HomeFAQsI'd like to clone a fragment amplified with Q5® High-Fidelity DNA Polymerase. Do I have to blunt-end clone?
FAQ: I'd like to clone a fragment amplified with Q5® High-Fidelity DNA Polymerase. Do I have to blunt-end clone?
Blunt-end cloning is recommended. However, if TA cloning is required, 3´A-overhangs can be added with a different polymerase. It is very important to remove all the Q5 High-Fidelity DNA Polymerase first by purifying the PCR product. The proofreading activity of Q5 is very strong, so any residual polymerase will degrade the A-overhangs as they are added. Taq DNA Polymerase or Klenow (exo-) DNA Polymerase are excellent options for A-overhang addition. We recommend ligating immediately so the 3´A-overhangs will not be lost during storage.
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