Owing to low precipitation and high inter-annual variability, the Loess hilly and valley region experiences frequent drought, which significantly affects plant growth. Studying the response mechanism of plants to water stress is important with respect to improving water-use efficiency (WUE) in this arid and semi-arid area. In the present research, Bothriochloa ischaemum (L.) Keng., a typical native species in the Loess hilly-gully region, was selected to investigate biomass accumulation and allocation, instantaneous WUE, stable carbon isotope composition (δ¹³C), and stable carbon isotope discrimination (Δ¹³C) in different organs under three water regimes: control [CK, 80% field capacity (FC)], mild water stress (MS, 60% FC), and severe stress (SS, 40% FC). Photosynthetic parameters of B. ischaemum were measured at 09:00-11:00 on sunny windless days (Aug 3, 10, and 13,2014), using a portable photosynthesis determination system (Li-6400; Li-Cor, USA). On Nov 11, when the experiment ended, the new leaf, old leaf, stem, fine root, and coarse root parts of seedlings were sampled carefully and weighed (with accuracy 0.001 g) after drying at 65℃. The root-to-shoot ratio was then calculated as the ratio of the belowground dry biomass to aboveground dry biomass. The δ¹³C of different plant parts was measured using a CCIA-36d-EP Carbon isotope ratio mass spectrometer (Los Gatos Research, USA). The Δ¹³C of different plant parts was calculated based on δ¹³C and the carbon isotope ratio of atmospheric CO₂. Many environmental factors that might affect WUE were analyzed to explore the response mechanisms of B. ischaemum under drought conditions. The following results were obtained. (1) The total biomass of B. ischaemum was significantly decreased under the SS condition, whereas the root-to-shoot ratio and fine root biomass were significantly increased under this water regime. (2) The δ¹³C and Δ¹³C in different plant parts showed different trends. Drought stress led to an increase in the δ¹³C value and decrease in the Δ¹³C value. There was no significant difference in the δ¹³C and Δ¹³C of different plant parts under the SS condition. The δ¹³C values under CK and MS conditions were in the order of fine root > coarse root > old leaf > new leaf > stem, and fine root > new leaf > old leaf > coarse root > stem, respectively. Furthermore, the Δ¹³C values under CK and MS conditions were all in the order of root < leaf δ¹³C and Δ¹³C values of new leaves had the highest correlation coefficients with respect to WUE, which indicated that the stable carbon isotope technique is suitable for the determination of the WUE of B. ischaemum. (4) The WUE was affected by different environmental factors under different water regimes. Leaf temperature, atmospheric water vapor pressure, and air temperature were the most important factors affecting WUE under CK, MS, and SS conditions, respectively. This research provides a theoretical basis for the feasibility of using the stable carbon isotope method to determine the WUE of B. ischaemum, and for elucidating the mechanism underlying the response of B. ischaemum to water stress.