Construction of eukaryotic expression vectors stably expressing pCDR1 Th epitope and CTLA4Ig fusion gene

Objective To construct an eukaryotic expression vector stably expressing pCDR1 Th epitope and CTLA4Ig fusion gene,and to identify its expression product.Methods Touchdown PCR was used to amplify CTLA4Ig cDNA and introduce pCDR1 Th epitope.The PCR product was ligated into the multiple clone site of eukaryotic expression vector pcDNA3.1(+)by gene recombination technique.After the attenuated Salmonella Typhimurium SL7207 containing the recombinant expression plasmids pcDNA3.1(+)-CTLA4-Ig-pCDR1 immunized BALB/c mice orally,spleens were removed to identify the recombinant proteins expression by immunohistochemistry assay.Results Restriction analysis and DNA sequence analysis showed that the pCDR1 Th epitope and CTLA4Ig cDNA had been successfully inserted into pcDNA3.1(+)eukaryotic expression vector.The fusion protein could be detected in the cytoplasm of spleen cells.Conclusion The eukaryotic expression vector stably expressing pCDR1 Th epitope and CTLA4Ig fusion gene is successfully constructed.