OBJECTIVES: Tooth agenesis is one of the most common developmental anomalies in humans. Genetic and environmental factors may be of etiological importance in this condition. Among genes involved in tooth morphogenesis, mutations in PAX9, MSX1, AXIN2, WNT10a, and EDA genes have been associated with tooth agenesis. The aim of our study was to investigate the relationship between the PAX9 gene variants and tooth agenesis in the Czech population. METHODS: The selected regions of the PAX9 gene were analysed by direct sequencing and compared with the reference sequence from the GenBank online database (NCBI). RESULTS: We found several novel variants in the PAX9 gene, e.g. insertion g.5100_5101insC (rs11373281) with simultaneous substitution g.5272C>G (rs4904155) in exon 1, and mutation g.10934C>T (Gly203Gly, rs61754301) in exon 3. In subjects with full dentition we observed polymorphisms g.10276A>G (rs12882923) and g.10289A>G (rs12883049) in IVS2 (intervening sequence 2) previously related to tooth agenesis in Polish study. CONCLUSIONS: In our study we excluded a direct effect of rs12882923 and rs12883049 polymorphisms on the dental agenesis in the Czech population. All described PAX9 genetic variants were present both in patients with tooth agenesis and controls. We expect that tooth agenesis in our cohort of patients is caused by mutations in regions different from PAX9 exons analyzed in our study.