Lysogenic bacteriophage M1 from Selenomonas ruminantium: isolation, characterization and DNA sequence analysis of the integration site

Department of Animal Science, University of Adelaide, Waite Campus, Glen Osmond 5064, Australia ABSTRACT Bacteriophage M1 from the ruminal bacterium Selenomonas ruminantium strain ML12 comprises a 30 nm icosahedral capsid, a 25 nm tail and 48 kb of linear dsDNA with cohesive ends. A restriction map of the phage genome has been constructed. The presence of bacteriophage M1 in the rumen has been demonstrated by PCR amplification and Southern blot analysis of DNA from rumen bacterial samples obtained from ten different sheep. Lysogeny was demonstrated by hybridization of M1 DNA to host chromosomal DNA and by identification and cloning of a 2.3 kb region of the phage containing the predicted attP domain which promotes chromosomal integration. DNA sequencing of the attP region demonstrated two major ORFs surrounding the predicted attP site and structural analysis of this region revealed a motif comprising three different inverted repeats surrounding a 12 bp palindrome. Analysis of the translated amino acid sequence upstream of the attP site demonstrated the presence of conserved residues found within integrase proteins of several temperate phages of different bacterial species. * Author for correspondence: John D. Brooker. Tel: +61 8 8303 7357. Fax: +61 8 8303 7114. e-mail: jbrooker@waite.adelaide.edu.au Keywords: bacteriophage, Selenomonas, integration