猪多杀性巴氏杆菌PlpP基因的原核表达及其免疫保护性的研究  

Prokaryotic expression of PlpP gene of Pasteurella multocida from swine and study on its immune protective efficacy in mice

作　　者：郝杰[1] 赵墩[1] 刘崇灵[1] 黄婕峰 余兴龙[1] 

机构地区：[1]湖南农业大学动物医学院,湖南长沙410128

出　　处：《中国兽医科学》2015年第6期584-588,共5页Chinese Veterinary Science

基　　金：长沙市科技成果产业化重点项目(K1308135-31)

摘　　要：为研究猪多杀性巴氏杆菌PlpP基因的免疫保护性,根据GenBank中登录的多杀性巴氏杆菌全基因组中的PlpP基因序列,对目的基因进行PCR扩增,并将PCR片段克隆至载体pET-28a(+)中,再将重组表达质粒pETPlpP转化至大肠杆菌BL21(DE3)中,用IPTG进行诱导。序列分析表明,得到了1条大小为945bp的基因片段,与预期大小相符。SDS-PAGE检测结果证实,该基因可以在BL21(DE3)中表达,表达产物为分子质量约45ku的融合蛋白。将纯化的重组蛋白pETPlpP制备成亚单位疫苗,并经皮下途径免疫小鼠,二免后第2周均以5LD50的A、B和D这3种荚膜抗原血清型的多杀性巴氏杆菌分别对小鼠进行攻毒,结果显示,A、B和D这3种血清型的保护率分别是16.67%、33.33%和66.67%。上述结果说明PlpP蛋白具有一定的免疫保护性。Based on the whole sequence of PlpP gene fromPasteurella multocidain GenBank,the PlpP gene was cloned and expressed in pET-28a(+)and its antigenicity was analyzed.Sequencing results showed that PlpP gene is 945 bp in length.SDS-PAGE confirmed that the recombinant protein is expressed in a form of inclusion bodies with about 34.7ku in molecular weight.In order to analyze the immune protective efficacy of PlpP gene,subunit vaccine was prepared with the purified recombinant protein pETPlpP and immunized subcutaneously to mice.On day 14post-second-immunization,the mice were challenged with 5LD50 of P.multocida CVCC444(Serotype A),CVCC434(Serotype B)and CVCC439(Serotype D),respectively.The results showed that the protection rates of PlpP on mice were 16.67%,33.33% and66.67% by P.multocidaserotypes A,B and D,respectively.In conclusion,the recombinant PlpP,to a certain extent,has the immune protective efficacy.

关 键 词：多杀性巴氏杆菌 PlpP基因 原核表达 抗原性 

分 类 号：S852.612[农业科学—基础兽医学]