The prokaryotic expression and purification of fusion protein 55kDa antigen from Pneumocystis carinii.

To generate 570bp gene fragment of Pneumocystis carinii 55kDa antigen,by constructing the prokaryotic expression vector pGEX-570,expression and purification of the fusion protein GST-p55/570.The gene fragment was amplified from P.carinii DNA by PCR and inserted into pGEM-T vector.The plasmids of pGEM-570 and pGEX-4T-1 were cutting by EcoR I and Xho I respectively,and then were retrieved to construct recombinant plasmid pGEX-570.The recombinant plasmids were identified by restriction endonucleases,PCR and the sequence analysis.Then it was transformed into E.coil BL21 for gene expression with IPTG induction.Through a great quantity induced expression,the recombinant proteins GST-p55/570 were purified by bag filter.Restriction endonucleases,PCR and the sequence analysis results indicated that the recombinant plasmid pGEX-570 had been constructed successfully.The recombinant plasmid was induced with IPTG to express a new fusion protein GST-p55/570 whose molecular weight was 47kDa.The proteins GST-p55/570 were purified by bag filter and identified by SDS-PAGE.In the present study,the recombinant plasmids pGEX-570 was successively constructed and expressed and the fusion protein GST-p55/570 was purified quite well.All the researches done in this paper provide foundation for further study on Pneumocystis carinii 55kDa antigen.