Circadian rhythms are observed in many physiological events in plant, and clock genes orchestrate the rhythms of expression of many genes. Precise environmental control of circadian rhythms provides a key technology for enhancing plant growth in artificial environments. In this study, we investigated the basic properties of a circadian rhythm for establishment of its control engineering in lettuce (Lactuca sativa L.). Bioluminescence of transgenic lettuce carrying an AtCCA1::LUC construct as a reporter of circadian gene expression was measured in young lettuce seedlings. We observed three basic properties; free-running circadian rhythms under constant conditions without day-night cycles, entrainment to red and blue light cycles with 12 h light-12 h dark period, and temperature compensation of a free-running period in three lettuce cultivars (Cisco, Cos, Greenwave). In addition, a light-quality dependence of the free-running period and sensitivity to blue-light cycles with small amplitude (20% of variance in average light intensity) and non-24 h period were also observed. The results in this study indicated that it is possible to control lettuce circadian rhythms by non-24 h period light cycles in LED illumination. It will play an important role in the research of control engineering for the circadian clock of lettuce in closed plant factories with artificial lighting.