DNase I (RNase-Free)

What it means: Purity Grade describes what the preparation is certified free of. RNase-free certified means the enzyme has been tested to contain no detectable ribonuclease activity.

Why it matters:

• The whole point of a DNase step in an RNA workflow is to destroy DNA and leave the RNA intact — a trace of RNase defeats it.
• RNase contamination is silent: the sample simply gives poor yields or low RIN downstream, with nothing pointing back to the DNase.
• Standard-grade DNase preparations frequently carry residual RNase and are unsuitable before reverse transcription or library preparation.

How to interpret here: This grade is what makes the enzyme safe to use on RNA — for gDNA removal before RT-PCR or RNA-seq. A non-RNase-free DNase should never be substituted in those steps.

Tip: Keep a dedicated aliquot for RNA work and never return a used tip to the stock tube.

What it means: Inactivation describes how the enzyme is stopped once digestion is complete. EDTA-chelatable means the enzyme depends on divalent cations, so chelating them shuts it down.

Why it matters:

• Carrying active DNase into a reverse transcription or PCR would degrade the cDNA and template you are trying to make.
• Chelation works at room temperature, so RNA is not exposed to the heat that a thermal inactivation step would require.
• Heat inactivation in the presence of divalent cations can itself fragment RNA, which is why the EDTA route is preferred here.

How to interpret here: Add 1 µl of 1 M EDTA (not supplied) and hold at room temperature for at least 10 minutes, or remove the enzyme by column purification with the Column-Pure RNA Miniprep Kit (Cat. No. D518).

Tip: Remember the added EDTA carries into the next reaction and chelates its Mg²⁺ too — account for it when setting up the RT or PCR.

What it means: Buffer Included describes the reaction buffer supplied with the enzyme. This product ships with 1.0 ml of 10X DNase I Reaction Buffer alongside 200 U of enzyme.

Why it matters:

• DNase I requires divalent cations, and the supplied buffer provides them at the concentration the enzyme was characterised in.
• Using an EDTA-containing buffer from another kit will chelate those cations and silently prevent digestion.
• A matched buffer removes composition as a variable when a gDNA removal step appears to have failed.

How to interpret here: Dilute the 10X buffer ten-fold in the digestion reaction. Do not substitute a generic nuclease buffer, and check that your RNA elution buffer is not already EDTA-based.

Tip: Thaw and mix the buffer completely before pipetting; salts settle during frozen storage.

What it means: Concentration is how much enzyme activity is packed per microlitre. At 2 U/µl with 200 U supplied, one unit degrades 1 µg of pBR322 DNA completely within 10 minutes at 37°C.

Why it matters:

• Knowing units per microlitre lets you dose by activity rather than by volume, which is what keeps digestion reproducible between lots.
• Over-dosing DNase wastes enzyme and adds more protein and glycerol to a reaction that will be carried forward.
• Under-dosing leaves residual genomic DNA, which shows up as false signal in a downstream qPCR.

How to interpret here: Scale the enzyme to the DNA load, not the sample volume. A typical RNA prep needs well under 1 U, so the 200 U pack covers a large number of gDNA removal reactions.

Tip: Always run a no-RT control after DNase treatment — it is the only way to confirm the genomic DNA is actually gone.