Cleaning seq. plates with "Bon Ami"??
bl275 at cleveland.Freenet.Edu
Wed Mar 30 00:41:18 EST 1994
In a previous article, balt at spieden.Stanford.EDU (Steven Balt) says:
>Hello, all! Our sequencing gels have been pouring rather poorly
>lately (no pun intended :) ) and we think we have narrowed down the
>source of the problem to our new glass plates.
>The glass plates have been scrubbed with "Bon Ami" brand cleanser
>instead of the normal ICN "7X" cleaning solution or Windex. Has anyone
>else experienced any weird phenomena after cleaning with Bon Ami??
>(In our case, the acrylamide solution accumulates in the _center_ of the
>glass plates instead of sliding down the sides, like it should.)
here is the procedure we use in our lab. we use the BRL S2 apparatus with
BRL plates and the IBI sharkstooth combs and spacers, since we dont like
the IBI apparatus or the BRL combs and spacers.
- soak brand new plates in 2M NaOH for 1-2 h and rinse thoroughly with
deionized water. place a sliver of tape on one side of each plate to
ensure consistent use of the same side of each plate for inside/outside.
- clean both plates with glass plus followed by isopropanol
- coat the small plate with Rain-X, a water repellent for windshields
available at many auto parts stores. none of that yucky disgusting silane
chloroform shit. put two coats of Rain-X on the small plate, let it
dry, then wash again with glass plus and isopropanol
- assemble the plate-spacer sandwich and clamp (tape is for sissies). we
use three small clips on either side. the plate sandwich is laid
horizontally on an empty pipet tip box or similar rectangular flat surface
a few inches from the benchtop
- prepare your gel mix and pour the gel horizontally, allowing the acryl
mix to enter by capillary action. we put the mix in a plastic cylinder
with lip, start on one corner of the gel, move to the other corner and then
back to the middle, allowing the gel mix to move as a single front down the
gel - its quick, easy and never fails. put some lab matting down to soak
up possible spills.
- put the comb in and clamp (we use two large clips placed so that they hold
the comb in place, but dont let the clips extend beyond the bottom of the
comb, i.e. clamp only where the clip is over comb or spacer, and never
where there is only glass)
- read a journal (a cool one) for 30-40 min and prepare your running buffer
(or just prepare several litres of Tris-Taurine (glycerol-tolerant) buffer
at once and store at RT)
- use a thin spatula to separate the comb from the gel plate, then ease
gently up. clean the surface of the gel plate which will contact the
aluminum plate to ensure even heat distribution. make sure its dry.
- set the gel in the apparatus and clamp tightly enough to avoid leaks, but
not to maximum tightness. pour in running buffer and wash out the top of
the gel with a pasteur pipet. start your prerun (we do 15-30 min at 40-60
W when using Tris-Taurine)
- get your sequenase reactions ready and wash yucky acrylamide from the
comb with water. rinse the top of the gel and insert the comb slowly until
the teeth just barely kiss the surface of the gel. tighten the gel until
- rinse the wells once again and load. run the gel.
- after the run, the plate treated with RainX will come off with minimal
effort. we then rinse with water and set the plate to dry for next time.
the larger plate goes into the fixing bath with the gel. once done, it
also gets rinsed.
- for the next run, simply clean with glass plus and isopropanol and youre
ready to go. the RainX treatment will last for 20 gels or more. fun fun.
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