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目的提高表达狂犬病毒糖蛋白（ＲＧ）的重组痘苗病毒的安全性。方法将编码中国狂犬病毒５ａＧ株糖蛋白的基因，插入痘苗病毒天坛株的ＴＫ区，获得重组病毒ＶＴＫＲＧ，通过两步同源重组，删除ＣＫ片段间与痘苗病毒毒力及宿主范围相关的基因，得到非复制型重组痘苗病毒ＶＴＫＲＧ△ＣＫ。结果经ＰＣＲ鉴定，ＣＫ间的核酸片段被成功删除，其缺失性状能稳定地遗传。ＷｅｓｔｅｒｎＢｌｏｔ及间接免疫荧光结果表明，ＶＴＫＲＧ△ＣＫ能有效地表达ＲＧ。该病毒在鸡胚成纤维细胞中繁殖，在人源细胞如ＴＫ１４３中几乎不增殖。动物实验证实，ＶＴＫＲＧ△ＣＫ免疫小鼠后能诱生中和抗体并能保护小鼠抵抗致死剂量狂犬病毒（ＲＶ）的攻击。结论ＶＴＫＲＧ△ＣＫ具有良好的免疫原性和更高的安全性。
Objective To improve the safety of recombinant vaccinia virus expressing rabies virus glycoprotein (RG). Methods The gene encoding Chinese rabies virus 5aG glycoprotein was inserted into the TK region of vaccinia virus Tiantan strain and the recombinant virus VTKRG was obtained. After two-step homologous recombination, the genes related to virulence and host range of the vaccinia virus were deleted, Get non-replicating recombinant vaccinia virus VTKRG △ CK. Results The results of PCR showed that the nucleotide sequence of CK was deleted successfully, and the deletion trait was stable. Western Blot and indirect immunofluorescence results showed that VTKRG △ CK can effectively express RG. The virus multiplies in chicken embryo fibroblasts and hardly proliferates in human cells such as TK143. Animal experiments confirmed that VTKRG △ CK immunized mice induced neutralizing antibodies and protect mice against lethal dose of rabies virus (RV) attack. Conclusion VTKRG △ CK has good immunogenicity and higher safety.