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The morphology of lettuce roots in hydroponics is affected by the flow environment of nutrient solutions. Regulating the flow rate therefore ensures quality and yield; however, the mechanisms underlying this impact have not been fully elucidated, and breakthroughs in regulatory methods are thus lacking. Herein, lettuce was grown in a hydroponic solution at three different flow rates, the root morphology indicators (such as root length and surface area) were evaluated, the hormone levels of roots grown under different flow conditions were measured, and the correlations between plant hormones and root morphology were analyzed. Transcriptomic analysis was conducted on roots at varying flow rates to investigate the differential expression of genes involved in the synthesis and signal transduction pathways of auxin, abscisic acid (ABA), ethylene, and jasmonic acid. Results showed that these plant hormones were closely related to root morphology. Compared to a non-flow environment, a flow environment promoted the synthesis of plant hormones in the roots and enhanced the signal transduction of certain hormones, such as auxin. However, it had a negative impact on the signal transduction of certain hormones, specifically ethylene. The flow of the nutrients regulated root morphogenesis by influencing the synthesis and transduction of plant hormones, which affected overall plant growth. During the same growth period, lettuce grown under suitable flow conditions had a higher fresh weight than lettuce grown under no-flow conditions. Although such flow would increase the electricity cost of operating water pumps, lettuce could be harvested in advance at appropriate flow rates, thereby reducing the energy consumed in plant factories through lighting and temperature regulation, which are the largest contributors to energy consumption. This study examined the impact of the nutrient solution flow environment as a positive stimulus for plant growth, and it provides a reference for strategically regulating the flow rate in plant factories to improve yield and save energy. Future research should explore the effects of other cultivation and flow methods on the growth of different plants and apply these findings to practical production.