A Drosophila melanogaster gene encoding a protein with greater than 80% sequence identity to the beta subunits of mammalian guanine nucleotide-binding regulatory proteins (G proteins) has been cloned. The gene, which was mapped to 13F on the X chromosome by in situ hybridization, was cloned from a Drosophila genomic library by using a bovine transducin beta-subunit cDNA probe. Genomic DNA blot hybridization analysis indicated that there is a single Drosophila G-protein beta-subunit gene. Multiple transcripts were detected throughout development; in adult flies the mRNA is expressed at higher levels in heads than in bodies. The proposed coding region is uninterrupted by introns, but there is evidence for differential mRNA splicing in the 5' nontranslated region.