Technique / Molecular Biology / PCR / Standard PCR


Taq inactivation



Taq inactivation

I am going to directly use the PCR mixture for restriction digestion,
but I fear the remaining Taq in the PCR mixture will fill in the 5' overhang
generated by restriction endonucleases. Is my worry necessary? Can Taq be
inactivated by heating at 94-96C for 30 min prior to restriction digestion? ... ...

Last update 05-Mar-2007, Rating of 1 votes.

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By Cathal on 05-Mar-2007
(For starters, this seems to be the wrong place to post questions: see the forum for that.)

I believe the different buffer used with restriction enzymes would be incompatible with taq. It pays to add the restriction enzyme buffer before the enzymes in this case, to be certain that taq isn't still active. However, conditions for restriction enzymes vary, and some might be compatible with taq.

If you're afraid that your reaction conditions might favour or allow taq to interfere with digestion, it's probably best to purify the DNA.
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