Summary: Genetic determinants of the temperate mycobacteriophage Ms6 required for chromosomal integration were identified. DNA sequence analysis of an -containing fragment revealed an ORF encoding a protein of 372 amino acid residues with a C-terminus similar to other conserved C-terminal regions typical of the phage integrase family. Comparison of the sequences of , and bacteria-prophage junctions and showed a 26 bp common core sequence, where recombination takes place, near the 5′ end of the integrase gene. Nucleotide sequence analysis of the chromosomal region showed that the core site overlaps the 3′ end of the tRNAgene. An integration-proficient plasmid vector was constructed and efficiently inserted at the tRNAgene of BCG and H37Ra. It was demonstrated that Ms6 and D29 integrative systems can be used in conjunction for inserting genes at multiple loci. The site-specific integration system of mycobacteriophage Ms6 is a new tool for mycobacterial genetic analysis and is poorly related to those of the L5 bacteriophage family.


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