HomeFAQsHow should I determine an appropriate annealing temperature for my reaction?
FAQ: How should I determine an appropriate annealing temperature for my reaction?
Please use NEB’s Tm Calculator to determine the appropriate annealing temperature for your primer pair and polymerase/buffer of interest. Unlike other calculators, the NEB Tm Calculator takes buffer components that affect melting temperatures into consideration when calculating the best annealing temperature. Other online calculators may underestimate the best Q5 annealing temperature.