Construction of eukaryotic expression vectors of HBV genes and transfection

Objective To construct eukaryotic expression vectors of human HBV genes and transfect theminto HepG2 cells for establishing stable HepG2 cell lines.Methods The HBV gene fragments were amplified from the HBV genome plasmid by PCR and were inserted into eukaryotic expression vector pcDNA3.1(+),respectively.After the identification by digestion and sequencing,the recombinant eukaryotic expression vectors were transfected into HepG2 cell respectively by lipofectamineTM 2000.After screening culture by G418,the stable transfected HepG2 cell lines were established,and the expression of HBV gene products were identified by flow cytometry and immunocytochemical staining.Results The eukaryotic expression vectors pcDNA3.1(+)/HBs,pcDNA3.1(+)/HBc,pcDNA3.1(+)/HBe,pcDNA3.1(+)/HBp,pcDNA3.1(+)/HB-preS1,pcDNA3.1(+)/HB-preS2,and pcDNA3.1(+)/HBx were constructed successfully.The stable transfected HepG2 cell lines were established and the HBV gene products were expressed successfully.Conclusion The construction of the eukaryotic expression vectors and the establishment of stable transfected HepG2 cell lines provide a solid foundation for further experimental studies on the function of HBV genes.