DNA vaccines have failed to induce satisfactory immune responses in humans. Several mechanisms of double‐stranded DNA (dsDNA) sensing have been described, and modulate DNA vaccine immunogenicity at many levels. We hypothesized that the immunogenicity of DNA vaccines in humans is suppressed by APOBEC (apolipoprotein B (APOB) mRNA‐editing, catalytic polypeptide)‐mediated plasmid degradation. We showed that plasmid sensing via STING (stimulator of interferon (IFN) genes) and TBK‐1 (TANK‐binding kinase 1) leads to IFN‐β induction, which results in APOBEC3A mRNA upregulation through a mechanism involving protein kinase C signaling. We also showed that murine APOBEC2 expression in HEK293T cells led to a 10‐fold reduction in intracellular plasmid levels and plasmid‐encoded mRNA, and a 2.6‐fold reduction in GFP‐expressing cells. A bicistronic DNA vaccine expressing an immunogen and an APOBEC2‐specific shRNA efficiently silenced APOBEC2 both in vitro and in vivo, increasing the frequency of induced IFN‐γ‐secreting T cells. Our study brings new insights into the intracellular machinery involved in dsDNA sensing and how to modulate it to improve DNA vaccine immunogenicity in humans.