earticle

영어

White Spot Syndrome Virus (WSSV) occurs worldwide and causes up to 100% mortality within 7-10 days in commercial shrimp farm, resulting in large economic losses to the industry [1]. WSSV contains five major structural proteins. Two of them, VP19 and VP28 are located in the
envelope [2]. The aim of this study is cloning of fusion VP19 and VP28 gene and expression this fusion protein for development of vaccines against WSSV. The DNA fragments encoding the entire VP19 ORF and VP28 ORF were amplified from genomic WSSV DNA by PCR. To produce the fused gene, VP19 and VP28 genes were digested by BamHI enzyme and fused by T4 ligase enzyme. The fusion VP (19+28) gene was digested and inserted into EcoR I and Sal I sites of the expression vector pGEX to construct the recombinant plasmid. For expression, the recombinant plasmid pGEXVP(19+28) was transformed into E.coli (BL21) and cultured in LB medium
containing 100μg/ml ampicillin in a shaking incubator at 37°C until OD600=0.6. At this point, the expression of fusion protein was induced by adding IPTG at a final concentration of 0.2mM. At 2h post-induction, cells were harvested. The harvested cell was disrupted by sonication in 1X PBS buffer. The expression of recombinant protein VP (19+28) was further confirmed by running samples on SDS-PAGE and Western Blot, and bands corresponding to 73kDa were observed in the pGEX-VP(19+28), no protein was found at the same position in control with plasmid of alone vector pGEX. For further work, these recombinant proteins will be use for production vaccine against WSSV infection in shrimp.