dyes in PCR reaction
Mary Clare Tracey
seebelow at for.my.address
Wed Oct 16 22:17:29 EST 1996
> luc nelles (lne at sgi.celgen.kuleuven.ac.be) wrote:
> > I want to include some loading dye in my PCR( and other) reactions in
> > order to skip the tedious addition of the loading buffer afterwards.
> > Does anyone have experience with any dyes in PCR reactions, restriction
> > digests or other? The same question goes for any density increasing
> > medium.
For PCR-SSP (sequence-specific primers) I use Cresol Red (10mg/ml
stock, 25µl/750µl master mix) and glycerol (125µl/750µl master mix). I
then combine the master mix 1:1 with the DNA solution. I'm doing 10µl
reactions in a PE 9600 and they work fine so long as I'm really sure
to seal the lids well. The saving in time and wrist-strain of dotting
out x number of blobs of loading dye is definitely worth it. The
Cresol Red does not interfere with the reaction and runs about 100bp
(I think, but don't quote me) on agarose.
When pipetting glycerol, cut the end of the pipette tip off to
increase the hole size and pipette v e r y slowly, both up and down.
Hope this helps
Mary Clare Tracey email: maryclareatkokako.wnmeds.ac.nz
Wellington **NOTE: change the at to @ for reply. Avoiding spam!**
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