Trophic niche is one of the important dimensions of niche theory. It reflects the nutritional needs of species, trophic level, functional status and nutritional relationship of interspecies in the ecosystem. The δ¹³C and δ¹⁵N in different tissues of animals reflect the species feeding information in different periods. Therefore, this is an important indicator for exploring feeding change, habitat resource utilization and trophic niche of interspecies. We analyzed the δ¹³C and δ¹⁵N ratio in hair and skeletal tissue of northern three-toed jerboa (Dipus sagitta) and mongolian five-toed jerboa (Allactaga sibirica) in 2017 in Alxa Desert, the Inner Mongolia, China. We compared the trophic level and niche change, feeding strategy and resource allocation mechanism of two rodent species. The results showed that, both at long-term (lifetime) and short-term (season) scales, two species presented an interspecific trophic niche separation. The trophic level of Dipus sagitta was significantly lower than Allactaga sibirica, but trophic niche area of Dipus. sagitta was larger than Allactaga sibirica. The feeding range and trophic niche area of Dipus sagitta and Allactaga sibirica showed a common trend of the maximum in spring, decreasing in summer, and increasing in autumn. The changes in trophic niches and feeding strategies of two rodents were consistent with the niche differentiation hypothesis. Two rodent species coexisted by trophic niche differentiation in Alax Desert.