The responses of Trifolium repens L. seedlings in growth, biomass and photosynthetic characteristics to 2 light intensity levels and 5 nitrogen application rate levels were hydroponically studied in growth chamber. The results showed that the stem and leaf biomass of Trifolium repens L. seedlings increased as the nitrogen concentrations increased from 5 to 15 mmol?L-1 and then decreased with the nitrogen concentrations moved from 20 to 25 mmol?L-1, but the root biomass and root/shoot ratio of Trifolium repens L. seedlings decreased with the increasing nitrogen concentration. The biomass of roots, stems, leaves and whole plant under low light decreased by 67.8%, 29.9%, 42.5% and 45.2%, respectively, and the root/shoot ratios decreased, but the specific leaf areas (SLA) was opposite. The root volumes of Trifolium repens L. seedlings decreased with the increasing nitrogen concentrations, when the root volume of Trifolium repens L. seedlings at the higher light intensity were higher than those at the lower light intensity. The root surface areas, root diameters and root lengths of Trifolium repens L. seedlings increased as the nitrogen concentrations from 5 to 15 mmol?L-1 and then decreased with the nitrogen concentrations moved from 20 to 25 mmol?L-1. The effects of light intensity on root length and root diameter of Trifolium repens L. seedlings significantly differed, but the root surface areas did not significantly differed. The photosynthetic rates (Pn) of Trifolium repens L. seedlings at the higher light intensity were higher than those at the lower light intensity, and the Pn increased as the nitrogen concentrations increased from 5 to 15 mmol?L-1 and then decreased as the nitrogen concentrations moved from 20 to 25 mmol?L-1. The effects of light intensity on stomatal conductance (Gs), intercellular CO2 concentration (Ci) and transpiration rate (Tr) did not significantly differed, but the effects of nitrogen concentration on Gs, Ci and Tr were significant. The effect of light and nitrogen as well their interaction on seedling growth and photosynthesis of Trifolium repens L. seedlings were significant. Light and nitrogen shortages all inhibited the growth capacity of the seedling, but the seedlings also had the capacities to acclimatize and adapt to the disadvantageous environments.