The interaction of nano-fertilizers with commercially important crops can be a promising solution to increase both crop yield and quality. This study investigated the effect of iron oxide nanoparticles (Fe3O4 NPs) on four-week-old garden cress (Lepidium sativum L.) seedlings. Iron is an essential micronutrient for plants but is not always available in sufficient quantities, which can lead to chlorosis and even plant death. The seedlings were grown hydroponically, with three concentrations (1 mg/L, 5 mg/L, and 10 mg/L) of the NPs, alongside a control group with no additions. During the experiment, the following methods were employed: measurement of stem and root length, spectrophotometry to determine chlorophyll absorbance and concentration, and the RAPD technique to assess the genotoxicity of Fe3O4 NPs. The study demonstrated a significant increase in the shoot length of cress at all concentrations compared to the control group (p < 0.05; p < 0.01). The light absorption and chlorophyll concentration levels in the experimental groups significantly increased compared to the control group (p < 0.001). Genotoxicity analysis revealed that the genotoxic impact of the NPs on the garden cress genome was only 10%, a statistically insignificant level. The findings suggest that Fe3O4 NPs exhibit low genotoxicity and have the potential to enhance the growth and chlorophyll content of cress seedlings in hydroponic conditions.