DNase agar is a differential medium, not a selective one. In its standard form, it contains no ingredients that inhibit or prevent the growth of any organisms. Instead, it distinguishes bacteria based on a single characteristic: whether they produce an enzyme that breaks down DNA. Any bacterium can grow on DNase agar, but only those that produce this enzyme will show a visible change in the medium.
How DNase Agar Works
The medium contains DNA as a substrate, along with a protein digest and sodium chloride to support bacterial growth. When an organism produces the enzyme deoxyribonuclease (DNase), it breaks the DNA in the surrounding agar into smaller fragments. This breakdown is invisible on its own, so an additional step is needed to reveal which colonies tested positive.
There are two common ways to read the results, depending on which version of the medium is used.
The first method uses hydrochloric acid (HCl). After bacteria have grown on the plate, a lab technician floods it with HCl. The acid causes intact DNA to clump together, turning the agar cloudy and opaque. Around colonies that produced DNase, the DNA has already been broken into small fragments that stay dissolved in acid. The result is a clear zone surrounding those colonies against a cloudy background. No clear zone means the organism is DNase-negative.
The second method skips the acid step entirely by incorporating a dye directly into the agar. DNase agar with methyl green starts out pale green because the dye binds to intact DNA. When DNase-producing bacteria break down the DNA, the dye is released and fades, creating a colorless zone around the colony. DNase agar with toluidine blue works similarly but produces a more dramatic color shift: DNase-positive colonies turn the surrounding blue agar bright pink due to a property of the dye called metachromasia. The toluidine blue version tends to be easier to read because the contrast between the pink reaction zone and the royal blue background is more obvious, especially for organisms that produce only small amounts of the enzyme. Methyl green can be harder to interpret with weak DNase producers because the pale green background doesn’t offer as much contrast.
Why It Is Differential, Not Selective
A selective medium contains agents that suppress certain organisms while allowing others to grow. A differential medium lets everything grow but provides a visual way to tell organisms apart. Standard DNase agar does not contain antibiotics, inhibitory dyes, or any other ingredient designed to block growth. Its only job is to reveal whether a colony can break down DNA.
That said, DNase agar can be made selective by adding extra ingredients. Researchers have created modified versions that incorporate antibiotics like vancomycin, trimethoprim, and amphotericin B to suppress competing bacteria and fungi. These modified plates are used in specialized clinical settings, for example to isolate specific pathogens directly from sputum samples. But the base formulation you encounter in a standard microbiology course or clinical lab is purely differential.
Common Uses in the Lab
The most frequent application of the DNase test is identifying Staphylococcus aureus. Most staphylococci that live on human skin are coagulase-negative and do not produce DNase. S. aureus, the more clinically significant species, typically does. In one study evaluating identification methods, the DNase test had a sensitivity of 75% and a specificity of 96%, meaning it rarely gave false positives. It produced only a single false-positive result in that evaluation, making it a reliable confirmatory tool during initial screening.
DNase production is also a key feature used to identify Serratia marcescens, a member of the Enterobacteriaceae family. Most of its close relatives do not produce DNase, so the test helps distinguish it from other similar-looking gram-negative bacteria.
What the Medium Contains
The standard recipe is straightforward. Per liter of distilled water, DNase agar contains 20 grams of a protein digest (pancreatic digest of casein), 2 grams of DNA, 5 grams of sodium chloride, and 12 grams of agar. The protein digest and salt support general bacterial growth, the DNA serves as the test substrate, and the agar solidifies the medium. When an indicator dye is included, it is added to this base formula before autoclaving.
Nothing in that composition targets specific organisms. The nutrients are broad enough to support a wide range of bacteria, which is exactly what you’d expect from a differential medium designed to compare how different species behave on the same plate.