Sustaining agricultural productivity and ensuring food security through adaption to climate change are main challenges in modern agriculture. Leveraging the connection between roots and their growth environment could be an effective approach to addressing these issues. Thus, this study aimed to quantify the root biomass production and depth distribution among different maize varieties under contrasting water regimes and to investigate the influence of variety and water regime on the relationship between the biomass allocation to above- and belowground biomass. Root traits (biomass, root distribution, length, diameter) to 100 cm soil depth and yields were assessed in 10 different maize varieties grown at four experimental sites, each with two contrasting water regimes. The average root biomass of maize down to 100 cm was found to be 3.2 ± 1.2 Mg ha−1 across all sites and treatments. Well-watered conditions led to 29% higher root biomass in the top soil (0-30 cm) compared to water deficit conditions. In the subsoil (30–100 cm) however, root biomass was 25% higher under water deficit and increased significantly with the number of water stress days. Varieties differed site-specifically by 59 to 71% for root biomass and by 10 to 48% for yield under well-watered conditions. Under water deficit conditions varieties differed by 63 to 72% for root biomass and by 20 to 74% for yield. Subsoil roots may play a pivotal role for water acquisition during water stress periods and may contribute more than topsoil roots and above ground biomass to the accrual of soil carbon.