Cloning of Chicken VEGF,Expression and Purification of Fusion Protein with GST

To obtain VEGF gene segments of chicken,express GST-cVEGF gene in E.coli and purify its fusin protein RT-PCR was used to amplify VEGF gene from chicken testis tissue.The PCR products were cloned into pMD 18-T vectors and sequenced.Then the gene fragments were subcloned into pGEX-4T-2 vectors,a prokaryotic expression vector with GST.Recombinant pGEX4T2-cVEGF was sequenced and transformed into E.coil BL21(DE3).The expression of GST fusion protein was induced by IPTG,and then purified by Ni-NTA Agarose.Finally,the purified proteins'molecular weight and specificity were detected by SDS-PAGE and western blot analysis.The result indicated that the recombinant vector pGEX4T2-cVEGF was constructed successfully.The GST-cVEGF fusion protein was expressed in E.coli BL21(DE3),and the molecular weight of the purified GST-cVEGF was identical to the expected values.The GST-cVEGF fusion protein purified was identified by Western blot.GST-cVEGF fusion protein may provide tools for further study of anti-tumor active immunity with this xenogeneic fusion protein vaccine in mouse tumor models.