H9N2禽流感病毒反向遗传系统的构建和鉴定  

Construction and characterization of reverse genetic system for H9N2 avian influenza

作　　者：刘丽琦[1] 董婕[1] 薄洪[1] 温乐英[1] 辛丽[1] 张烨[1] 李梓[1] 董丽波[1] 王敏[1] 郭元吉[1] 舒跃龙[1] 

机构地区：[1]中国疾病预防控制中心病毒病预防控制所流感室、传染病预防控制国家重点实验室,北京100052

出　　处：《中华实验和临床病毒学杂志》2009年第1期41-43,共3页Chinese Journal of Experimental and Clinical Virology

基　　金：MOH/WHO H9N2合作课题（WP/2006/CHN/CSR/1.5/002） 感谢MOH/WHO H9N2合作课题的资助,感谢美国CDC惠赠的表达质粒.

摘　　要：目的建立H9N2亚型禽流感病毒反向遗传系统，为人禽流感疫苗研制以及传播和致病机制等方面的研究提供技术平台。方法使用RT-PCR方法获得禽流感H9N2亚型病毒A，Guangzhou／333／99（H9N2）的8条全长基因节段，然后克隆到双表达载体pCI-pol I中，获得H9N2禽流感病毒的8个基因节段的8质粒系统。将构建好的8质粒共转染293T细胞后，收获上清接种鸡胚，然后对鸡胚尿囊液进行鉴定；对拯救的病毒进行鉴定。结果8质粒系统转染293T细胞后可以成功拯救出H9N2禽流感病毒，血凝效价可达到2^9／50μl，生长特性与野生型病毒类似。结论成功建立了H9N2禽流感病毒反向遗传系统。Objective To provide a technology platform for vaccine development as well as the research on transmission and pathogenesis, the reverse genetic system for H9N2 avian influenza virus was established. Methods Eight full-length cDNAs of avian influenza virus A/Guangzhou/333/99 （H9N2）were amplified by RT- PCR and separately cloned into the transcription/expression vector, pCI-poll. The 8 plasmids DNA was cotransfected into 293T cell, the cell supernatant was collected and inoculated into embryonated eggs, the rescued virus from the allantoic fluid was identified by hemagglutinination assay. Results The avian influenza H9N2 virus was successfully rescued by 8 plasmids co-transfection in 293T cells. The hemagglutinination titer of the rescued virus is up to 2^9/50 μl and its growth curve remained relatively as to the wild-type virus. Conclusion The reverse genetic for avian influenza H9N2 subtype virus has been established successftdly.

关 键 词：禽流感 流感病毒A型 鸟 遗传学技术 

分 类 号：S852[农业科学—基础兽医学]