FW: Tris acetate PAGE gels

Deanne Bell dbell at fresno.ars.usda.gov
Mon Aug 2 15:32:52 EST 2004


I can give you a protocol for a 10% Polyacrylamide Gel that I have used
for DNA Visualization.  I think it would work or at least it is a
starting point.

Prepare 20X Stock TAE Buffer
500 ml water
48.44 gm TRIS	
7.44 gm EDTA
11.42 ml Glacial Acetic Acid


Prepare 15 ml of a 10% Polyacrylamide Gel (enough for 2 Bio-Rad mini
gels)
765ul	20X TAE
75ul	10% ammonium persulfate catalyst
De-gas solution for 5-10 minutes on aspirator
12ul	TEMED

I was using a Bio Rad mini protean II electrophoresis rig
Use a 5ml syringe to pour the gel to within 5mm of the top of the glass
plate
Set under lamp 40 - 60 minutes
If the gel will set overnight, overlay the top with a layer of mineral
oil
Prepare 1 Liter of 1X TAE from 50ml of 20X Stock TAE
Place ~500 ml 1X TAE Buffer and a small stir bar into the lower
electrophoresis chamber
Place the electrophoresis chamber into a tray of ice on top of a stir
plate
Add ~ 250ml 1X TAE Buffer to the upper chamber
Electrophorese at 100V / 98mAmps 


Hope this helps to get you started
You may have better luck if you look up TAE Buffer
Deanne Bell


-----Original Message-----
From: owner-methods at hgmp.mrc.ac.uk [mailto:owner-methods at hgmp.mrc.ac.uk]
On Behalf Of "Jayakumar, R"
Sent: Friday, July 30, 2004 2:35 PM
To: methods at hgmp.mrc.ac.uk
Subject: Tris acetate PAGE gels

HI..
    does anybody know how to make Tris-acetate PAGE gels for the
separation of large molecular weight proteins?  I could not get any
information from any website or books for that matter.  if anybody knows
anything about the procedure or any pointers to any useful website,
would be greatly appreciated.
   thanks in advance
Jai


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