抗羊口疮病毒蛋白ORFV086多克隆抗体的制备及其应用

Abstract

Abstract: This study was aimed to clone and express ORFV086 protein of Orf virus (ORFV) in prokaryocytes and prepare polyclonal antibody anti-ORFV086 which was used to detect the expression of ORFV086 protein. The ORFV086 gene was amplified from genome DNA isolated from ORFV by PCR, then cloned into prokaryotic expression vector pET-33b (+) to construct a recombinant expression vector pET33b-086. The pET33b-086 was transformed into E.coli BL21 (DE3) pLys and induced by IPTG from which the fusion protein was identified by SDS-PAGE. The purified protein was injected into New Zealand rabbits and the polyclonal antibody was prepared, which was identified by the indirect ELISA and Western blotting methods. The results showed that the recombinant protein mainly existed in the inclusion body with the expected molecular weight of about 100 ku. After purified by cutting the gel slices, target protein was used as immunogen to prepare polyclonal antibody. The titer of the polyclonal antibody was 1:128000. Western blotting method revealed that the purified polyclonal antibody had a specific affinity for the reorganizational and natural ORFV086 protein. Thus, the rabbit anti-ORFV086 polyclonal antibody was successfully prepared, providing a tool for further investigation on the role of ORFV086 protein infection.

Key words: Orf virus; ORFV086 protein; prokaryotic expression; inclusion body; polyclonal antibody

CLC Number:

S852.65

WANG Xiao-ping, HAO Wen-bo, LUO Shu-hong, NING Zhang-yong. Preparation and Application of the Polyclonal Antibody against Orf Virus ORFV086 Protein[J]. , 2014, 41(11): 7-13.

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Preparation and Application of the Polyclonal Antibody against Orf Virus ORFV086 Protein

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