Introduction: Hydroponics enables precise control over nutrient delivery, optimized water requirements and growing conditions. The combination of nanotechnology and hydroponics paves the way towards sustainable agriculture with less environmental footprints. We investigated the effects of nano urea on the model plantArabidopsis thalianain hydroponics. Methods: A growth experiment in a nitrogen-free hydroponic medium compared the effects of a liquid nano urea formulation (NUF) marketed by Indian Farmers Fertilizer Cooperative (IFFCO) to an equimolar bulk urea. Transcriptome analysis identified the molecular mechanisms of growth enhancement. Dynamic light scattering and transmission electron microscopy confirmed the negative surface charge and sub-100 nm size of NUF, correlating its uptake and distribution in the plant. Results and discussion: A two-week growth in the hydroponic medium with 70 μM NUF led to a 20% higher biomass and 16% higher chlorophyll content than a medium with 70 μM urea. Higher doses of NUF inhibited growth, whereas higher equivalent urea doses did not. NUF led to the differential expression of more genes than urea at 12 h to seven days of treatment. Nitrogen assimilation, growth, photosynthesis, and stress tolerance genes showed higher transcript levels in NUF than in urea. On the other hand, NUF led to greater suppression of many negative growth-regulating genes. After seven days of treatment, chlorophyll biosynthesis genes got up-regulated, while chlorophyll catabolism genes got down-regulated at higher levels by NUF than by urea, correlating with the higher chlorophyll content of NUF-treated seedlings. In conclusion, NUF outperformed equimolar urea for the growth promotion ofA. thalianaat a low concentration in hydroponics, leading to a greater regulation of genes for nitrogen metabolism and chlorophyll biosynthesis. Our results suggest a potential use of NUF as a nitrogen fertilizer for hydroponic agriculture.