PCR from blood

dnaboy at my-dejanews.com dnaboy at my-dejanews.com
Wed Sep 23 22:02:54 EST 1998


In article <6u6nof$of5 at pmgm.Stanford.EDU>,
  jladasky at pmgm.Stanford.EDU (John Ladasky) wrote:
> In article <Pine.OSF.3.91.980918111106.30177B-100000 at leofric>,
> S. Patel <apy019 at coventry.ac.uk> wrote:
> >
> >Dear all
> >
> >I have been extracting DNA  from clinical blood samples collected using
> >vacutec tubes. I have been experiencing some problems with inhibition in
> >PCR. Does anyone have any advise on DNA extraction from blood ie to
> >remove inhibition or to improve DNA recovery. Any comments would be most
> >helpful.
>
> 	I seem to recall that Ernest Beutler's lab discovered that
> heparin inhibits PCR.  I've been getting all of my blood samples drawn
> into tubes containing EDTA, rather than heparin, for this reason.
> Standard DNA isolation protocols give PCR-capable DNA from blood samples
> containing EDTA anticoagulant.
>
> --
> Rainforest laid low.
> "Wake up and smell the ozone,"
> Says man with chainsaw.					- John Ladasky
>

My experience is that ACD vacutainer tubes work well with PCR.	I have also
heard about Heparin problems, though I've never specifically run into them. 
You might also want to try changing your extraction method.  Gentra makes a
kit which seems to work well, called DNA Puregene.  If nothing else though,
you might just try an extra wash of the WBC pellet after lysing the RBCs. 
PBS should keep the WBCs stable while washing out any leftover Heparin.


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