¡¾Abstract¡¿ AIM: To construct a new cell culture system for producing HBV in vitro.METHODS: Fullª²length HBV DNA was cloned into pcDNA3 vector (named pcDNA3ª²3HBV). HepG2 cells were transfected with pcDNA3ª²3HBV and screened with antibiotic G418. HBsAg and HBeAg were identified by ELISA and HBcAg was identified by immunocytochemical staining. S gene mRNA expression was tested by RTª²PCR and HBV DNA in the supernatant of transfected cells was tested by PCR. RESULTS: The plasmid pcDNA3ª²3HBV was constructed successfully. After stable transfection, HBsAg and HBeAg could be detected in the supernatant of transfected cells. HBcAg positive staining was located mainly in the cytoplasm. S gene mRNA expression was verified. S gene and preª²S gene could be detected in the supernatant by RTª²PCR. The copies of HBV genome can reach 1¡Á108 copies/L detected by realª²time quantitative PCR. CONCLUSION: Recombinant plasmid pcDNA3ª²3HBV could be expressed, transcribed and replicated in HepG2 cells. This transfectionª²based cell culture system could produce high copies of HBV genome.

    ¡¾Keywords¡¿ hepatitis B virus; genome, viral; cloning, moleculor; transfection; cell culture; gene expression ......