Using Glycogen to prec DNA???

[Simon Dawson]

> I am trying to purify a PCR fragments, around 600 bp from gel.  I can get 
> a decent amount out of the gel, but I lose it during the precipitation 
> step.  I want to use this product for direct cycle sequencing, so yeast 
> tRNA as an aid to precipitating the DNA was suggested to me to be a bad idea.
> I have heard of glycogen used to prec oligonucleotides.  Does anyone know 
> of the right concentration of glycogen to use.  And does it interfer with 
> sequencing afterwards?  Or does anyone know of another method to get a 
> better yield.  
> Thanks.  Seanna    (sannis at uoguelph.ca)
> 

Hiya Seanna,
           I use glycogen on a regular basis for precipitating small
fragments of DNA for all sorts of protocols. We use the RNase/DNase
free glycogen available from Boehringer-Mannheim. I think it used
to come at a conc. of 20mg/ml. I use it at a final concentration of 
approx. 20 microgrammes/ml I think. Something like that. I precipitate
as normal after adding glycogen i.e with NaOAc/EtOH.
  As far as it interfering with anything...I'm not sure about 
sequencing but it is certainly inert in ligations, cDNA reactions in
fact I don't think we've come across a problem with it re: messing
up enzymic reactions.
  Hope that helps! :)

                         Simon.

Simon Dawson
Dept biochemistry
Queens Medical Centre
Nottingham
UK

Internet email: mbxspd at unicorn.nott.ac.uk