We cloned a Drosophila melanogaster ras gene (Dmras64B) on the basis of its homology to the ras oncogen from Harvey murine sarcoma virus. This gene mapped at chromosomal position 64B on the left arm of the third chromosome. Sequencing of Dmras64B revealed extensive amino acid homology with the proteins encoded by the human and Saccharomyces cerevisiae ras genes. The coding region of the Drosophila gene is interrupted by two introns located in different positions with respect to its human counterpart. Dmras64B encodes three different RNAs (1.6, 2.1, and 2.6 kilobases long) that are constantly expressed throughout the development of the fly.The transformed phenotype of tumor cells has been associated with the activation of normal cellular oncogenes by a variety of different processes. In the case of the ras genes, which were originally isolated from the Harvey and Kirsten murine sarcoma viruses, this activation results from somatic mutation, giving rise to a protein (p21) with an altered amino acid sequence (1,14,(16)(17)(18)(19)21). Elevated expression of the ras proteins can also transform animal cells (2, 6). Nothing is known, however, about the molecular mechanisms by which the p21 ras-encoded protein causes the transformed phenotype.To gain more insight into the biochemical and cellular roles of the ras gene family, we have begun a study of these genes in Drosophila melanogaster, taking advantage of the fact that they are so highly conserved during evolution (15). A Charon 4A Drosophila genomic library (obtained from T. Maniatis) was screened by using the BglII DNA fragment containing the Harvey murine sarcoma virus ras (Ha-ras) gene as a-probe (3, 8). The hybridization was done in 30% formamide-0.75 M NaCl-0.075 M sodium citrate-0.2% polyvinylpyrrolidone-0.2% bovine serum albumin-0.2% Ficoll-1.0% sodium dodecyl sulfate-denatured calf thymus DNA (100 ,ug/ml) at 37°C for 12 h. The filters were then washed twice with 0.75 M NaCl-0.075 M sodium citrate at room temperature and twice with the same buffer at 60QC. Of the several positive clones obtained, one, designated X-2A, was studied in detail. Figure 1 shows the restriction map of a PstI subfragment, obtained from the X clone, which contains all the sequences homologous to the Ha-ras oncogene. This DNA fragment was sequenced by using the protocol of Maxam and Gilbert (11) (see Fig. 1 for a diagram of the sequencing strategy), and the amino acid sequence of the different open reading frames was compared with that of the human and Saccharomyces cerevisiae ras oncogenes. We found three regions of homology interrupted by two stretches of nonhomologous sequences. These regions of nonhomologous sequences are likely to be introns because protein termination codons exist in all three possible reading frames. The precise location of the introns within the protein-coding regions was assigned on the basis of three different criteria: the existence of a consensus donor and acceptor splice site sequence, the maintenance of an open * Corresponding author. reading frame, and...