A comprehensively understanding of the trade-offs between ecosystem services is a basic requirement for effective ecosystem management and extensively regional development. However, current ecosystem service-related research is less concerned about the gradient effect in ecosystem service trade-off relationships. Taking the Daning River Basin in the Three Gorges Reservoir area as the study area, this study aims to explore the spatial variability characteristics for ecosystem services under three environmental gradients of elevation, precipitation, and vegetation cover fraction, and to analyze the gradient effects of ecosystem services and their trade-offs. Four ecosystem services, including food production, carbon storage, soil retention, and water yield, were assessed and mapped in 2018 by applying the Carnegie Ames Stanford Approach, Revised Universal Soil Loss Equation, and Integrated Valuation of Ecosystem Services and Tradeoffs model. Root mean squared deviation was used to explore the gradient effects of ecosystem service trade-off relationships. The results show that ecosystem services in the Daning River basin were significantly spatial heterogeneity. Specifically, food production was spatially distributed in the southern and central areas and was lower in the northern area, carbon storage decreased from northwest to southeast, high soil retention and water yield were mainly distributed in the west. Elevation, precipitation, and vegetation cover fraction had significantly influenced on ecosystem services. Generally, carbon storage and soil retention were increased with elevation, precipitation, and vegetation cover fraction increase, whereas food production decreased. From the perspective of ecosystem service trade-off relationships, the trade-off relationship between carbon storage and water yield was weak, soil retention and food production showed a moderate trade-off relationship. For most of the ecosystem services, their trade-off relationships were always strong. There was no obvious rule of ecosystem service relationships along the elevation gradient because of interactions among multiple environmental gradients. The increasing elevation only significantly enhanced the trade-off relationships of carbon storage-soil retention, and soil retention-food production, but weakened the relationships between carbon storage and water yield. Moreover, increasing vegetation cover fraction significantly enhanced ecosystem service trade-offs, whereas the increased precipitation had an obvious negative effect. Future ecological restoration projects intended to coordinate relationships among ecosystem services and promote ecological function, should pay more attention to ecosystem service trade-offs in different regions of elevation, precipitation, and vegetation cover fraction gradients. Overall, this study provides a special study perspective of ecosystem service and trade-offs, and the results obtained in this study could also be used to guideline the planning strategy of ecological restoration.