中国动物检疫2016，Vol.33Issue(12)：85-88,4.DOI:10.3969/j.issn.1005-944X.2016.12.023
A型猪流感病毒TaqMan探针荧光RT-PCR检测方法的建立
Development of a Real-time RT-PCR Assay with TaqMan Probe for Detecting Swine Inlfuenza A Virus
摘要
Abstract
A Taqman probe-based real-time RT-PCR was developed with a pair of primers and a probe designed according to the conserved region of M gene sequence of swine influenza A virus(SIV). Results showed the assay was specific to detect subtype H1N1,H3N2 and H9N2 of swine influenza A virus and had no cross-reaction with classical swine fever virus(CSFV),porcine reproductive and respiratory syndrome virus(PRRSV),porcine epidemic diarrhea virus (PEDV)and transmissible gastroenteritis virus(TGEV). The real-time RT-PCR assay has a broad linear detection range for RNA standard control of M gene of SIV(SIV-M-RNA)from 3.8×101 copies/µL to 3.8×108 copies/µL,the standard curve equation was Y=-3.4365X+40.091,and the correlation coefficient of the assay was 0.999 8. This assay could detect 38 copies/µL of SIV-M-RNA at the lowest level. Three different concentrations of SIV-M-RNA were used to test the repeatability,and the coefficients of variation(CVs)of both inter-assay and intra-assay were less than 1.5%, showing good repeatability. 938 nasal swab samples were tested for SIV detection by the assay. No positivereaction was found for all 860 samples from imported pigs. 17 of 78 samples from domestic pigs were found to be positive for SIV detection. These results suggested that the newly established real-time RT-PCR would provide a rapid,sensitive and specific detection for swine influenza A virus.关键词
A型猪流感病毒/TaqMan探针/荧光RT-PCRKey words
swine influenza A virus/TaqMan probe/real-time RT-PCR分类
农业科技引用本文复制引用
王建华,董志珍,赵祥平,陈小金,肖妍,王玉玲,谭旭菲,赵丹,张俊哲,陈本龙,王乃福..A型猪流感病毒TaqMan探针荧光RT-PCR检测方法的建立[J].中国动物检疫,2016,33(12):85-88,4.基金项目
天津市科技支撑项目（13ZCZDNCO1300）；天津市滨海新区惠民项目 （）
