Important Tips for Q5® Polymerase

Here are some quick tips for getting the most out of NEB's Q5® High-Fidelity DNA Polymerase.


Here are some important tips for working with Q5 high-fidelity DNA polymerase. Q5 often requires higher annealing temperatures than other DNA polymerases.

First, determine the optimal annealing temperature for your primer set. We always recommend starting with the NEBTm calculator, found on the NEB website.

Second, Q5 is optimized for speed, so extension times of 20 seconds per kb are recommended. It is important to avoid extension times that are greater than one minute per kb.

Third, a denaturation temperature of 98 degrees is recommended with Q5 high-fidelity DNA polymerase.

Fourth, we recommend using a final concentration of 0.5 micromolar for each primer in your reaction.

Lastly, only one unit of Q5 high-fidelity DNA polymerase is recommended per 50 microliter reaction. Following these tips will ensure successful amplifications with Q5 DNA polymerase.

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