Help!!!Gel box cause for RNA degradation???

Dan Baggott dbaggott at reed.edu
Sat Sep 30 16:18:32 EST 1995


I'll second the use of hydrogen peroxide.  We can get away with using 3% 
(diluted in DEPC-treated water) for approx. an hour followed by a rinse with 
DEPC-treated water.  Ambion makes a detergent called RNaseZap that also works 
well for removing RNase activity from glassware and plastic.  (They 
periodically seem to have free offers where you can get a trial size bottle.)

For what it's worth, we make up RNase-free gels in baked glassware using 
buffer that was made with DEPC-treated water and autoclaved.  Molds and gel 
boxes are treated with H2O2 or are RNaseZapped and we add 40% glycerol 
(DEPC-treated) to our samples for loading.  I usually like to treat the gel 
mold prior to each use (although I don't always and I haven't had problems 
the times I've skipped treatment) but the box/running buffer last a while 
before needing to be replaced/recleaned.  And depending on how paranoid I'm 
feeling, if RNA is getting degraded I'll just rinse out the box with 
DEPC-treated water and replace the buffer with fresh RF buffer without 
cleaning the box.  Perhaps some of these precautions are unecessary, but it 
does the trick.

Dan Baggott (who is not associated with Ambion)
Reed College, Portland, OR

mantei at neuro.biol.ethz.ch (Ned Mantei) wrote:
>In article <cad596-2909951158560001 at aragorn223.acns.nwu.edu>,
>cad596 at lulu.acns.nwu.edu wrote:
>
>> Our lab has been having problems with total RNA preps, i.e. total degradation.
>> We have narrowed the source down to the gel box (which was treated
>> overnight with 0.5N NaOH to remove RNases, followed by complete rinsing,
>> to no avail).  Any suggestions.  Buffers, loading dye, and agarose are all
>> OK.
>
>We treat the gel box, tray, and slot former with 5--10% hydrogen peroxide
>for about 30 minutes. Rinse several times with autoclaved water. Autoclave
>gel plus buffer (ethidium bromide added without autoclaving, if you use
>it).
>
>-- 
>Ned Mantei
>Neurobiology, Swiss Federal Institute of Technology
>CH-8093 Zurich, Switzerland
>mantei at neuro.biol.ethz.ch; fax: +41-1-633-1046





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