Abstract: Objective Recombinant MVM VP2 protein was synthesized by baculovirus expression system and the immunogenicity of the recombinant VP2 protein was analyzed. Method The optimized MVM VP2 gene sequence was cloned into the eukaryotic expression vector pFastBac1, then the recombinant vector pFastBac1-MVM VP2 was transformed into DH10Bac susceptible E. coli to form recombinant plasmid. The recombinant plasmid were transfected into insect cells Sf9. Recombinant baculovirus rBacMVM VP2 was harvested after significant cellular lesions were observed. Sf9 cells were inoculated with recombinant virus rBac-MVM VP2 for 48 and 72 h. Immunogenicity of the recombinant protein was tested by IFA and Western blot. Result Recombinant baculovirus rBac-MVM VP2 could successfully express recombinant MVM VP2 protein after infecting Sf9 cells. Western blot and IFA analysis showed that the recombinant protein had good immunogenicity and high expression at 72 h of virus infection. After protein purification, a high purity recombinant protein VP2 was obtained. Conclusion In this study, MVM VP2 protein was successfully expressed by insect cell-baculovirus system and the recombinant VP2 protein has good immunogenicity, which lays a foundation for the study of VP2-related functions and the establishment of clinical diagnosis method.

Key words: minute virus of mice, VP2 protein, baculovirus expression

摘要： 目的 利用杆状病毒表达系统合成重组小鼠细小病毒(MVM) VP2 蛋白,并对其免疫原性分析。 方法 将优 化后的 MVM VP2 序列克隆至表达载体 pFastBac1,命名为 pFastBac1-MVM VP2,再将其转化至 DH10Bac 感受态大 肠杆菌中形成重组杆粒,提取重组杆粒经 PCR 鉴定正确后转染昆虫细胞 Sf9,镜下观察到细胞明显病变后收获重组 杆状病毒 rBac-MVM VP2。 Sf9 细胞接重组病毒 48 和 72 h 后,用间接免疫荧光 ( IFA) 和蛋白免疫印迹 ( Western blot)法验证重组蛋白的免疫原性。 结果 构建的 rBac-MVM VP2 重组杆状病毒感染 Sf9 细胞后,可成功表达 MVM VP2 重组蛋白,经 Western blot 和 IFA 检测分析,重组蛋白具有较好的免疫原性且在病毒感染 72 h 时表达量较高。 经蛋白纯化后可得到纯度较高的 VP2 重组蛋白。 结论 本研究利用昆虫细胞-杆状病毒系统成功表达 MVM VP2 蛋白并具有良好的免疫原性,为 VP2 相关功能的研究和临床诊断方法的建立奠定基础。 

关键词: 小鼠细小病毒, VP2 蛋白, 杆状病毒表达