Chemically robust conductive p-type boron-doped diamond (BDD) films are an important electrode material and have been widely applied in electrochemistry. In this study, BDD films are taken as a two-dimensional (2D) electrode in a electrophoresis tank system instead of the conventional one-dimensional platinum wire electrode. The theoretical simulations by finite element numerical analysis reveal that the 2D BDD electrodes have relatively high intensity and uniformity of electric field in the tank. Experimentally, the 2D BDD electrodes with smaller size show excellent properties for the separation of DNA fragments. The advantages of the 2D BDD electrodes with chemical inertness, sustainability, high intensity and uniformity electronic field, as well as reduced small size of electrophoresis tank would open a possibility for realizing new generation, high-performance biological devices.