Human cytomegalovirus (HCMV) establishes severe disease in fetus, newborn and immunocompromised individuals. Polytope DNA vaccine strategy allows us to choose conserved and immunodominant epitopes from different antigens that can stimulate cellular and humoral immune responses simultaneously. In this study, a synthetic chimeric gene fragment was subcloned in to DNA vaccine vector pcDNA3.1+. The recombinant vector was transferred in to suitable eukaryotic cell line HEK 293T and the expression level of polytope construct from HEK 293T-infected cells was determined by western blot. These results show that there was no mutantion in target segment and recombinant vector showed significant levels of expression. Base on these results, using a proper procedure for design can cause expression and stability of polytope peptide.