猪圆环病毒Ⅱ型ORF1基因的克隆与原核表达  

Cloning and Prokaryotic Expression of ORF1 Gene of Porcine Circovirus Type 2

作　　者：曾朔[1] 吴星星[1] 冉多良[1] 陈丽娟[1] 胡雯婷[1] 

机构地区：[1]新疆农业大学动物医学学院,乌鲁木齐830052

出　　处：《新疆农业大学学报》2011年第4期307-310,共4页Journal of Xinjiang Agricultural University

基　　金：新疆维吾尔自治区重点产业紧缺人才专业大学生创新项目(jqrcp2201028)

摘　　要：根据GenBank中猪圆环病毒Ⅱ型(PCV2)ORF1基因序列,设计合成了一对引物,扩增出PCV2新疆株ORF1基因片段。将扩增出的ORF1基因(945 bp)插入到pMD18-T载体中,筛选获得重组质粒,命名为pMD18-ORF1。将ORFl酶切产物亚克隆到原核表达载体pET-32a中,获得重组质粒,命名为pET-32a-ORF1。用IPTG进行诱导表达,收集菌液进行SDS-PAGE检测,结果显示PCV2-ORF1在pET-32a中获得了高效融合表达,其表达蛋白分子量约为54 kDa,Western blotting分析结果表明,获得的融合蛋白可与PCV2阳性血清发生特异性反应,具有良好的免疫原性。This study was conducted to design,a pair of primer according to PCV2 ORF1 gene sequence in the Genbank.The ORF1 gene was amplified from suspected PMWS sample via polymerase chain reaction（PCR）,and the gene was cloned into pMD18-T vector.Then the ORF1 gene was subcloned into prokaryotic expressing vector pET-32a,the recombinant plasmid was named pET-32a-ORF1,and was induced by IPTG.The results of SDS-PAGE and Western-blotting indicated that the ORF1 gene was expressed in a high level,and the recombinant fusion protein was about 54 kDa.The recombinant fusion protein has peculiar reaction to PCV2 positive serum which has immunological activity.

关 键 词：PCV ORF1 克隆 原核表达 

分 类 号：S851.659.2[农业科学—预防兽医学]