Fig. 1. Expression of HBx and HBx–GFP as functionally active forms. (A) Expression pattern of GFP and the HBx–GFP fusion protein. HepG2 cells were transfected with pEGFP and pEG-HBx-GFP plasmid. At 24 h post-transfection, cells were photographed by fluorescence microscopy (magnification ×200). (B) Expression of HBx and HBx–GFP analyzed by western blotting. HepG2 cells were transfected with pEG-HBx or pEG-HBx-GFP, and at 10 h post-transfection cells were treated with TNF-α for 20 h. Equal amounts of total cell lysates were analyzed by 12% SDS–PAGE. (C and D) Transcriptional transactivity of HBx and HBx–GFP in the CMV promoter-driven CAT reporter assay in HepG2 cells. At 24 h after co-transfection (0.5 µg each), whole-cell lysates were prepared and normalized for CAT assay with the calorimetric method (C) and TLC method (D).