Answer to Question re DNA stability

ELADER at AMBION.COM ELADER at AMBION.COM
Thu Sep 17 20:50:46 EST 1998


In article <6tlkhd$3b8 at mserv1.dl.ac.uk>,
  "RAYMOND CHAWLA (BIOCHEMISTRY) PG" <RCHAWLA at ollamh.ucd.ie> wrote:
> Hello fellow scientists,
>
> I have a very perplexing problem and would be grateful for any
> assistance/advice you can offer me.

Often when you PCR many cycles past the plateau, the Taq can not extend all
the denatured templates . This leaves you with ss DNA. I see this all the
time since my products are radiolabelled. If you run the stuff in a PAGE Urea
gel, you only get one band.

Putting a fragment in water is never a good idea. Besides not being buffered
and not having EDTA to chelate divalent cations, the lack of salt
destabilizes the double stranded DNA. It's like washing your southern blot in
water - very stringent wash indeed!

Eric


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