Mycobacteriophage L5 is a temperate phage of the mycobacteria that forms stable lysogens in Mycobacterium smegmatis. We show here that the 183-amino-acid product of L5 gene 71 confers immunity to L5 superinfection, is required for maintenance of the lysogenic state and contains a helix-turn-helix DNA-binding motif--properties associated with repressors of temperate phages. We have utilized these observations to demonstrate the use of L5 gene 71 as a selectable marker for genetic transformation of the mycobacteria. Significantly, the use of L5 gene 71 as a selectable gene avoids the requirement for antibiotic-resistance genes providing an important tool for manipulation of the pathogens Mycobacterium tuberculosis and Mycobacterium avium, and for the construction of recombinant BCG vaccines.