We are in the process of aligning fragments of the 40 Mb Dictyostelium genome along the chromosomes after cloning them in Yeast Artificial Chromosomes (YACs). We have chosen a five fold redundant mapping set of 1016 YACs from a larger set of 2500 YACs that appear to give a faithful representation of the total genome. By probing blots of an arrayed set of electrophoretically separated YACs, we can group them into sets of 4 or 5 overlapping YACs that all carry a specific gene. The markers are positioned to within 20 by restriction mapping the cloned fragments. The YACs have inserts that average 200 kb and so can be recognized by probes from linked genes. Contigs are sequentially extended as new genes are added. Linkage is confirmed by megabase mapping restriction sites around the genes in the YACs and comparing the composite map to the genomic restriction map. Contigs are assigned to one of the 7 chromosomes that range from 3 Mb to 9 Mb when they carry a genetically mapped marker. We are also generating an independent genetic map from an isogenic set of Dictyostelium strains in which a plasmid carrying ApaI and SstII sites is randomly integrated to generate Restriction Fragment Length Polymorphisms (RFLPs). A set of 150 isogenic RFLP strains probed with our marker genes will map each chromosome to within a few hundred kb. Comparison of the RFLP maps to those generated from ordered YAC fragments will provide the necessary confirmation to ensure the accuracy of the assignments. An ordered set of a few hundred YACs covering the Dictyostelium genome will allow rapid isolation and mapping of new genes as well as provide a ready source of genomic clones for the community. It will be important to demonstrate that a total genome map can be efficiently finished. Moreover, the approaches and techniques that are successful for the Dictyostelium genome should be useful to projects focused on other genomes including that of humans.