DNA fragments actively transcribed at different developmental stages of Drosophila melanogaster were cloned. In situ hybridization was used to study their distribution in the polytene chromosomes. The clones were dot-hybridized to poly(A)+ RNA isolated at different developmental stages. Fifty-four of 67 clones were unique, 37 of them were localized. An irregular distribution of the cloned sequences was revealed. Twenty-three clones are localized in chromosome 3, six clones in the X-chromosome, and only eight in chromosome 2. There are transcriptionally active regions where, on a relatively small area of the chromosomes, a number of clones are localized (sections 18-19, 69-71, 82-85, 95-97). RNAs transcribed from the cloned sequences code for about 0.005 to 0.2% of the total poly(A)+ RNA.