In recent decades, the Qinghai Tibet Plateau (QTP) has experienced a rapid urban expansion, which has brought serious threats to the local ecology and environment. Location factors (e.g. topography, transportation and river, etc.) will affect the spatial pattern of urban expansion, and then change its impact on regional ecology and environment. Therefore, it is of great significance to understand the characteristics of the location factors of urban expansion for the protection of the ecology and environment on the QTP. In this study, we analyzed the impact of location factors on the urban expansion of the QTP at multiscale, that is, the whole region, the biome and the ecoregion. First, based on the long-term urban land dataset, we analyzed the urban expansion of the QTP. Second, the random forest model was used to analyze the location factors of urban expansion on the QTP. We found that the QTP experienced rapid urban expansion during 1990-2020. The urban land area of the whole plateau increased from 277.4 km² to 974.9 km², an increase of 2.5 times. Elevation was the most important location factor affecting the urban expansion of the QTP, and its importance was 30.1%. However, the importance of elevation to urban expansion declined from 1990 to 2020. In contrast, the railway factor was the location factor with the largest change in importance to urban expansion, increasing its importance by a factor of 3.9 from 2.15% to 10.56%. The study also found that the construction of the railway not only promoted the urban development of the QTP, but also affected nearly a third of the endangered species in the region. Therefore, in the future urban development process of the QTP, attention should be paid not only to the role of railways in driving urban expansion, but also to reducing the impact of railways on surrounding biodiversity, so as to promote regional sustainable development.