Despite the existence of various types of vaccines and the involvement of the world’s leading pharmaceutical companies, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) remains the most challenging health threat in this century. Along with the increased transmissibility, new strains continue to emerge leading to the need for more vaccines that would elicit protectiveness and safety against the new strains of the virus. Nucleic acid vaccines seem to be the most effective approach in case of a sudden outbreak of infection or the emergence of a new strain as it requires less time than any conventional vaccine development. Hence, in the current study, a DNA vaccine encoding the trimeric prefusion-stabilized ectodomain (S1+S2) of SARS-CoV-2 S-protein was designed by introducing six additional prolines mutation, termed HexaPro. The three-dose regimen of designed DNA vaccine immunization in mice demonstrated the generation of protective antibodies.