Position-configurable, reproducible, vertically aligned nanosheets assemblies (ANAs) arrays are fabricated by polymer-templated electrodeposition method at room temperature. Here, nanoimprint lithography is utilized to fabricate polymer template on the fluorine-doped tin oxide substrate for the purpose of evenly tuning the location of Ag nanostructures. Subsequently, vertically aligned ANAs can be achieved at the bottom of each hole via electrodeposition in a mixed aqueous solution of AgNO3 and citric acid. To obtain uniform ANAs array, we have systematically investigated the factors that influenced the electrodeposition. It was found that the formation of uniform ANAs arrays is strongly depended on the seeding layer, citric acid concentration, electrodeposition potential and time. The as-synthesized ANAs array exhibited a remarkable SERS activity and Raman signal reproducibility to rhodamine 6G, a concentration down to 10(-13) M can be identified. Our results revealed that the ANAs array is a highly desirable candidate as the reliable enhancer for high performance SERS analysis.