The productivity of short‐rotation coppice (SRC) plantations with poplar (Populus spp.) strongly depends on soil water availability, which limits the future development of its cultivation, and makes the study of the transpirational water loss particularly timely under the ongoing climate change (more frequent drought and floods). This study assesses the transpiration at different scales (leaf, tree and stand) of four poplar genotypes belonging to different species and from a different genetic background grown under an SRC regime. Measurements were performed for an entire growing season during the third year of the third rotation in a commercial scale multigenotype SRC plantation in Flanders (Belgium). Measurements at leaf level were performed on specific days with a contrasted evaporative demand, temperature and incoming shortwave radiation and included stomatal conductance, stem and leaf water potential. Leaf transpiration and leaf hydraulic conductance were obtained from these measurements. To determine the transpiration at the tree level, single‐stem sap flow using the stem heat balance (SHB) method and daily stem diameter variations were measured during the entire growing season. Sap flow‐based canopy transpiration (E
c), seasonal dry biomass yield, and water use efficiency (WUE; g aboveground dry matter/kg water transpired) of the four poplar genotypes were also calculated. The genotypes had contrasting physiological responses to environmental drivers and to soil conditions. Sap flow was tightly linked to the phenological stage of the trees and to the environmental variables (photosynthetically active radiation and vapor pressure deficit). The total E
c for the 2016 growing season was of 334, 350, 483 and 618 mm for the four poplar genotypes, Bakan, Koster, Oudenberg and Grimminge, respectively. The differences in physiological traits and in transpiration of the four genotypes resulted in different responses of WUE.