Abstract: Traditional villages represent a concentrated expression of the preservation and transmission of traditional culture within the context of rural revitalization, conveying important social, historical, and cultural values. Based on the perspective of spatial genes, this study selected 67 typical traditional villages in the Wuling Mountain Area in Southwest China as the case study. Between 2021 and 2024, by identifying and extracting spatial genes through the semi-structured interviews, Laddering Technique, and Landscape Pattern Index, and applying the geo-detector, the study explored the morphological characteristics and influencing mechanisms of traditional Tujia villages from the perspective of spatial genes of ecological-production-living. The findings are as follows: 1) the spatial genes of traditional Tujia villages in the Wuling Mountain Area exhibit distinct patterns. Ecological genes, categorized by natural environment and layout, demonstrate a transition from clustered to dispersed patterns from southern to northern regions. Production genes, categorized by location and cultivation patterns, show concentrated agricultural lands in the south and fragmented in the north. Living genes, divided into house plans, facades, and public buildings, reveal a higher prevalence of courtyards, stilted houses, and public buildings in northern areas. 2) The spatial genes have evolved through the combined influence of four key factors: natural environment, socioeconomic development, policy systems, and ethnic culture. 3) The integration of multi-source data and the application of both qualitative and quantitative approaches provide a comprehensive framework for analyzing traditional village form and their influencing mechanisms. This methodology offers valuable insights for developing sustainable strategies for the conservation of traditional Tujia traditional villages in this region.