Members of the T-box family are known to play critical roles in the embryonic development of most animal species. Recently, we have isolated its new mammalian member, Tbr2, from mouse embryonic brain. We have also shown that the expression patterns of Tbr2 and the closely related Tbr1 appear to be reciprocal in the developing brain; Tbr2 is expressed in mesencephalon and rhombencephalon, but expression of Tbr1 is restricted to telencephalon. To investigate possible structural and functional relationships of Tbr2 and other T-box containing genes, we analyzed genomic organization of the murine Tbr2 gene. The Tbr2 gene is composed of six exons (1353, 155, 122, 159, 62 and 1035bp), and five introns (920, 643, 602, 85 and 2036bp). This exon/intron organization is very similar to that of Tbr1. We also analyzed the 3.9kb sequence of the 5' promoter region flanking the Tbr2 gene and the corresponding region of the Tbr1 gene. The sites for Brn-2 and Tst-1 were found in the promoter of Tbr2 but not Tbr1. On the contrary, there were eight HNF-3beta binding sites in the Tbr1 gene promoter but only three in the Tbr2 promoter. The differential presence of putative binding sites for these brain-specific transcription factors may explain the reciprocal expression of Tbr1 and Tbr2. Furthermore, a single chromosomal locus for mouse Tbr2 was assigned to 9F3 by fluorescence in-situ hybridization 1.