The preparation of a high-resolution, physical map of the human genome is one of the current challenges of molecular genetics today. The overall approaches and the methodologies to complete this task have been developed to some extent, but few mapping strategies have gone from model systems to actual genomic mapping. We propose here to 1) develop the technology necessary to perform this task. 2) implement its use by preparing a map of a limited region of the genome and 3) investigate the application of this map to problems in human genetics. We will concentrate on the improvement of techniques for the production of libraries containing clones localized to the region of interest, and on the implementation of novel restriction cleavage strategies that pro- duce a physical map of the long arm of chromosome 5 at 300-kb resolution. The map will consist of unique probes of known order and distance that have been linked by megabase restriction mapping. Emphasis will be placed on bands 5423-31, an area of biomedical significance, containing genes that are involved in mutagen-induced leukemia. Lastly, we will investigate the use of the physical map to identify structural alterations in the human genome, addressing questions about the general reproducibility of this map from individual to individual, and the use of this map to identify translocations at 5q31 in the cells of patients with leukemia.