摘要
针对H7N9亚型欧亚谱系AIV的血凝素(HA)基因和神经氨酸酶(NA)基因的序列保守区分别设计特异性引物和TaqMan探针。通过优化反应条件,建立了H7N9亚型禽流感病毒实时荧光定量RT-PCR检测方法。利用该方法对泰安市活禽市场11个月的空气状况进行监测。结果显示:H7和N9基因的最低检测限度分别为100copies/μL和101 copies/μL,检测H7和N9基因的R2值均为0.999。批内和批间试验的变异系数均小于3%。采用该技术,活禽市场空气样品3/114份检测出H7N9阳性。结果表明:本试验建立的H7N9亚型AIV实时荧光定量RT-PCR方法与其他方法相比具有快速、特异和敏感的优点,可为H7N9亚型AIV的监测及早期诊断提供理论依据。
The aim of this study was to develope a rapid,sensitive and specific real-time RT- PCR method for detection of H7N9 subtype avian influenza virus（AIV）. Two pairs of primers and probes specific to the hemagglutinin（HA） gene and neuraminidase（NA） gene were designed ac- cording to the HA gene sequences of H7 subtype and the NA gene of N9 AIV available the Eura- sian lineage in GenBank, respectively. The reaction conditions were optimized, the real-time RT- PCR assay of HTN9 subtype was edtablished. Eleven months air samples of live poultry markets were supervised with this method. The results showed that the detection limit of H7 was 10^0 cop- ies/μL and N9 was 10^1 copies/μL,the Re index of HTN9 subtype virus displayed was 0. 999. The intra- and inter- assay coefficients of variation were all less than 3%. Furthermore,11 months air samples were tested,and out of 114,3 positive air samples for HTN9 AIV were identified by the established assay, other months is negative. The established H7N9 subtype AIV real-time RT- PCR method,compared with other method, is a rapid, specific and sensitive method for surveil- lance and early diagnosis of H7N9 subtype AIV.
出处
《中国兽医学报》
CAS
CSCD
北大核心
2017年第1期78-82,96,共6页
Chinese Journal of Veterinary Science
基金
国家自然科学基金资助项目(31470258)
国家病原微生物重点实验室开放基金项目(SKLPBS1449)
国家环境保护环境微生物利用与安全控制重点实验室年开放基金资助项目(SMARC2013D001)
