I have gel-purified PCR product from a Q5 high-fidelity amplification to which I want to add the A tails for TOPO-TA cloning. The protocols I find all say to use Taq polymerase and dATP, but can I use a Taq polymerase mastermix instead. I know it has all four nucleotides, but shouldn’t it add the A tail also? (I have Colorless GoTaq Hotstart mastermix 2X on hand.)
Uwe Borgmeyer
Adding all nucleotides is no problem. See :
- Zhou, M.Y., Clark, S.E. and Gomez-Sanchez, C.E. 1995. Universal cloning method by TA strategy. BioTechniques. 19:34-35.
or:
https://bitesizebio.com/19709/ta-da-the-magic-of-taq-and-ta-cloning/
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Shane Denecke
Hello David,
A master mix shouldn't be a problem. I've used GoTaq master mix for A-tailing and it always worked well for me.
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