Evaluation of various root traits and determination of their genetic control in response to water deficit stress is crucial for improving and stabilizing grain yield worldwide. The present study aimed at evaluating the gene action, heritability and genetic variances of grain yield and root morphological traits under water deficit conditions in mature maize. Seeds of seven maize genotypes, including B73 (maternal inbred line), MO17 (paternal inbred line), F1 (SC704), F2, BC1, BC2 and F3 were sown in PVC tubes and grown under field water capacity. The maize plants were then subjected to three water regimes of 100% FC, 55% and 75% available water depletion at 5–6 leaf stage and continued to silk emergence in 2018 and 2019 (two years). The generation mean analysis results revealed that non-additive effects were greater than the additive effects. A high narrow sense heritability and additive variance were found for root dry weight and seminal root number, suggesting that selection in the parents’ inbred lines or early segregating generations could be an efficient method for genetic improvement of the traits under deficit irrigation conditions. Contrarily, selection for grain yield, root volume, primary root length, root surface area, root diameter and root fresh weight should be delayed for several generations until a high level of gene fixation is attained. Based on correlation analysis and percent change of the studied traits under deficit irrigation conditions, SC704 and MO17 could be used as a source for water deficit tolerance in maize breeding programs. Overall, the results suggested that the selection of root traits linked to better plant growth has great potential to alleviate water deficit stress, contributing to sustainable maize production in drought-prone areas.