What makes gels smile?
ashendel at aclcb.purdue.edu
Wed Dec 11 12:38:01 EST 1996
On 10 Dec 1996 13:59:15 GMT, Dr E. Buxbaum <EB15 at le.ac.uk> wrote:
>brett at BORCIM.WUSTL.EDU (brett) wrote:
>>You tried slow?!? Smiling comes from uneven heat distribution within the
>>gel, the heat being focused in the center of the gel. This lowers the
>>resistance in the center, and boosts the current. As the nucleic acids
>>(and dyes) migrate with this current, they move faster in the center of
>>the gel. The only solution is to redistribute the heat.
>Yes, that's the way I heard the story. Which just leaves me wondering:
>How come that my gels are always frowning? (And I run them watercooled)
You water cool DNA sequencing gels??????? With urea in them?
I was under the impression that these gels need to be run at about 60
degrees C for the DNA denaturation to be effective and the samples to
run properly. That is the whole point of prerunning the gel, to warm
it up before loading the samples. That is why they are best run under
constant power, so the heat output, and hence the gel temperature, is
constant. For the S2 BRL apparatus we have, using 0.4 mm non-wedge
spacers, I think that is 60 watts of power. More power, the plates get
too hot and crack. Less, and the correct temp is not reached and the
urea is not effective. Running with constant voltage starts out hot
and gets cooler during the run.
For those without a thermometer (the kind that use thermal color
coding or thermal-chemical opacity on thin plastic film and sold for
placing on window glass work nicely when stuck onto the front glass
plate), 60C is very warm to hot to the touch (almost too hot to keep
you hand on it).
I suspect that your gels are running too cool. Try running them
without the water cooling and under constant power.
Purdue University, West Lafayette, IN
ashendel at purdue.edu
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