Prokaryotic expression of the PCV2 ORF4 gene and preparation of polyclonal antibodies against ORF4--《Acta Agriculturae Zhejiangensis》2014年01期

《Acta Agriculturae Zhejiangensis》 2014-01

Prokaryotic expression of the PCV2 ORF4 gene and preparation of polyclonal antibodies against ORF4

WANG Jing-xiu;GAO Zhang-zhao;JIANG Yong-hou;WU Hai-gang;RAO Pin-bin;QUAN Yan-ping;XU Hui;College of Life Sciences,Zhejiang Sci-Tech University;Centre of Animal Disease Control of Zhejiang Province;

Porcine circovirus 2( PCV2) is the primary causative factor of postweaning multisystemic wasting syndrome( PMWS) in swine. The first three open reading frames have been described while ORF4 is the fourth ORF in PCV2 genome and its function is still unknown. In this study,ORF4 gene was cloned into the vector of pGEX-4T-1 to construct recombinant plasmid pGEX-4T-ORF4. The recombinant plasmids confirmed by sequencing were transformed into BL21 competent strain and induced by IPTG. The purified GST-ORF4 protein was subcutaneously injected into the rabbits for preparation of the anti-GST-ORF4 antibody. Blood was collected four weeks later and the titer of polyclonal antibody was assessed by indirect enzyme-linked immune sorbent assay( ELISA). The results showed that the GST-ORF4 protein was successfully expressed by SDS-PAGE analysis and could be specifically detected with resulting polyclonal antibodies against GST-ORF4 protein by Western blot. The ELISA result indicated that the titer of the antibody was over 1∶ 12 800 and the antibody can be used for further functional study of ORF4 which may facilitate understanding the pathogenesis of PCV2 and interaction between PCV2 and host.

【Fund】：国家自然科学基金项目(31101831);; 浙江省自然科学基金(Y3090166)
【CateGory Index】： S852.651

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