AbstractMost of the Brazilian soils, especially in Cerrado biome, have high acidity and toxic aluminium, making the application of limestone an indispensable technique to obtain high yield in soybean crop. The hypothesis of this study was that contrasting base saturation provides changes in water‐use efficiency (WUE) and agronomic traits of soybean populations. Thus, we evaluated the WUE and agronomic performance of F3 soybean populations under different base saturation levels. Each experiment was conducted in a randomized complete blocks design with three replicates and 10 F3 populations (P1, P2, P3, P4, P5, P6, P7, P8, P9 and P10) for two crop seasons. Segregating populations were obtained by bulk method. In the first experiment, F3 populations were evaluated without correction of base saturation (V = 30%), while in the second experiment, limestone was applied three months before sowing to increase base saturation to 60%. The traits evaluated were as follows: calcium, magnesium and potassium contents, water‐use efficiency (WUE), plant height, first pod height, main stem diameter and grain yield. Our findings indicate that WUE and agronomic performance of F3 soybean populations depends on the saturation condition. We identified the P4 and P9 populations as the most promising for use in breeding programmes aiming at higher tolerance to abiotic stress caused by low base saturation in soil.