Intact cells of Agrobacterium tumefaciens were examined for ability to take up biologically active LR-4 phage deoxyribonucleic acid (DNA) from the surrounding medium. DNA incorporation as measured by subsequent plaque formation (transfection) failed to occur when the bacteria were grown in defined minimal salts media, and was restricted to a 4-hr period in the early log phase of growth in enriched media. In the latter case, maximal transfection frequencies were obtained after a 25- to 30-min incubation with 22.5 mug of phage DNA/ml. Higher DNA concentrations or longer incubation times were inhibitory. Transfection was completely inhibited by deoxyribonuclease but not by ribonuclease, trypsin, or phage-specific antisera.