Geothermal resources, as a renewable and clean source of energy, are attracting widespread attention globally. In China, most medium to high enthalpy geothermal resources are developed in the Tibetan Plateau, especially in the rift zone of Southern Tibet. To further investigate the genesis mechanisms of geothermal resources, this study collected geothermal spring samples from the Cuona-Woka rift zone in Southern Tibet. Hydrochemical and isotopic characteristics were analyzed to reveal the origin, evolution, reservoir temperature, and circulation mechanisms of the geothermal waters. The exposed temperature of the geothermal spring ranges from 34 to 67 °C. Compared with HCO3-Ca·Na and HCO3-Na type samples, HCO3·Cl-Na and HCO3·SO4-Na type samples have higher concentrations of Cl− and trace elements. The geothermal springs are recharged by a mixture of meteoric water, snow-melt water, and magmatic water. The recharge areas had an elevation range from 5091 to 6087 m, with temperatures from −5 to −10 °C. The hydrochemical processes are dominated by silicate and carbonate dissolution, and positive cation exchange, with local gypsum dissolution. Solute geothermometers, silica-enthalpy mixing models, and geothermal conceptual model indicate that there exist shallow geothermal reservoirs (temperature = 137–162 °C) mixed by surficial cold groundwater and initial deep geothermal reservoirs (temperature = 196–212 °C), respectively. Finally, two genesis models of geothermal waters are proposed: the deep melt mixing and heating model (Type A) and the high-temperature steam heating model (Type B). The achievements of this study would provide valuable insight into geothermal research and exploitation in the Tibetan Plateau.