The physical characteristics of the genome of Puccinia graminis f. sp. tritici, the wheat stem rust fungus, were determined by reassociation kinetics. The results indicate that the haploid genome contains 67 Mb and consists of three classes of DNA sequences: (1) 64% unique; (2) 30% repetitive; and (3) 4% foldback. The repetitive sequences have a total complexity of 390 kb and are repeated an average of 52 times. The base composition was 45.3% G+C based on an analysis of the DNA melting temperature. The average amount of DNA per ungerminated urediniospore by diphenylamine assay, corrected for losses during extraction, was 435 fg. This was three times the expected value (147 fg) for dikaryotic spores with nuclei in the G1 phase of the cell cycle, an indication that the spores were in G2.