[Construction and expression of eukaryotic expression plasmid pIRES-neo-HBAg carrying HBV fusion antigen gene].

OBJECTIVE

To construct a eukaryotic expression plasmid harboring HBV fusion antigen gene, and to express it in 293T cells.

METHODS

The HBV fusion gene fragment was amplified by PCR from the plasmid pVAX1-HBV containing HBV fusion gene. After purified, the product was cloned into pMD18-T vector. The recombinant plasmid was confirmed by endonuclease digestion and sequencing analysis, and then subcloned into eukaryotic expression vector pIRES-neo. Then the recombinant expression plasmid pIRES-neo-HBAg was transfered into 293T cells by Lipofectamine(TM); 2000. The expression of HBV fusion antigen was identified by Western blotting, flow cytometry and immunofluorescence cytochemistry.

RESULTS

The eukaryotic expression vector pIRES-neo-HBAg was constructed successfully. The expression of fusion antigen could be detected in the pIRES-neo-HBAg transfected 293T cells by Western blotting, immunofluorescence cytochemistry and flow cytometry.

CONCLUSION

The eukaryotic expression plasmid pIRES-neo-HBAg is successfully constructed and the fusion antigen is expressed in 293T cells.