This study represents the physical map of the genome of the fruit fly, Drosophila melanogaster, as constructed by in situ hybridization and various methods for identifying overlapping clones in YACs and bacteriophage PI vectors. The current status of the physical map is presented with emphasis on resolution, biological content, and integration with the genetic map. The genomic localization's of 1200 YAC and 2400 Pl clones are tabulated, along with detailed protocols for the genetic manipulation of the clones. Also discussed is the general utility of the set of mapping clones for research in molecular genetics, development, neurobiology, and evolutionary biology.