Two specific DNA binding sites for the enzyme dihydrofolate reductase from Lactobacillus casei have been located by means of an immunoprecipitation assay within a 2900-base pair L. casei DNA fragment containing the L. casei dihydrofolate reductase structural gene, which was previously cloned into pBR322. The inserted L. casei DNA was mapped using restriction endonucleases, and the location and orientation of the structural gene coding for L. casei dihydrofolate reductase were determined. The two specific binding sites map at the 5' end of the structural gene, approximately 100 base pairs upstream from the start of the coding region.