Phosphorene, a monolayer of black phosphorus, has emerged as one of the most promising two-dimensional (2D) nanomaterials for various applications in the post-graphene-discovery period due to its highly anisotropic structure and novel properties. In order to apply phosphorene in biomedical fields, it is crucial to understand how it interacts with biomolecules. Herein, we use both molecular dynamics (MD) simulations and experimental techniques to investigate the interactions of phosphorene with a dsDNA segment. Our results reveal that dsDNA can form a stable binding on the phosphorene surface through the terminal base pairs and adopt an upright orientation regardless of its initial configurations. Moreover, the binding strength of dsDNA with phosphorene is found to be mild and does not cause significant distortion in the internal structure of dsDNA. This phenomenon is attributed to the weaker dispersion interaction between dsDNA and phosphorene. Further analysis of the free energy profile calculated by the umbrella sampling technique suggests that the puckered surface morphology significantly reduces the adsorption free energy of DNA bases to phosphorene. Compared to graphene, phosphorene is found to show a milder attraction to DNA, which is confirmed by our electrophoresis experiments. We believe that these findings provide valuable insight into the molecular interactions between phosphorene and dsDNA which may prompt further investigation of phosphorene for future biomedical applications.