Expression of the Combined orf7 and orf5 Genes of Porcine Reproductive and Respiratory Syndrome Virus in E.coli.

According to the nucleotide sequence of Porcine reproductive and respiratory syndrome virus (PRRSV), two pairs of primers were designed and used to amplified the orf7 and orf5 genes by RT-PCR, respectively. By using the EcoRI、SpeI and Hind Ⅲ sites, both genes were cloned into pMD-18T vector in turn, then sequencing and comparing the homologies of the cloning genes. The combined orf7 and orf5 genes was subcloned into a prokaryotic expressing vector pGEX-KG, the recombinant plasmid named pGEX-KGNE was constructed and transformed into E.coli BL21.The results of SDS-PAGE and Western-blot indicated that the combined genes of orf7 and orf5 and Glutathion-S-transferase (GST) was expressed in E.coli BL21 as a fusion protein GST-NE about 64kDa in molecular weight, and this fusion protein showed reactive activity immunologically. This study lays a foundation for the development of the serodiagnostic methods and vaccines for PRRSV.