禽多杀性巴氏杆菌脂蛋白E基因的原核表达及其对小鼠的免疫原性  

Expression of the Lipoprotein E Gene of Avian Pasteurella multocida and Its Immunogenicity in Mice

作　　者：崔卫涛[1] 胡思顺[1] 王延昭[1] 周晓芬[1] 李自力[1] 肖运才[1] 毕丁仁[1] 

机构地区：[1]华中农业大学动物医学院/农业微生物学国家重点实验室,武汉430070

出　　处：《湖北农业科学》2011年第4期854-857,共4页Hubei Agricultural Sciences

基　　金：湖北省科技攻关计划资助(2006AA202A05)

摘　　要：根据禽多杀性巴氏杆菌ATCC12 948株脂蛋白E基因(plpE)序列设计一对特异性引物,经PCR从禽多杀性巴氏杆菌C48-1株中扩增获得了全长plpE基因,大小约1kb。将plpE基因片段插入原核表达载体pET-28a中,构建了重组表达质粒pET-plpE。转化大肠杆菌BL21(DE3),在IPTG诱导下表达获得了重组蛋白plpE。SDS-PAGE结果显示,重组蛋白plpE约为37.8kD,与预期大小相符。Western blot检测结果表明,重组蛋白plpE能与C48-1菌体阳性血清发生特异性反应,用重组蛋白plpE免疫小鼠,二免后2周用10LD50 C48-1菌株攻毒即获得了100%的保护,表明该蛋白具有良好的免疫原性,为进一步的交叉保护性试验研究和禽霍乱亚单位疫苗研制奠定了基础。According to the plpE nucleotide sequence of avian Pasteurella multocida ATCC12948,a pair of primers was designed and amplified by PCR.And a lipoprotein E（plpE） gene was got from Pasteurella multocida C48-1 strain.It was cloned into the prokaryote expression vector pET-28a and used to recombine plasmid pET-plpE.Recombinant plpE could be expressed in E.coli BL21（DE3） via IPTG induction.The expressed product was identified by SDS-PAGE and Western-blotting.The results showed that fusion protein plpE was about 37.8kDa and it could react with antisera of C48-1.Subsequently,some six-week-old mice were immunized with 50μg of purified plpE.Two weeks after enhanced immunization,mice were challenged with injection of 10LD50 of strain C48-1.The survival rate was 100%.These results provided a good groundwork for further research on the cross-protective test and development of subunit vaccine against fowl cholera.

关 键 词：禽巴氏杆菌 原核表达 免疫原性 

分 类 号：S852.612[农业科学—基础兽医学]