猪传染性胸膜肺炎放线杆菌毒素apxⅡA基因的真核表达及其核酸疫苗  

Eukaryotic Expression and DNA Immunization of apxⅡA Gene of Porcine Actinobacillus pleuropneuromoniae

作　　者：贝为成[1] 严琳[1] 何启盖[1] 肖少波[1] 陈焕春[1] 

机构地区：[1]华中农业大学动物医学院动物病毒室,武汉430070

出　　处：《农业生物技术学报》2005年第5期616-619,共4页Journal of Agricultural Biotechnology

基　　金：国家自然科学基金(No.30200011);湖北省科技攻关项目(No.2001AA201B02)基金资助

摘　　要：利用PCR从自行分离鉴定的猪传染性胸膜肺炎放线杆菌(Actinobacillus pleuropneuromoniae,APP)血清7型菌株基因组DNA中扩增apxⅡA基因,构建了一个利用人类巨细胞病毒(Human cytomegalo virus HCMV)启动子表达apxⅡA基因的真核表达质粒pCDapxⅡA,体外转染PK-15细胞,处理后的样品进行SDS-PAGE和Western blot分析,证实了免疫原性蛋白apxⅡA在细胞中表达。用表达质粒pCDapxⅡA作为核酸疫苗免疫BALB/c小鼠酶联免疫吸附试验,ELISA检测小鼠血清中特异性抗猪胸膜肺炎放线杆菌的抗体,结果第2次免疫后其抗体滴度都在1:128以上。初步证实,用apxⅡA作为核酸疫苗免疫动物可以激活机体产生免疫应答反应。apxⅡA gene was amplified fi＇om genomic DNA and eukaryotic expression recombinant plasmid, pcDapxⅡA containing toxin apxⅡA gene of Actinobacillus pleuropneumoniae was constructed. The expression of foreign gene was promoted by immediate-early enhancer/promoter of Haman cytomegalo virus （HCMV）. The plasmid was transfected into PK-15 cell in vitro. Expressed protein was detected by SDS-PAGE and Western blotting assay. The recombinant plasmid was also used as DNA vaccine to inject BALB/c mice intramuscularly with 100 μg/mouse. The immunization was boosted at the 2^nd week after first injection. The antibody title were measured by indirect ELISA and higher than 1：128. Collecting these results showed that the apxⅡA gene as DNA vaccine to immunity animal could induce protective immunity to body.

关 键 词：猪传染性胸膜肺炎放线杆菌 apxⅡA基因 表达 核酸疫苗 免疫应答 

分 类 号：S858.286.3[农业科学—临床兽医学] Q781[农业科学—兽医学]