鸡IFN-γ基因在大肠杆菌中的表达及表达产物对鸡的免疫保护  

Expression of chicken IFN-γ gene in Escherichia coli and immunological protection of the expressed protein for chickens

作　　者：王立红 高春萍[2] 邢克智[3] 王红英[4] 郭永军[3] 

机构地区：[1]鼎正动物药业有限公司,天津300402 [2]清华大学深圳研究生院,广东深圳518055 [3]天津农学院养殖与生态重点实验室,天津300384 [4]天津市禽病诊断培训中心,天津300402

出　　处：《中国兽医科学》2008年第9期777-781,共5页Chinese Veterinary Science

基　　金：天津市科技攻关培育项目(05YFGPNC01900);天津市科技创新能力与环境建设平台项目(06TXTJJC14200)

摘　　要：将鸡干扰素-γ(ChIFN-γ)成熟肽基因编码区序列亚克隆到原核表达载体pQE30上,并转化大肠杆菌M15感受态细胞。经诱导后,以镍金属亲和层析法对表达蛋白进行纯化,采用SDS-PAGE与Western-blot对表达蛋白进行鉴定。结果显示,经诱导的转化子在体外获得了高效表达,表达蛋白约占菌体蛋白的23%。0.5ng/mL以上的重组ChIFN-γ(rChIFN-γ)能显著抑制鸡新城疫病毒对细胞的侵染与伤害;但是,细胞先感染病毒后,再以不同剂量rChIFN-γ作用于细胞,则不具有对细胞的保护作用,说明rChIFN-γ对细胞的保护是通过调节机体免疫能力、干扰病毒入侵实现的。1ng/mL的rChIFN-γ可显著提高新城疫疫苗的免疫效价,具有较强的免疫协同作用。表明,原核表达的rChIFN-γ具有显著的生物学活性。The mature peptide coding sequence of chicken IFN-γ（ChIFN-γ） gene was cloned into pQE30 vector and then transformed into Escherichia coli M15. The recombinant with ChIFN-γ gene was expressed with induction of IPTG and the expressed protein was identified by SDS PAGE and Western blotting using 6 × His-tag monoclonal antibody. Under denaturing conditions, the expressed rChlFN-γ was purified using the immobilized metal affinity chromatography technology with Ni NTA agarose,and refollittle protection for CEFs challenged with NDV,indicating that the main mechanism of rChIFN -γ against NDV was to regulate the immunological ability and to interfer with virus invasion. In addition, 1 ng/mL rChIFN-γ could improve significantly the antibody level of the NDV vaccine, and the SPF chickens were protected against NDV invasion. These results revealed that the expressed rChIFN-γ had significant biological activity.

关 键 词：鸡 干扰素-γ基因 体外表达 生物学活性 

分 类 号：Q511[生物学—生物化学] S852.4[农业科学—基础兽医学]