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Restriction Endonucleases Undergo Rigorous QC Testing to Ensure Optimal,,,Performance


Reliable restriction enzymes satisfy all your research needs

Ronda Allen Kim Kuplent
Stratagene

Restriction endonucleases are used in many molecular biology protocols, making the functional purity of critical importance. Each restriction enzyme preparation must be free of contaminants that can potentially interfere with either DNA cleavage or subsequent reactions that use digested DNA (i.e., ligation or sequencing reactions). Such contaminants might include other site-specific endonucleases that produce unwanted DNA fragments, exonucleases that destroy the integrity of the cut site by cleaving nucleotides from the resultant 5 or 3 termini of the fragments, and phosphatases that inhibit ligation reactions by removing the 5 phosphate from DNA ends. To ensure that Stratagenes enzymes consistently meet the needs of the research community, we impose strict quality control procedures in analyzing every enzyme preparation. In most instances, the testing conditions are designed to be more rigorous than typical laboratory applications. In this article, we describe the assays used to establish the functional purity of restriction endonuclease preparations.

Unit Activity Determination

Prior to assaying samples for contaminants, the unit activity of each enzyme sample is accurately determined. One unit of activity is defined as the amount of enzyme required to completely digest 1 g of substrate DNA in 1 hour at the appropriate temperature under the optimal assay conditions. This information is provided on the Certificate of Analysis provided with each restriction endonuclease. Restriction enzyme activity is substrate-dependent; therefore, when digesting a new substrate, we recommend titrating the amount of enzyme added. The unit activity of a repr
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