Using a mouse GDF-9 cDNA as a probe, over 20 kb of sequence encompassing the GDF-9 gene was isolated from a mouse 129SvEv genomic library. Sequence analysis of the exons, exon-intron boundaries, and 5'- and 3'-non-translated regions was used to establish the structure of the mouse GDF-9 gene. The GDF-9 gene is encoded by two exons separated by a 2.9 kb intron. Multiple putative transcription start sites are detected between 31 and 57 bp upstream of the start site of translation and a putative polyadenylation signal lies 342 bp 3' of the end of translation. Knowledge of the GDF-9 gene structure will enable us to further understand the role of GDF-9 in ovarian physiology and development.