The large family of signal transducing proteins known as G proteins are heterotrimers that dissociate into an independent alpha-subunit and beta gamma-subunit complex after ligand binding or other stimulation. For G alpha, at least 30 distinct sequences representing 10 different classes have been identified. On the other hand, cDNAs for only three G beta-subunit genes have been isolated so far. All three of the G beta genes have been chromosomally mapped in the human, but only two in the mouse. Using a human retinal cDNA for the third G protein beta-subunit, we have mapped the corresponding gene, termed Gnb-3, to mouse Chromosome 6 with somatic cell hybrids and have positioned it distal to but near the marker Raf-1 by analysis of the progeny of three genetic crosses.