Background and Aims Soil waterlogging often causes oxygen deficiency in the root systems of plants and severely inhibits plant growth. Formation of aerenchyma - interconnected spaces that facilitate the movement of gases between and within the aerial and submerged parts of plants - is an adaptive trait for coping with waterlogged conditions. Soybean (Glycine max) forms porous secondary tissues known as aerenchymatous phellem (AP), which are derived from the outermost cell layer of phellogen. To understand what factors other than waterlogging are involved in phellogen and AP formation, we examined how their formation in soybean seedlings was affected by darkness, CO2 deficiency and blockage of phloem transport. Methods Aerenchymatous phellem and phellogen formation were expressed as area ratios in cross-sections of hypocotyl. CO2 was depleted by use of calcium oxide and sodium hydroxide. Phloem transport was blocked by heat-girdling of hypocotyls. Sucrose levels were measured by spectrophotometry. Key Results Under light conditions, waterlogging induced the accumulation of high concentrations of sucrose in hypocotyls, followed by phellogen and AP formation in hypocotyls. Phellogen formation and AP formation were inhibited by darkness, CO2 deficiency and blockage of phloem transport. Phellogen formation and AP formation were also inhibited by excision of shoots above the epicotyl, but they recovered following application of sucrose (but not glucose or fructose application) to the cut surface. Conclusions The results demonstrate that sucrose derived from leaves is essential for AP and phellogen formation in soybean hypocotyls under waterlogged soil conditions. Maintenance of a high sucrose concentration is thus essential for the development of phellogen and AP and the differentiation of phellogen to AP.