Construction of HBV infectious replicon and its significance

Objective To construct expression vector of HBV infectious replicon and establish hepatitis B virus(HBV) replication model in vitro,and observe the dynamic changes of viral replication and its sensitivity to antiviral durg.Methods 1.3 fold overlength genome of HBV,from pHBV-dimer carring two head to tail copies of HBV genome,was cloned into pCR 2.1 by combined PCR and restriction enzyme digestion.The recombinant plasmids containing 1.3 copies of HBV genome were transfected into Huh7 cell line.The levels of HBsAg and HBeAg in supernatant of Huh7 cells were measured by ELISA,intracellular HBV replicative intermediates and intracellular HBV transcripts were analyzed by Southern Blot and Nor thern Blot respectively.Antiviral effect of adefovir was evaluated in vitro.Results The recombinant expression plasmid of HBV infectious replicon was constructed successfully.HBV gene carried in pHBV 1.3 could efficiently replicate and express in Huh 7 cells.A defovir inhibited HBV replication in this cell model of HBV.Further more,the inhibition effect depended on the concentration of durg.Conclusion This recombinant HBV replicon,with initiates viral replication efficiently in transfected hepatoma cells,is expected as a novel tool in the field of HBV replication and antiviral research.