A selective medium containing 25 to 50 μg per ml of tunicamycin was devised to isolate micromonosporae from soil samples, making possible simple, preferential isolation of a variety of Micromonospora. When a large amount of Gram-negative bacteria was present in a sample, alkaline treatment (0.01 N NaOH, 5-10 minutes at 15°C) was employed to reduce the numbers. Using the tunicamycin agar medium, 1, 585 strains of presumably different micromonosporae were obtained from 400 soil samples collected from various regions around the world. In average, 4 different Micromonospora strains could be isolated from one soil sample. This tunicamycin method made possible a concentrated screening method for new antibiotics from Micromonospora.