人肠道病毒71型高致病性毒株及其体外适应株cDNA感染性克隆的构建  

Construction of cDNA Infectious Clones of EV71 Highly-Pathogenic and Cell-Culture-Adapted Strains

作　　者：张永欣[1] 李晓宇[1] 黄宇明[2] 周永东[3] 毕胜利[3] 岑山[1] 

机构地区：[1]中国医学科学院医药生物技术研究所,北京100050 [2]首都医科大学附属北京地坛医院,北京100015 [3]中国疾病预防控制中心病毒病预防控制所,北京100052

出　　处：《病毒学报》2014年第6期605-613,共9页Chinese Journal of Virology

基　　金：十二五"新药创制"科技重大专项基金(2012ZX09301-002-004);十二五"艾滋病和病毒性肝炎等重大传染病防治"科技重大专项基金(2013ZX10004601-002);国家自然科学基金(81401675)

摘　　要：高致病性EV71毒株的感染易伴发神经系统症状,是导致手足口病死亡病例的主要原因,但目前临床上仍缺乏防治EV71感染的特效药物及安全有效的疫苗。为进一步明确EV71的致病机制,本项工作以高致病性的EV71-HP临床分离株及其经体外细胞培养适应的EV71-CCA毒株的基因组RNA为模板,应用反向遗传学方法构建了二者的感染性克隆。研究表明HP株在细胞中的增殖速度明显快于CCA株。两株基于感染性克隆的拯救病毒分别保持了与各自母本株基因组序列的一致性,同时也保留了母本株增殖表型的差异,包括病毒的增殖速度以及病毒的温度敏感性。本工作将促进对EV71的致病机制以及对EV71疫苗的研究。The highly-pathogenic EV71 strain is the primary cause of mortality in hand-foot-and-mouth disease（HFMD）associated with neurological symptoms,for which no clinically effective drugs or vaccines exist.This study aimed to construct infectious cDNA clones of the EV71highly-pathogenic strain and the cell-culture adapted strain using a reverse genetics approach.The genomic RNAs of EV71 parent strains were used as the templates for RT-PCR amplification,and then the PCR products that overlapped the fulllength genome were connected into the pBR322 vector to produce infectious clones of pEV71（HP）and pEV71（CCA）,respectively.The results showed that the HP strain propagated much more quickly than the CCA strain.The rescued viruses derived from the infectious clones not only maintained their consistency with their parent strains in terms of genomic sequences,but also retained their respective biological phenotypes.This research will contribute to our understanding of EV71 pathogenesis and the development of novel vaccines against HFMD.

关 键 词：肠道病毒71型 感染性克隆 拯救病毒 增殖表型 温度敏感性 

分 类 号：R373.2[医药卫生—病原生物学]