牛坏死杆菌43K OMP多克隆抗体的制备

Polyclonal Antibody Preparation of 43K OMP from Fusobacterium Necrophorum

为了评价牛坏死杆菌外膜蛋白43K OMP 的免疫原性,试验利用大肠杆菌原核表达系统对牛坏死杆菌43K OMP基因进行表达,纯化目的蛋白免疫兔,制备牛坏死杆菌43K OMP多克隆抗体,并对其进行鉴定。对基因序列进行预测分析后,针对大肠杆菌稀有密码子优化合成该基因序列,克隆至pET-32a 原核表达载体上,经异丙基硫代-β-D-半乳糖苷(IPTG)诱导表达及镍柱进行纯化,获得重组蛋白大小约为59 ku。重组蛋白经Western blotting 鉴定后免疫兔,制备多克隆抗体。Western blot 检测牛坏死杆菌43K OMP多克隆抗体与43K OMP重组蛋白具有很好的免疫原性。研究成功制备了牛坏死杆菌43K OMP 多克隆抗体,为以43K OMP蛋白为基础的疫苗研究奠定基础。

In order to analyze the immunogenicity of bovine necrosis bacillus outer membrane protein(43 K OMP),the 43 K OMP gene of Fusobacterium necrophorum was expressed in E.coli prokaryotic expression system,and the purified protein was used to immunize the rabbits,polyclonal antibodies against 43 K OMP was prepared and identified.The recombinant protein was synthesized and cloned into the prokaryotic expression vector of p ET-32 a.The recombinant protein was induced by isopropylthioside β-D galactoside(IPTG)and purified by nickel column.The length of the recombinant protein was about 59 ku. Polyclonal antibody was prepared by immunization of recombinant protein with western blot.Western blot detection results showed that the 43 K OMP polyclonal antibody and 43 K OMP recombinant protein had better immunogenicity.The polyclonal antibody against bovine 43 K OMP laid a foundation for the further research of 43 K OMP protein vaccine.