Wizard PCR purification from agarose gel
shsu at HEART.MED.UTH.TMC.EDU
Mon Mar 18 15:32:47 EST 1996
How big is the fragment you want to purifiy? If that is too small, close to
200 bp, you need to incubate it much longer or even put into cold room to
have better binding. I purified a 100 bp fragment by incubating in ice for 5
min, after melt the gel in 65 C and mixed with resin, and that worked
perfectly. Agarose of 1.0% gives me no problem, I don't know about 2% gel
you can dilute it to test. Final vol. or or dilution mix is not important,
about 3.4X resin with your melted gel (1.0%) will bind well. Good Luck!
>Tae Gwon Oh wrote:
>> ruth e.w. walker wrote:
>> > I am unable to purify my PCR product from the gel using the Wizard kit.
>> > It works well when I purify directly from the PCR rxn. I use 2% agarose,
>> > which melts within 4 min at 65C in presence of the resin. Is that too
>> > long? Too short? What else could be wrong, given that it does work with
>> > the PCR rxn, done concurrently?
>> Did you use TAE buffer ? It's important. After heat in 65C, then cool to
>> r.t. I can help you.
>I doubt if TAE is that important. We use TBE, have used the Wizard resin to
>purify DNA fragments and it isn't a problem. How are you doing the rest of the
>procedure. You might try adding TE to a final volume of 500 ul, then add the
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