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.
Help us celebrate our 50th anniversary! We have hidden 1,000 golden butterflies and are waiting for you to find them. They can be anywhere that you find NEB! Beginning April 15th, be sure to visit our website and tables at tradeshows and events you are attending. Visit our social media channels frequently for tips on where we have hidden the butterflies – and once you find one, either click or scan the code to be eligible for a 50th anniversary prize pack, as well as a grand prize trip to NEB headquarters in Ipswich, MA!.
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