In the two decades since their initial discovery, DNA vaccines technologies have come a long way. Unfortunately, when applied to human subjects inadequate immunogenicity is still the biggest challenge for practical DNA vaccine use. Many different strategies have been tested in preclinical models to address this problem, including novel plasmid vectors and codon optimization to enhance antigen expression, new gene transfection systems or electroporation to increase delivery efficiency, protein or live virus vector boosting regimens to maximise immune stimulation, and formulation of DNA vaccines with traditional or molecular adjuvants. Better understanding of the mechanisms of action of DNA vaccines has also enabled better use of the intrinsic host response to DNA to improve vaccine immunogenicity. This review summarizes recent advances in DNA vaccine technologies and related intracellular events and how these might impact on future directions of DNA vaccine development.