To test the principle that genetically engineered epitopes in a plasmid DNA can efficiently induce specific immunity, a minigene cassette encoding cytotoxic T lymphocyte (CTL), helper T and B cell epitopes from herpes simplex virus (HSV) was constructed and placed in an expression vector named pcMini. Following immunizations with pcMini, mice developed epitope-specific CTLs comparable to the response induced by live HSV. Less effective but detectable antibody, lymphoproliferation, and T cell cytokine responses were also produced. In addition, pcMini-primed mice elicited a recall response upon restimulation with recombinant vaccinia virus expressing HSV antigen. The protection provided by minigene vaccination was significant, although not as efficient as live virus vaccine. The DNA minigene approach may prove useful to define and induce immune responses against minimal antigenic determinants.