The gully region of the Loess Plateau is a landform with an extremely uneven distribution of water resources. In the dry season, water shortages seriously affect the property and lives of residents. During the rainy season, excessive rainfall leads to a series of geological disasters such as landslides and collapses, mudslides, and soil erosion. The subsurface flow in the gully of the Loess Plateau provides a new way to solve this problem. The premise of redistributing water resources by using subsurface flow effectively is to find out the temporal and spatial distribution of water in gullies and its replenishment methods. However, there have been no related reports about it in recent years. In this study, a typical gully (Jiulongquan Gully, Yan’an, Shaanxi Province, China) was studied by long-term positioning monitoring and isotope tracing. The direction and proportion of hydrological cycle transformation in each water body were quantitatively analyzed using a multi-terminal mixed model. The gully water transformation relationship and the contribution rate of water resource types to gully water were identified. The result shows that the subsurface flow in slope soil was mainly concentrated in the upper soil from 0-100 cm, while the subsurface flow in the gully was mainly generated in the upper soil from 0-120 cm. For deep soil-water, there was no significant difference in the water sources at different locations of gullies. The main recharge ratios of precipitation, surface water, and groundwater to the deep soil-water were 55.39%-60.10%, 10.48%-21.85%, and 22.76%-29.42%, respectively. For shallow soil, the difference was more obvious. Compared with the upstream of the gully, the proportion of precipitation supply for the subsurface flow in the midstream of the gully increased from 38.82% to 56.42%, and the proportion in the downstream of the gully increased from 2.54% to 17.89%. Upstream of the gully, the main recharge ratios of precipitation, surface water, and groundwater to the subsurface flow were 38.82%, 58.64%, and 2.54%, respectively. For the midstream of the gully, the main recharge ratios of precipitation, surface water, and groundwater to the subsurface flow were 56.42%, 41.34%, and 2.24%, respectively. For the downstream of the gully, the main recharge ratios of precipitation, surface water, and groundwater to the subsurface flow were 8.56%, 48.51%, and 42.93%, respectively. By installing the intercepts on the slope (100 cm) and in the gully (120 cm) and the reservoirs upstream and downstream of the gully, it is expected that the subsurface flow can be used to effectively trap soil-water and regulate water distribution in the gully.