Mesenchymal stem cells (MSCs), which can differentiate into tri-lineage (osteoblast, adipocyte, and chondrocyte) and suppress inflammation, are promising tools for regenerative medicine. MSCs are phenotypically diverse based on their tissue origins. However, the mechanisms underlying cell-type-specific gene expression patterns are not fully understood due to the lack of suitable strategy to identify the diversity. In this study, we investigated gene expression programs and chromatin accessibilities of MSCs by whole-transcriptome RNA-seq analysis and an assay for transposase-accessible chromatin using sequencing (ATAC-seq). We isolated MSCs from four tissues (femoral and vertebral bone marrow, adipose tissue, and lung) and analysed their molecular signatures. RNA-seq identified the expression of MSC markers and both RNA-seq and ATAC-seq successfully clustered the MSCs based on their tissue origins. Interestingly, clustering based on tissue origin was more accurate with chromatin accessibility signatures than with transcriptome profiles. Furthermore, we identified transcription factors potentially involved in establishing cell-type specific chromatin structures. Thus, epigenome analysis is useful to analyse MSC identity and can be utilized to characterize these cells for clinical use.