Maintaining root function is crucial for favorable plant growth under flooding. The genetic variation in the response of root development to flooding is unclear, because measurement of root growth is time consuming, especially with numerous lines. To overcome the methodological problems and to reveal the effect of flooding on root development and its genetic variation, we developed a new capillary watering system without soil medium and raised cotyledon-stage seedlings of 92 soybean lines with and without flooding. After 7 days of flooding, dry weights (DW) and root characteristics were determined and the results were compared with those in non-flooded plants. The root DW decreased linearly with decreasing total root length and root surface area, and the degree of damage varied greatly among lines. Short-term flooding inhibited root elongation and branching, but not in flood-tolerant lines.