猪繁殖与呼吸综合征病毒ORF5/6/7基因真核载体的构建  

Construction of Eukaryotic Expressing Vector from ORF5/6/7 of Porcine Reproductive and Respiratory Syndrome Virus

作　　者：康艳梅[1] 赵明秋[1] 沈海燕[1] 琚春梅[1] 陈金顶[1] 

机构地区：[1]华南农业大学兽医学院,广东广州510642

出　　处：《华南农业大学学报》2012年第2期256-258,263,共4页Journal of South China Agricultural University

基　　金：广东省科技计划项目(2007A020300064);广东省自然科学基金创新团队项目(5200638);教育部"长江学者和创新团队发展计划"创新团队项目(TRT0723)

摘　　要：针对PRRSV ORF5～ORF7设计3对特异性引物,以PRRSV GD-XH株和GD08-1株为模板进行RT-PCR扩增,并将扩增产物克隆到真核表达载体pEGFP-N1中,获得重组质粒pEGFP-N1-(ORF5～ORF7),然后将重组表达质粒转染Marc-145细胞.通过荧光显微镜观察显示,重组质粒pEGFP-N1-(ORF5～ORF7)和载体pEGFP-N1均有绿色荧光出现.结果表明:成功构建了表达PRRSV GD-XH株和PRRSV GD08-1株的GP5、M、N蛋白的真核表达质粒pEGFP-N1-GD-XH-GP、pEGFP-N1-GD-XH-M、pEGFP-N1-GD-XH-N、pEGFP-N1-GD08-1-GP5、pEGFP-N1-GD08-1-M、pEGFP-N1-GD08-1-N.Specific primers aiming to ORF5-ORF7 of porcine reproductive and respiratory syndrome（PRRSV） were designed.The target segments with restricted enzyme digestive sites were obtained by RT-PCR with PRRSV GD-XH and GD08-1 as template.All the segments were inserted into vector pEGFP-N1 to obtain six recombinant plasmids：pEGFP-N1-GD-XH-GP5,pEGFP-N1-GD-XH-M,pEGFP-N1-GD-XH-N,pEGFP-N1-GD08-1-GP5,pEGFP-N1-GD08-1-M,pEGFP-N1-GD08-1-N.The recombinant plasmid was transfected into Marc-145 cell.Fluorescence microscopy showed that EGFP were expressed in all transfected cells.

关 键 词：猪繁殖与呼吸综合征病毒 ORF5/6/7 PEGFP-N1 

分 类 号：S852.65[农业科学—基础兽医学]