目的 构建表达狂犬病毒糖蛋白基因(GP)的重组人腺病毒。方法 克隆狂犬病毒疫苗株GP基因并测序，将其插入E3区缺失的Ad5载体，置于E3区早期启动子的控制下，通过同源重组，蚀斑纯化，筛选表达GP的重组人腺病毒。用ELISA法检测表达的糖蛋白，快速荧光灶抑制试验(RFFTT)法测定此重组病毒免疫小鼠后产生的中和抗体。结果 获得的3aG株基因的开放读码框为1575个核苷酸，编码524个氨基酸；经同源重组和蚀斑纯化获得了E3区表达狂犬病毒GP基因的重组人腺病毒；其表达产物能特异性地被抗狂犬病毒人免疫血清所识别；免疫小鼠后能产生一定水平的中和抗体。结论 成功构建了E3区表达狂犬病毒GP基因的重组人腺病毒，此重组腺病毒能有效诱导产生特异性中和抗体。%Objective To analysis Chinese rabies vibies virus vaccine strain 3aG glycoprotein(GP) gene and furtherproduce GP by E3-deleted human adenovirus recombinant.Methods Chinese rabies virus vaccine strain 3aG glyco-protein gene was cloned by RT-PCR and its sequence was determined by DNA sequencing.Cotransfection was per-formed to obtain adenovirus recombinant.The expressed glycoprotein was examined by ELISA and its immunogenicitywas evaluated by testing neutralizing antibody level of mice inoculated with the recombinant virus.Results DNA se-quencing showed that the open reading frame of GP gene contains 1 575 nucleotides and five of the deduced amino acidsare different from the previous report.The recombinant adenovirus containing GP gene in E3 region was obtained by co-transfecting 293 cells and rounds of plaque purification. ELISA assay demonstrated that the GP gene can be efficientlyexpressed and rapid fluorescent focus inhibition test (RFFTT) showed it can also elicit GP specific neuralizing antibody.Conclusion Ciinese rabies vurus vaccine strain 3aG glycoproteein was successfully expresssed by E3 - deleted humanadenovirus recombinant snd sprvigic nrutralizing snyinody can be elicited in mice sfter immunixsyion by the recombinant.
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