A physical map of the chromosome of Lactococcus lactis subsp. lactis DL11 was constructed by using the contour-clamped homogeneous electric field mode of pulsed-field gel electrophoresis in one- and two-dimensional separations to analyze restriction digests of high-molecular-weight genomic DNA. The map, which shows all the observed NotI and SmaI sites (six and 21, respectively) and 8 of approximately 30 SalI sites, is circular and yields a total size of 2.58 megabase pairs for the L. lactis subsp. lactis DL11 chromosome. By using rDNA from Mycoplasma capricolum to probe Southern blots of pulsed-and fixed-field digestion patterns, six putative rRNA operons were identified in L. lactis subsp. lactis DL11 and placed on the map of the chromosome. Five of these loci are clustered in a region representing only 20% of the chromosome. The presence of a SmaI site in each of the putative operons allowed the direction of transcription of each operon to be deduced.