Abstract We report identification of a novel site-specific DNA recombination system that functions in both in vivo and in vitro, derived from lysogenic Staphylococcus aureus phage ϕMR11. In silico analysis of the ϕMR11 genome indicated orf1 as a putative integrase gene. Phage and bacterial attachment sites ( attP and attB, respectively) and attachment junctions were determined and their nucleotide sequences decoded. Sequences of attP and attB were mostly different to each other except for a two bp common core that was the crossover point. We found several inverted repeats adjacent to the core sequence of attP as potential protein binding sites. The precise and efficient integration properties of ϕMR11 integrase were shown on attP and attB in Escherichia coli and the minimum size of attP was found to be 34 bp. In in vitro assays using crude or purified integrase, only buffer and substrate DNAs were required for the recombination reaction, indicating that other bacterially encoded factors are not essential for activity.