中国草食动物科学2025，Vol.45Issue(3)：39-45,7.DOI:10.3969/j.issn.2095-3887.2025.00.003
施马伦贝格病毒N蛋白的原核表达及其单克隆抗体制备
Prokaryotic Expression of Schmallenberg Virus N Protein and Preparation of Its Monoclonal Antibody
摘要
Abstract
Schmallenberg virus(SBV)is a new orthobunyavirus that primarily infects ruminants such as cattle,sheep and goats.The nucleocapsid protein(N)of SBV has been shown to be an ideal target antigen for serological testing.In this study,the full-length coding sequence of SBV N gene was cloned into pET-28a(+)vector,and the recombinant plasmid pET-28a-SBV-N was constructed,which was transformed into Roseseta competent cells,and IPTG induced the expression of N protein.His-SBV-N protein was purified by Ni-NTA agarose affinity chromatography to obtain N protein to immunize BALB/c mice,and after four immunizations,the splenocytes of mice with high titer were fused with SP2/0 cells to prepare monoclonal antibodies against SBV N protein(McAbs),and the hybridoma cells secreting monoclonal antibody were screened by indirect ELISA.Finally,three hybridoma cell lines that stably secreted anti-N monoclonal antibody were obtained,which were named 2B1,2B2 and 3F2,respectively.The antibody isotype was identified as IgG2a.The results of Western blot and ELISA showed that the three monoclonal antibodies had good reac-tivity with recombinant SBV N protein in enzyme-linked immunosorbent assay,and the expression of recombinant N protein and the successful preparation of monoclonal antibodies provided important materials for the serological diag-nosis of SBV.关键词
施马伦贝格病毒/核衣壳蛋白/原核表达/细胞融合/单克隆抗体Key words
Schmallenberg virus/N protein/prokaryotic expression/cell fusion/monoclonal antibodies分类
畜牧业引用本文复制引用
仇槿昕,侯辉,孙法壮,冯耀宇,李维克..施马伦贝格病毒N蛋白的原核表达及其单克隆抗体制备[J].中国草食动物科学,2025,45(3):39-45,7.基金项目
甘肃省科技重大专项(22ZD6NA001) （22ZD6NA001）
