Here are the ingredients involved in a Restriction Digest, before they are all mixed together. The only reagent not shown is a bottle of sterile, distilled water. The reagents shown are typically found in concentrated form, so the water is used to bring everything to its "working concentration." Not only that, but the glycerol in the stock enzyme tube must be diluted, because high levels of glycerol (>10%) inhibit Restriction Enzymes.

Because so many molecular biology reagents are shipped, stored, and incubated in microfuge tubes like those shown in the ice bucket here, it is extremely important to keep all of your tubes clearly labeled. In a Restriction Digest like this one, the tube on the left (DNA) would be the purified, concentrated DNA; the center tube (B) holds the Buffer; and the last tube (E) is the Restriction Enzyme.

A typical Restriction Digest is in a final volume of 10 or 20 microliters (usually denoted as µl). Buffers are usually shipped in a concentration 10 times that at which it will be used (called 10X). And the RE, which has an activity measured in Units where 1 Unit is the amount of enzyme needed to digest 1 µgram of DNA in 1 hour, is shipped at 10 Units/µl and stored temporarily at 1 Unit/µl. So if, for example, the final volume is 10 µl, the following volumes of reagents would be added to the reaction tube:

• 1 µgram DNA (typically 1-5 µl)
• 1 µl 10X RE Buffer
• 1 µl Restriction Enzyme
• Water to 10 µl

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