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PRRSVGD08-1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 Peg-FP-N1 to obtain six recombinant plasmids:Pegfp-Nl-GD-XH-GP5, Pegfp-Nl-GD-XH-M, Pegfp-Nl-GD-XH-N, PEGFP-N1-GD08-1-GP5, PEGFP-N1-GD08-1-M, Pegfp-Nl-GD08-l-N. The recombinant plasmid was transfected into Marc-145 cell. Fluorescence microscopy showed that EGFP were expressed in all transfected cells.
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