Carbon isotope discrimination (Δ) has been proposed as a method for evaluating transpiration efficiency in C3 plants. It is important to study the association of Δ in different parts with the long‐term water use efficiency (WUE). In this paper, plant height, Δ in leaf water soluble carbohydrates (ΔWSC), photosynthetic gas exchange parameters, Δ in aboveground shoot (ΔAGS), stem/leaf weight ratio, aboveground biomass (AGB), evapotranspiration, and WUE were determined in a collection of 10 alfalfa (Medicago sativa L.) genotypes under three water treatments (230 [T1], 460 [T2], and 700 mm irrigation [T3]) in a Ningxia central desert steppe in 2012 and 2013. Significant interactions were found for AGB and WUE among year, treatment, and genotype, whereas significant interactions of year × treatment and year × genotype were only found for ΔAGS but not for ΔWSC. Generally, more water input increased the values of the traits tested except for WUE and single‐leaf intrinsic WUE (Pn/Gs). More irrigated water input resulted in increase in AGB, ΔAGS, and ΔWSC and decrease in Pn/Gs. The greatest WUE was recorded under T2 in 2012. Significant and positive correlations were observed between AGB and ΔAGS (r = 0.588, 0.574, and 0.386 under T1, T2, and T3, respectively). Under T2 and T3, ΔWSC correlated negatively with WUE (r = −0.383 and 0.602, respectively). Ningmu No. 1 was found to have the greatest WUE (18.34 kg ha−1 mm−1) and yield (17.71 Mg ha−1) across water treatments and years. The findings suggest that in the semiarid areas applying irrigation of 230 to 700 mm yr−1, Δ in aboveground shoots or in leaf water‐soluble carbohydrates at the flowering stage seems to be a useful indicators reflecting aboveground biomass and integrated WUE for alfalfa.