Construction of Plant Expression Plasmid of Chimera SBR-CT~(ΔA1)

Objective The purpose of this study is to construct plant expression plasmid containing the gene encoding chimera SBR-CT+{ΔA1}.Methods The target gene fragment P2, including the gene-encoded chimera SBR-CT+{ΔA1} (3 498～5 378 bp), was obtained by standard PCR amplification. The PCR products were ligated with pGEM-easy vector through TA clone to form plasmid pTSC. The plasmid pTSC and plasmid pPOKII were digested by restricted endonuclease BamHI and KpnI, and the digested products were extracted and purified for recombination. Then the purified P2 and plasmid pPOKII were recombined by T4 DNA ligase to form recombinant plasmid pROSC; inserting bar gene into the plasmid and form pROSB plasmid. The recombined plasmids were isolated and identified by restricted endonuclease cutting and Sanger dideoxy DNA sequencing.Results P2 gene was linked to pPOKII plasmid and formed recombinant plasmid pROSC. The DNA sequence and orientation were corrected. And bar gene was inserted into pPOSC and form recombinant plasmid pROSB.Conclusion Plant expression vector pROSC and pROSB containing the gene encoding chimera SBR-CT+{ΔA1}, which may provide useful experiment foundation for further study on edible vaccine against caries have been successfully constructed.