Objective To construct the recombinant modified vaccinia Ankara vaccine expressing HBsAg and investigate its antigenicity and immune effect by different strategies.Methods Target gene was cloned into plasmid pSC1l,the recombinant plasmid pSC11-S was transfected into BHK-21 cells that infected with MVA.Homologous recombination occurred between MVA and pSC1 1-S.The recombinant virus MVA-S was selected by several rounds of blue/clear plaques.Target gene was identified by PCR.The expression of target gene was analyzed by Western Blot.The BALB/c mice were immunized with single immunization and consecutive immunization strategy.The level of specific cellular response was measured by ELISpot assays.Results The recombinant virus expressing target gene was confirmed by PCR and Western Blot showed that MVA-S could elicit specific cellular response.There were no significant differences among two kinds of DNA vaccine and recombinant MVA-S.The prime/boost regimen could induce higher cellular response than single DNA vaccine immunization.High doses of MVA-S boost with the same DNA vaccine prime could induce highest response.There was no significant difference between different DNA vaccines (pVR-S,pcDNA-S) followed by same MVA-S boost.Conclusions MVA-S expressing target was constructed successfully,and it could induce specific cellular immune response which was higher with the strategy of DNA prime/MVA-S boost than that of single immunization.