The distribution of metal nanoparticles on the surface of a surface enhancement Raman scattering (SERS)-active substrate plays a prominent part in not only the enhancement of Raman vibration signal, but also the spectrum uniformity. Here, a facile method to fabricate SERS substrates with excellent homogeneity and low cost was proposed, in which a lyotropic liquid crystal soft template was introduced for the coordinated growth of the silver nanoflowers in the process of electrochemistry deposition. Simulation was carried out to illustrate the dominated influence of the distance of electrodes on the deposited nanoparticle number. Two kinds of conductive materials, silver plate and indium tin oxide (ITO) glass, were chosen as the anode, while the cathode was fixed as ITO glass. The simulated conjecture on the effect of electrode flatness on the uniformity of deposited nanoparticles in silver is experimentally proved. More importantly, it was demonstrated that with a relatively smooth and flat ITO glass anode, a SERS substrate featuring higher spectrum uniformity could be achieved. This work is of great significance to the actual applications of the SERS substrate for quantitative detection with high sensitivity.