We developed a facile approach to synthesize thin-film nano-templated composite (TFNt) nanofiltration membrane with high water permeability, high NaCl/MgSO4 selectivity and strong antimicrobial properties. A polydopamine (PDA) coating on a polysulfone support was used as a nano-template to generate silver nanoparticles (AgNPs) in situ with high loading and high uniformity. A subsequent interfacial polymerization reaction of piperazine and trimesoyl chloride was performed on this nano-template substrate to form the TFNt membrane. The TFNt membrane had significantly increased both the water permeability and salt rejection compared to the control thin-film composite (TFC) membrane as well as a thin-film nanocomposite (TFN) membrane prepared by the conventional way of loading AgNPs directly during the interfacial polymerization process. Furthermore, the TFNt membrane showed better antimicrobial properties than both the TFC and the conventional TFN membranes. The current work presents an exciting approach to fabricate novel nanofiltration membranes using nano-templates, which provides important insights for high performance NF membrane synthesis.