Getting rid of Taq polymerase, HELP

jim hartley jhartley at access2.digex.net
Thu Sep 28 06:35:56 EST 1995


I heard a talk recently on this topic.  In an experiment set up to answer 
this question, phenol extraction worked 
very well.  Interestingly simply precipitating the PCR product with 
isopropanol and ammonium acetate worked pretty well too.  Qiagen and 
glass purification weren't so good.  Gel purification before restriction 
wasn't very good either (electroelution was used to get the fragment out 
of the gel slice), so apparently Taq sticks to PCR products. The experiment 
generated a PCR 
product that was part of a drug resistance gene, which was then cut and 
cloned into a plasmid which would theoretically regenerate the drug 
resistance.  Filling in the end by residual Taq activity would change the 
reading frame and / or make the fragment unclonable.  

On Wed, 27 Sep 1995, Petri Kursula wrote:

> I know I've seen it somewhere... how do you get rid of Taq before 
> restriction digestion of the PCR products? Should I just extract with 
> phenol and ethanol precipitate or what? People that have experience with 
> this , please help, otherwise I'll lose my mind :-)
> 
> Pete
> 
> 
> 
> 



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