乙型肝炎病毒e抗原真核表达载体的构建及在酵母中的表达

To investigate the biological functions of HBeAg protein, PCR was performed to amplify the gene of HBeAg from the plasmid pCP10 containing the whole fragment of ayw subtype of HBV.The PCR product was cloned into pGEM-T vector and was identified by DNA sequencing.Then, the gene of HBeAg was digested from pGEM-T vector and cloned into yeast expression plasmid pGBKT7. The reconstructed plasmid pGBKT7-HBeAg was transformed into yeast cell AH109 and screened on the synthetic dropout nutrient medium, (SD/-Trp) .The yeast protein was isolated and analyzed with sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and Western blotting.The results of SDS-PAGE and Western blotting assay showed the HBeAg was expressed in yeast cells and relative molecular weight of the expressed product was about 43000 Da.