Sequence homology between the genomes of a hamster papovavirus (HaPV), polyoma virus (Py), and Simian virus 40 (SV40) has been studied by filter hybridization and electron microscopy under conditions of varying stringency. Hybrids between the HaPV and SV40 DNAs could be demonstrated only under nonstringent conditions. The region of highest homology was mapped in the early region of the SV40 genome. Extensive homology was detected between the genomes of HaPV and Py under stringent hybridization conditions, indicating at least 80% base matching in the regions of strongest sequence homology. These sequences were localized within both the early region and the late region of the Py genome. The homologous DNA segments mapped in the Py and the SV40 genomes are among the most strongly conserved regions in the polyoma (miopapova)-virus group.