Spatial definitions of study areas for specific projects are of crucial importance for these projects. It is necessary to come out from the aims of the project for spatial definition of mountain and under-mountain areas in South- Bohemian Region. There are many ways of solution and the definition should be strictly connected with the structured goals of this project. The methods and usage of criteria for definition of study areas will depend on aim identification. There are several possibilities of spatial definition: one- or more-criteria selection, base data sources or complex data sources, findings of one or more sciences – especially geomorphology, geography and agricultural regionalization.