Construction of HBV whole-genome 1. 3 ploid HepG2 cell model and expression of HBV biomarkers

Objective To construct an HepG2 cell model containing HBV whole-genome 1. 3 ploid ( HBV 1. 3P) , and to investigate the expression of HBV biomarkers. Methods HepG2 cells were transfected with the 1. 3-ploid whole-genome HBV DNA sequence using the adenovirus vector to construct an HBV 1. 3P-HepG2 cell model. Agarose gel electrophoresis was used to identify HBV 1. 3P recombinant adenovirus plasmids, and the ABI 3730 sequencer was used to confirm whether the recombinant adenovirus plasmids had correct HBV 1. 3P sequence. The optimal multiplicity of infection ( MOI) of HepG2 cells infected by the HBV 1. 3P adenovirus was determined under a fluorescence microscope. Quantitative real-time PCR was used to measure the expression of HBV DNA and ccc DNA in the HBV 1. 3P-HepG2 cell model at 0-9 days, and chemiluminescence and immunofluorescence assay were used to measure the expression of HBs Ag and HBe Ag, respectively, in supernatant and cells. An analysis of variance was used for comparison of continuous data between multiple groups, and the SNK-q test was used for comparison between any two groups. Results When MOI was 40, the efficiency of HepG2 cells being infected by HBV 1. 3P recombinant adenovirus reached above 90%. On the second day of infection, the expression of the biomarkers HBV DNA, ccc DNA, HBs Ag, and HBe Ag was detected in supernatant and cells, and the levels of these biomarkers reached peak values at 4-6 days and gradually decreased after 7 days. Conclusion This HBV 1. 3P-HepG2 cell model can express HBV biomarkers stably, which lays a foundation for future research on HBV.