
广西特聘专家专项(2011B020)
广西科技攻关重大专项(14121003-4-2和1222003-2-4)
