Cloning and Prokaryotic Expression of ORFV 115 Gene

    Objective   ORFV 115 was cloned for a study on the gene.

    Method    Oligo 7 software was used to design and select primers for specific amplification of the gene. Using the genome of ORFV FJ-ND as template, the sequence was obtained by PCR. The gene was then cloned into pGEX-6p-1 to obtain the recombinant plasmid pGEX-6P-115 for sequencing and identification prior to transformation to RosettaganmiB (DE3) receptor cells. Expression conditions of the recombinant fusion protein were optimized with respect to IPTG concentration and induction time. SDS-PAGE was used to analyze the expressed target protein.

    Result   ORFV 115 was 450 bp encoded 149 amino acids and could be expressed in RosettaganmiB. It was approximately 42 kDa in molecular weight and mainly expressed as inclusion body.

    Conclusion   ORFV 115 was successfully cloned, the pGEX-6P-115 prokaryotic expression plasmid constructed, the recombinant protein purified, and the expression conditions optimized to facilitate further studies on the mechanism and role it plays in the infection on host animals.