As part of Coachella Valley Conservation Commission’s Wildlife Connectivity Assessment for Interstate 10 (I-10) and Closely Associated Transportation Infrastructure in the Banning Pass project, UC Davis’ Road Ecology Center evaluated wildlife use of existing structures and assessed traffic noise and light disturbance across a 15-mile segment of I-10 from Cabazon to California Route 62. Biodiversity data were collected using camera traps, eDNA, animal sign tracking, and roadkill records. Of 135 terrestrial mammal and herpetofauna species predicted within one mile of the highway, 101 were documented through >22,000 field observations. Connectivity actions were prioritized based on wildlife data. Noise and light models and field data were used to model the extent of I-10’s impacts, while land ownership informed the feasibility of connectivity improvements. Combined analyses produced recommendations identifying priority areas for wildlife crossing infrastructure and noise and light mitigation. While all existing structures had site specific recommendations, the existing Whitewater River and Stubbe Canyon Wash undercrossings were highlighted for enhancement based on their high biodiversity and land ownership. A new overcrossing site east of the Whitewater River was identified. This project demonstrates the value of empirical data in guiding targeted wildlife connectivity investments and provides a model for similar assessments.