CAJ | 학술논문

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29/50μl,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.
목적 건립H9N2아형금류감병독반향유전계통,위인금류감역묘연제이급전파화치병궤제등방면적연구제공기술평태.방법 사용RT-PCR방법획득금류감H9N2아형병독A/Guangzhou/333/99(H9N2)적8조전장기인절단,연후극륭도쌍표체재체pCI-pol Ⅰ중,획득H9N2금류감병독적8개기인절단적8질립계통.장구건호적8질립공전염293T세포후,수획상청접충계배,연후대계배뇨낭액진행감정;대증구적병독진행감정.결과 8질립계통전염293T세포후가이성공증구출H9N2금류감병독,혈응효개가체도29/50μl,생장특성여야생형병독유사.결론 성공건립료H9N2금류감병독반향유전계통.
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(HgN2)were amplified by RT-PCR and separately cloned into the transcription/expression vector, pCI-poll. The 8 plasmids DNA was co-transfected 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 HgN2 virus was successfully rescued by 8 plasmids co-transfection in 293T cells. The hemagglutinination titer of the rescued virus is up to 29/50 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 successfully.