The Drosophila melanogaster ovo gene is required for survival and differentiation of female germline cells, apparently playing a role in germline sex determination. We recovered 60 kb of genomic DNA from its genetic location at 4E1,2 on the X chromosome. A transcription unit coding for an apparently female-specific germline-dependent 5-kb poly(A)+ RNA size class is located substantially in a 7-kb region, within which three DNA-detectable lesions for mutations that inactivate the ovo function are located at two sites approximately 4 kb apart. The breakpoint of a deficiency that removes the neighboring lethal complementation group shavenbaby (svb) but leaves the ovo function intact maps approximately 5 kb to the molecular left of the leftmost ovo mutant site. A class of mutations that inactivates both the svb function and the ovo function affects genomic DNA between the two ovo sites. Sequences required for the two genetic functions are partly overlapping. In spite of this overlap, P element-mediated gene transfer of a 10-kb genomic DNA segment containing the 5-kb poly(A)+ RNA transcription unit rescues the female sterility phenotypes of ovo mutations, but not the svb lethality.