The production performance of genetically improved farmed tilapia (GIFT, Oreochromis niloticus) and freshwater prawn (Macrobrachium rosenbergii) in periphyton‐based systems were studied in farmers' ponds at Mymensingh, Bangladesh. Fifteen ponds (200–300 m2 area and 1.0–1.5 m in depth) were used to compare five stocking ratios in triplicate: 100% GIFT, 75% GIFT plus 25% prawn, 50% GIFT plus 50% prawn, 25% GIFT plus 75% prawn and 100% prawn. Ponds were stocked at a total density of 20 000 GIFT and/or prawn ha−1. Bamboo poles (mean diameter 6.2 cm and 5.5 pole m−2) were posted in pond bottoms vertically as periphyton substrate. Periphyton biomass in terms of dry matter (DM), ash‐free DM and chlorophyll a were significantly higher in ponds stocked with prawn alone than in ponds with different combinations of GIFT and prawn. Survival of GIFT was significantly lower in ponds stocked with 100% GIFT (monoculture) whereas, that of prawn was significantly higher in its monoculture ponds indicating detrimental effects of GIFT on prawn's survival. Individual weight gains for both species were significantly higher in polyculture than in monoculture. The highest total fish and prawn yield (1623 kg GIFT and 30 kg prawn ha−1) over 125–140 days culture period was recorded in ponds with 75% GIFT and 25% prawn followed by 100% GIFT alone (1549 kg ha−1), 50% GIFT plus 50% prawn (1114 kg GIFT and 68 kg prawn ha−1), 25% GIFT plus 75% prawn (574 kg GIFT and 129 kg prawn ha−1) and 100% prawn alone (157 kg ha−1). This combination also gave the highest economic return. Therefore, a stocking ratio of 75% GIFT plus 25% prawn at a total density of 20 000 ha−1 appeared to be the best stocking ratio in terms of fish production as well as economics for a periphyton‐based polyculture system.