Porcine reproductive and respiratory syndrome virus(PRRSV)has brought huge economic losses to the pig industry.Therefore,it is particularly important to detective a rapid diagnosis method for the virus as soon as possible.The structural protein GP4 encoded by the ORF4 gene of PRRSV is necessary for virus infection.In order to prepare GP4 monoclonal antibodies(MAbs),the ORF4 superior sequence was selected to optimize synthesis and then connected to pET-32a prokaryotic expression vector.The constructed recombinant plasmid pET-32a-ORF4 was verified through double enzyme digestion and transformed into BL21(DE3)competent cells for protein expression with IPTG induction.The expressed protein was purified and confirmed in SDS-PAGE and Western blotting.The GP4 protein was expressed in supernatant form and the size was about 30 kDa.The purified protein was used to immunize mice for preparation of MAbs.As result,a total of 4 IgG subtype-positive hybridomas were obtained and subcloned for production of specific MAbs.The availability of MAbs to GP4 protein laid the foundation for future research and development of PRRSV diagnostic methods.
关键词
猪繁殖与呼吸综合征病毒/GP4蛋白/原核表达/单克隆抗体
Key words
Porcine reproductive and respiratory syndrome virus/GP4 protein/prokaryotic expression/monoclonal antibody
维普
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