The exploration and fabrication of nano-biointerfaces have fundamental significance and practical importance in many fields including chemistry, biology, and materials science. Recently, the integration of DNA with graphene has been substantially advanced. It is well known that single-stranded (ss) DNA can interact with graphene (or graphene oxide) via π–π stacking. However, for the case of DNA duplex/graphene, the studies are still not conclusive. Most work does not address the question of whether or how dsDNA is attracted to graphene oxide (GO). Here the interaction of DNA/GO is systematically investigated and its nanobiological effects, molecular recognition, and biosensing are explored. It is demonstrated that GO can adsorb DNA duplexes, which is possibly facilitated by partial deformation of the double helix on GO. Additionally dsDNA on GO shows specific effects on enzymatic degradation, which could be effectively cleaved by DNA enzyme I and restriction endonucleases as EcoR I, whereas it is highly resistant to degradation by Exo III. An improved understanding of the behavior of these GO/DNA entities will facilitate the development of applications in biomedicine, biosensing, and bionanotechnology.