狂犬病病毒HEP-Flury-mEG株的拯救与鉴定  

Rescue and characterization of recombinant rabies virus HEP-Flury-mEG

作　　者：郑学星[1,2] 杨松涛[1,2] 王化磊[1,2] 薛向红[1,2] 盖微微[1,2] 赵永坤[1,2] 郭霄峰[3] 夏咸柱[1,2] 

机构地区：[1]军事医学科学院军事兽医研究所,吉林长春130062 [2]吉林省人兽共患病预防与控制重点实验室,吉林长春130062 [3]华南农业大学兽医学院,广东广州510642

出　　处：《中国病原生物学杂志》2012年第12期881-884,共4页Journal of Pathogen Biology

基　　金：公益性行业(农业)科研专项(No.201103032);"十一五"国家科技支撑计划重点项目(No.2010BAD04B03);人兽共患病研究教育部重点实验室开放基金

摘　　要：目的拯救狂犬病毒HEP-Flury-mEG株,为疫苗制备奠定基础。方法将ERA株G蛋白333位毒力位点修饰为谷氨酸后替换至HEP-Flury株基因组。重组感染性克隆质粒和4个辅助质粒,共转染BHK-21细胞,拯救重组病毒。结果 IFA鉴定成功拯救出了HEP-Flury-mEG株狂犬病病毒。重组病毒G基因经XholⅠ酶切,片段大小为1 071bp和520bp,与预期结果相符。重组病毒在BHK-21细胞中传代4次,滴度维持在1×107.5 FFU/ml。重组病毒经常规负染后在电镜下为弹状粒子,长度和直径与亲本株一致。结论获得了重组狂犬病毒HEP-Flury-mEG株。该病毒滴度高,形态完整,传代稳定,为进一步研究狂犬病毒病毒的生物学特性和新型基因工程疫苗奠定了基础。Objectives To rescue a strain of rabies virus,named HEP-Flury-mEG,,and provide a basis for vaccine development.Methods G of the HEP-Flury strain was replaced with the mutant G gene of the ERA strain in which Arg333 was substituted with a Glu residue.BHK-21 cells were cotransfected with the full-length infectious clone and four other helper plasmids,and then the recombinant virus was rescued.Results IFA results showed that the HEP-Flury-mEG strain was successfully rescued.The recombinant G gene resulted in two fragments after XhoI digestion with a respective size of 1 071 bp and 520 bp,just as expected.The titer of HEP-Flury-mEG remained at 1×107.5 FFU/ml after four passages.HEP-Flury-mEG was observed with electron microscopy after negative staining.Cells were bullet-shaped and both their length and diameter were the same as those of the parental strain.Conclusion Recombinant HEP-Flury-mEG was obtained in a high titer with an intact morphology and genetic stability,providing important insight into the biological characteristics of rabies virus and development of novel genetic vaccines.

关 键 词：狂犬病病毒 HEP-Flury株 ERA株 反向遗传 糖蛋白 

分 类 号：R373.9[医药卫生—病原生物学]