Recently, osteogenic precursor cells were isolated from human dental follicles, which differentiate into cementoblast- or osteoblast-like cells under in vitro conditions after the induction with dexamethasone or insulin. However, mechanisms for osteogenic differentiation are not understood in detail. In a previous study, real-time RT-PCR results demonstrated molecular mechanisms in dental follicle cells (DFCs) during osteogenic differentiation that are different from those in bone-marrow-derived mesenchymal stem cells. We analysed gene expression profiles in DFCs before and after osteogenic differentiation with the Affymetrix GeneChip(R) Human Gene 1.0 ST Array. Transcripts of 98 genes were up-regulated after differentiation. These genes could be clustered into subcategories such as cell differentiation, cell morphogenesis, and skeletal development. Osteoblast-specific transcription factors like osterix and runx2 were constitutively expressed in differentiated DFCs. In contrast, the transcription factor ZBTB16, which promotes the osteoblastic differentiation of mesenchymal stem cells as an up-stream regulator of runx2, was differentially expressed after differentiation. Transcription factors NR4A3, KLF9 and TSC22D3, involved in the regulation of cellular development, were up-regulated as well. In conclusion, we present the first transcriptome of human DFCs before and after osteogenic differentiation. This study sheds new light on the complex mechanism of osteogenic differentiation in DFCs.