hekp with sigmacoat and sequencing gels
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Youre-mailaddress
Wed Jul 12 15:37:49 EST 1995
In article <Pine.SOL.3.91.950711134256.15100A-100000 at chip>,
szjmahr at peseta.ucdavis.edu says...
>
>I am relatively new to sequencing. at first when pouring my gels i
would
>clean them with detergent and a sponge, lay them flat on a tt rack and
>with no tape, pour the gel - this worked great until once when taking
the
>apparatus apart the gel stuck to both plates and made a mess of it. to
>try to prevent this i tried using sigmacoat on one plate and disaster
>struck (huge pockets where the acrylamide just WOULD NOT go - the
>location of which where not the same when i tried this 3 times in a
>row). for my fourth attempt icleaned the plates using every dish
washing
>solvent in the lab and did not use sigma coat - the same thing to a
>somewhat lesser degree happened again (i even tried taping halfway
>through and then bringing the plates vertical but the huge bubbles
>wouldn't budge despite my pouding on the plates) i have heard mixed
>reveiws on sigma coat
> 1. will i ever be able to get the stuff off my plates
> 2. does anyone know of a way to use the stuff so that it works
reliably
> 3 does any one have alternative ideas for helping me out????
>
>
>
>sincerly - jamahr at ucdavis.edu
>
I use SigmaCote straight from the bottle, using a disposable pipettor,
(perhaps a total of 6 drops/plate) and distributing evenly with a
Kimwipe. I let the plates air dry. I routinely coat both plates, then
hold the sandwich at about a 30 degree angle and pipet the acrylamide
solution in with a disposable transfer pipet into the top corner, letting
capillary action and gravity fill the plates. Sometimes it takes awhile
to fill, sometimes it fills very rapidly. A BubbleBuster (AT Biochem) is
the best way to get rid of bubbles--and to use to start the filling if
the SigmaCote is really working. I use a 2% NaOH solution to get rid of
the SigmaCote--although normally I just wash and reuse my plates. The
SigmaCote usually lasts 3 plus times before I have sticking problems
again. I prefer SigmaCote to Gel Slick (AT Biochem) because I prefer to
have a slow-pouring gel than one that sticks, tears, and ripples when
transferring to filter paper. Good luck.
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