The Hybrid sterility 1 (Hst1) gene affects fertility of male hybrids between certain laboratory strains (such as C57BL/10) and some Mus musculus musculus mice by causing a breakdown of spermatogenesis at the stage of primary spermatocytes. In the process of positional cloning of the Hst1 gene, we generated a contig of bacterial artificial chromosomes and subsequently a low coverage sequence of the candidate region of the 129S1/SvImJ strain. Development of new genetic markers allowed us to narrow the Hst1 region from 580 to 360 kb. The products of two genes from this region, TATA-binding protein (Tbp) and proteasome subunit beta 1 (Psmb1), accumulate during spermatogenesis. These proteins have been described previously as having conserved C-terminal sequences and species-specific N-termini. We evaluated the candidacy of these genes for Hst1 by allelic sequencing and by real-time semiquantitative reverse-transcription PCR of testicular mRNAs. © 2005 The Linnean Society of London, Biological Journal of the Linnean Society, 2005, 84, 637–641.