||Establishment of an effective supplementary lighting procedure is necessary to increase the value of leaf lettuce grown using a hydroponic method involving a low production cost. In leaf lettuce extracts, quercetin, one of the flavonoids, was isolated and identified. It was investigated that quercetin has important functions that can be used as a dietary supplement. Flavonol synthase (FLS) is a key enzyme involved in quercetin biosynthesis, catalyzes the conversion of dihydroquercetin to quercetin. Therefore, we determined the sequence of the flavonol synthase gene (FLS) in red leaf lettuce. We harvested leaf lettuce grown using supplementary light sources, such as ultraviolet radiation B (UV-B), ultraviolet radiation A, blue, and red lamps during the night. It is noteworthy that FLS expression and the quercetin content were particularly increased to a greater extent in young leaves than in mature leaves when UV-B and blue light were used simultaneously at night. We suggest that UV-B with blue light is used simultaneously at night for producing leaf lettuce with high quercetin content.