鸡源性HMGB1蛋白的原核表达及多克隆抗体制备  

PROKARYOTIC EXPRESSION AND ANTIBODY PREPARATION OF CHICKEN HMGB1 PROTEIN

作　　者：曲昱蓉 詹媛[1] 仇旭升[1] 丁铲[1] QU Yu-rong;ZHAN Yuan;QIU Xu-sheng;DING Chan(Shanghai Verinary Research Institute,CAAS,Shanghai 200241,China)

机构地区：[1]中国农业科学院上海兽医研究所,上海200241

出　　处：《中国动物传染病学报》2018年第2期68-73,共6页Chinese Journal of Animal Infectious Diseases

基　　金：国家自然科学基金(31372421和31530074);公益性行业(农业)科研专项(201303033)

摘　　要：从鸡胚原代细胞CEF中扩增出鸡HMGB1基因,构建重组原核表达质粒p ET-28a-ch HMGB1,并将p ET-28a-ch HMGB1转入大肠杆菌BL21(DE3)中,于37℃进行诱导表达,经SDS-PAGE分析表明该重组蛋白可以在大肠杆菌中高效表达且以可溶性蛋白的形式存在。蛋白通过Ni-NAT纯化树脂亲和纯化,并作为免疫原制备鼠抗ch HMGB1多克隆抗体血清。该多克隆抗体同时具有ELISA、Western blot和IFA效价,且特异性识别禽源细胞内HMGB1蛋白。The chHMGB1 gene was amplifi ed from CEF cells and recombinant prokaryotic expression vector pET-28a-chHMGB1 and eukaryotic expression vectors FLAG-chHMGB1 were constructed accordingly.The recombinant protein chHMGB1 was expressed at 37℃in E.coli BL21(DE3).The SDS-PAGE analysis showed that the recombinant chHMGB1 was effi ciently expressed in E coli and existed in the form of soluble protein.The recombinant chHMGB1 was then purified by affinity chromatography on Ni-NAT for preparation of polyclonal antibodies.The resulting polyclonal antibodies showed specifi c reaction with chHMGB1 in ELISA,Western blot and IFA.

关 键 词：鸡源性HMGB1蛋白 原核表达 多克隆抗体 

分 类 号：S852.42[农业科学—基础兽医学]