Ninety kilobase pairs (kb) of DNA have been isolated from the 3B region of the X chromosome of Drosophila melanogaster. Previous cytogenetic analyses have placed a gene required for rhythmic behavior (per) in this chromosomal interval. Physical characterization of a series of chromosomal rearrangements altering per locus activity indicates that DNA affecting behavioral rhythms is found in a 7.1-kb HindIII fragment. A single 4.5-kb poly(A)(+) RNA is transcribed from this DNA in wild-type pupae and adult flies. The transcript is eliminated by a per mutant that retains some rhythmic activity, but this mutant substitutes two novel transcripts, 11.5 kb and 0.9 kb. It is suggested that the new poly(A)(+) transcripts provide residual per locus activity.