Over-expression in E.coli and purification of PTI-1 protein

AIM To construct human GST/PTI-1 gene recombinant plasmid, over-express GST/PTI-1 fused protein in E. Coli and purify it. METHODS Expressing vectorPTI-1/pGEX4T-1 was constructed by using plasmid pUC19/PTI-1, and then it was transformed into DH5α and identified by restriction analysis. The expressed product was analyzed by SDS-PAGE and purified by GST affinity chromatography. RESULTS Five positive clones of recombinant plasmids with pGEX4T-1 were screened and SDS-PAGE showed that a Mr 69 000 fused protein was over-expressed in E.coli, its purity being 43.2%. The purity of GST/PTI-1 fused protein obtained by GSH was 79.72%. CONCLUSION GST/PTI-1 fused gene recombinant plasmid was successfully constructed, over-expressed in E.coli and purified. The study makes it possible and easier to obtain GST/PTI-1 fused protein.