The beta-galactosidase of Lactobacillus sake DSM 20017 is encoded by two genes located on its chromosome. These genes designated lacL and lacM were cloned in Escherichia coli NM 554 on an 8.65 kbp HindIII fragment inserted in vector pRB473. Deletion analysis of the originally cloned fragment revealed that both genes are required for the formation of a functional beta-galactosidase. lacL and lacM are transcribed as a single transcript of approximately 2.9 kbp starting 34 bp upstream of the translational start codon. The proteins derived from lacL and lacM share only 18-59% homology with other beta-galactosidases. The genes encoding the beta-galactosidase are scattered with multiple direct and inverted repeats of 9-12 bp. However, comparison with the plasmid-encoded Leuconostoc lactis beta-galactosidase revealed equal distribution of conserved amino acid residues and suggests that the genes have a common origin. Specific deletions or insertions resulting from the presence of the repeats were not observed. The L. sake beta-galactosidase was phenotypically expressed in E. coli NM 554 and Lactobacillus curvatus LTH 1432. Its two genes can be used to replace antibiotic reporter genes to develop food-grade vectors and alpha-complementation systems for self-cloning in meat lactobacilli.