Human cDNA probes for two new types of transforming growth factor-beta, TGF-beta 2 and TGF-beta 3, were used for mapping their cognate genes on human and mouse chromosomes by Southern blot analysis of somatic cell hybrid lines and, for the human loci, also by in situ chromosomal hybridization. For TGF-beta 2, a single site was found on the long arm of human chromosome 1, band 1q41, and on mouse chromosome 1, most likely in the known conserved syntenic region. For TGF-beta 3, the major site of hybridization, both on Southern filters and direct chromosome preparations, was at 14q24 in humans. This region is homologous in part to mouse chromosome 12, to which the murine beta 3 locus was mapped. These results indicate a wide dispersion of the TGF-beta gene family, with genes for TGF-beta 1 previously mapped by us to human chromosome 19q and mouse chromosome 7 and for inhibins alpha, beta B and beta A to human chromosomes 2q33-qter, 2cen-q13 and 7p15-p13, respectively.