Salt stress is a vital factor limiting nitrogen uptake and cotton growth in arid regions. The mechanisms underlying salt stress tolerance in cotton plants under high soil salinity have not been fully elucidated. Therefore, the aim of this study was to examine the proportion and mechanism of cotton nitrogen uptake under salt stress using the ¹⁵N isotope labeling technique. Cotton plants were grown in four undisturbed saline soils (1, 3, 6 and 9 dS m⁻¹), and the experiment was designed using the ENVIRO-GRO (E-G) model. The results showed that the dry matter of roots, stems and leaves of the cotton parts in slightly saline soil (C2, 3 dS m⁻¹) was not significant compared with the non-saline soil (C1, 1 dS m⁻¹). The cotton fruit grown in low-salinity soil (C2, 3 dS m⁻¹) had significantly higher dry matter than that grown in the other treatments, implying that cotton plants grown in 3 dS m⁻¹ soil have the best nitrogen uptake and salt tolerance. Cotton plants grown in weakly (C3, 6 dS m⁻¹) and moderately (C4, 9 dS m⁻¹) saline soils exhibited premature senescence. The distribution of total nitrogen and nitrate content in cotton was the best explanatory variable of total ¹⁵N recovery, of which cotton ¹⁵N recovery was between 26.1% and 47.2%, and soil ¹⁵N recovery was between 7.7% and 14.9%. Our findings provide guidance for further exploitation and utilization of saline soil resources and sustainable development of the agricultural soil ecosystem in arid regions.