DNA quantitation method wanted

Jeff crowther at rschp1.anu.edu.au
Wed Mar 9 01:41:31 EST 1994


In article <3253.2d73f345 at mbcl.rutgers.edu>, marian at mbcl.rutgers.edu wrote:
..........
>     I am looking for a quick and effiecient method for quantitation
> of DNA.  I want to use the method to control for cell number in
> cell cultures.  I need most of the cell layer, so I only have a
> small percentage of the sample availabe for quantitation.  I am
> aware of Invitrogen's "sticks", but these are not very cost-efficient.
> I am condsidering Maniatis' protocol for etbr quantitation of
> droplets on saran wrap, but have serious reservations about the 
> relaibility of this method.  Any suggestions?

I have used the Maniatis method for quantitation of DNA in protein preps.
Rather than using saran wrap I use a 1% agarose mould of 10x5 4mm wells.
The DNA/EtBr in appropriate amounts are mixed in small ed's and then put
into
wells. Run a set of std's from known DNA conc on the gel block.
Advantages-
DNA drops are of uniform size due to wells as are std's
No/low background from gel
Low diffusion at 1% over a moderate length of time
I find quantitation is not too bad from a reasonable number of stds but
depends on the level of precision you want
If you want to get carried away Scan gel/photo and do densitometry on it.I
have not done this so don't know how accurate it is.

Hope it helps. If you have any further queries e-mail me
crowther at rsc3.anu.edu.au

Regards
Jeff C.
Centre for Molecular Structure and Function
Research School of Chemistry
Australian National University

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