南方农业学报2013，Vol.44Issue(7)：1215-1219,5.DOI:10.3969/j:issn.2095-1191.2013.7.1215
禽流感病毒H1、H3亚型和M基因三重PCR检测方法的建立
Simultaneous detection of H1, H3 subtypes and M gene of avian influenza virus by multiplex PCR
摘要
Abstract
[Objective]A method of detecting avian influenza virus(AIV) and separating H1 and H3 subtype was determined to provide references for effectively preventing and controlling AI.[Method]Three sets of specific primers were designed according to the conserved sequences of AIV matix (M) gene,and the hemagglutinin (HA) genes of H1 and H3 subtype AIV.A multiplex PCR assay system was developed and optimized for rapid and simultaneous detection of these genes.The specificity and sensitivity of this method was evaluated.This method was used to test the AIV infection from clinical samples.[Result] It was shown that all samples containing H1 and H3 subtype AIV could be amplified into three specific bands by this multiplex PCR.The lengths of these bands were 302 bp H1 AIV HA,453 bp(M) and 626 bp(H3 AIV),respectively.Samples containing H1 or H3 subtype AIV could be amplified into two specific bands,which were 302 bp and 453 bp as well as 453 bp and 626 bp,respectively.Samples containing other subtypes AIV could be amplified into a 453 bp specific band.However,no specific band was amplified from other avian pathogenic virus.As little as 7.00 pg of H 1 subtype AIV,3.62 pg of M subtype and 20.60 pg of H3 subtype AIV could be detected by this multiplex PCR.Thirty-six clinical samples (36/100,36％) were found to be infected with AIV,of which 4 were H1 subtype AIV,9 were H3 subtype AIV and 23 were other type AIV.[Conclusion]Detecting H1,H3 and M subtypes of avian influenza by multiplex PCR was appropriate,rapid,sensitive and specific.关键词
禽流感病毒/H1亚型/H3亚型/M基因/三重PCRKey words
avian influenza virus / H1 subtype / H3 subtype / M gene / multiplex PCR分类
农业科技引用本文复制引用
郭捷,谢芝勋,罗思思,刘加波,谢丽基,庞耀珊,范晴,邓显文,谢志勤..禽流感病毒H1、H3亚型和M基因三重PCR检测方法的建立[J].南方农业学报,2013,44(7):1215-1219,5.基金项目
广西科技攻关项目(桂科重1222003-2-4,桂科攻10100014-5,桂科专项11-3)；广西特聘专家专项项目(2011B020) （桂科重1222003-2-4,桂科攻10100014-5,桂科专项11-3）
广西自然科学基金项目(2011GXNSFA018088) （2011GXNSFA018088）
