Orc5p is a subunit of the origin recognition complex in the budding yeast Saccharomyces cerevisiae, which has been shown to play a critical role in both chromosomal DNA replication and transcriptional silencing. We have cloned cDNAs from both human and fission yeast Schizosaccharomyces pombe that encode proteins homologous to the budding yeast and Drosophila Orc5p. Human Orc5p showed 35.1, 22.3, and 19.4% identity to the Drosophila, S. pombe, and S. cerevisiae Orc5p, respectively. We have localized the human ORC5 gene (ORC5L) to chromosome 7 using Southern and PCR analysis of DNA isolated from a panel of human/rodent somatic cell hybrids and mapped the gene locus to 7q22 using fluorescence in situ hybridization. We have identified a YAC clone that contains human ORC5L and maps to chromosome band 7q22.1. We have identified the S. pombe ORC5 gene and located it in a cosmid mapped on chromosome II.