The core gene promoter of duck hepatitis B virus (DHBV) has been localized and an enhancer element has been found in a region of the DHBV genome immediately upstream of the core promoter. This enhancer was able to activate expression from both the core promoter and the S promoter of DHBV, as well as from the heterologous thymidine kinase and simian virus 40 early promoters, but not from the DHBV PreS promoter, which was active both in the absence and in the presence of the enhancer. The activity of the enhancer showed a preference for cell cultures of hepatic origin, suggesting a possible tissue preferencein vivo. © 1991 Academic Press, Inc.