To evaluate the potential use of a 24-hour photoperiod for transplant production in a closed system, tomato (Lycopersicon esculentum Mill.) plug transplants were grown for 17 days either under a 24-hour photoperiod with a photosynthetic photon flux (PPF) of 200 μmol·m-2·s-1 or under a 16-hour photoperiod with a PPF of 300 μmol·m-2·s-1, resulting in the same daily integrated PPF (17.3 mol·m-2). Air temperatures were alternated between 28 °C during the first 16 hours and 16 °C for the subsequent 8 hours of each day. Fresh weight, dry weight and leaf area were 41%, 25%, and 64% greater, respectively, under the 24-hour photoperiod than under the 16-hour photoperiod. Physiological disorders (e.g., chlorosis and/or necrosis) were not observed under the 24-hour photoperiod, probably due to the alternating air temperature. Floral development of plants originating from both treatments did not differ significantly. Electric energy use efficiency of the closed system was 9% greater under the 24-hour photoperiod than under the 16-hour photoperiod. These results suggest that using a 24-hour photoperiod with relatively low PPF can reduce both initial and operational costs for transplant production in a closed system due to the reduction in the number of lamps.