The nonpathogenic human adeno-associated virus (AAV) has developed a mechanism to integrate its genome into human chromosome 19 at 19q13.4 (termed AAVS1), thereby establishing latency. Here, we provide evidence that the chromosomal signals required for site-specific integration are conserved in the mouse genome proximal to the recently identified Mbs85 gene. These sequence motifs can be specifically nicked by the viral Rep protein required for the initiation of site-specific AAV DNA integration. Furthermore, these signals can serve as a minimal origin for Rep-dependent DNA replication. In addition, we isolated the mouse Mbs85 proximal promoter and show transcriptional activity in three mouse cell lines.