Construction of an eukaryotic plasmid encoding HBsAg and screening of a cell line stably expressing clones

Objective To establish an eukaryotic vector encoding Hepatitis B Antigen(HBsAg),pcDNA3-HBsAg,and to screen HBsAg positive clones from mouse hepatic cancer cell line H22.Methods The HBsAg gene was amplified from PUC18-HBV by PCR.pCDNA3-HBsAg was prepared by inserting the fragment into the plasmid pcDNA3.Then the recombinant vector was identified by restriction enzyme and sequencing.Next pcDNA3-HBsAg was transfected into the H22 cell line and the positive clones(H22/HBsAg) were screened with G418.Finally,the expression of HBsAg in H22/HBsAg was detected by RT-PCR.Results The fragment of HBsAg was amplified and pcDNA3-HBsAg was prepared successfully.No errors were found both in the sequence and ORF of the acquired fragment.The expected fragment of HBsAg(678bp) was amplified from H22/HBsAg by RT-PCR.Conclusion:An eukaryotic plasmid encoding HBsAg(pcDNA3-HBsAg) has been constructed and HBsAg was transfected stably into H22 cell.