Fixing sequencing gels

Kilroy, Gail - Dr. York 2545 kilroyge%pbrc at mhs.pbrc.edu
Tue Aug 2 15:41:45 EST 1994


>010Date: 29-Jul-94 11:27:33 -0600
>From: HORTON @ XGATE (Robert Horton) {horton at molbio.cbs.umn.edu}
>To: METHODS- @ XGATE {methods-and-reagents at net.bio.net}
>001Subject: Re: Fixing sequencing gels
>
>  To: methods-and-reagents at net.bio.net
>  From: horton at molbio.cbs.umn.edu (Robert Horton)
>
>Chris Greene (cgreene at bimcore.emory.edu) wrote:
>: In article 1125646336B at ra.uvic.ca, cupton at sol.uvic.ca ( Chris Upton) writes:
>: > In response to an article in Biotechniques (FEB '94, Yang & Waldman), that
>: > suggested it wasn't necessary to fix 35S seq gels but that the Saran Wrap
>: > should be left on - I persuaded a grad student next door to try this :-)
>: > 
>: > He found it worked fine!! (to think of all the time I spent fixing gels!!)
>: > 
>: > BUT, he also took off the Saran Wrap!!  & there was _NO_ problem with the
>: > gel sticking to the film!!
>: > 
>: > This may be because he dried the gels very well or low local humidity.
>: > I guess it's something to test yourself.
>: > 
>  <snip> 
>
>: I have been exposing my sequencing films in this way for about two
>: years now, and it does indeed work fine.  One thing you must be careful
>: of is that the gel must be dry if you remove the plastic film.  I'm
>: sure that when the humidity levels rise once again you will finds lots
>: of little black dots all over your film where the gel stuck.  This
>: tends to make the film rather unsightly but can still be read quite
>: easily.
>
>  <snip>
>
>I do it this way, and rarely have problems - you need to be quite sure the
>gel is dry, though. This is in Minnesota, where the humidity is not usually
>severe. Our gels are lifted onto 3MM paper, and dried at 80oC for about 60
>-90 minutes, using a water aspirator. On a humid day, it may be important
>to expose the gel to film right away, so it doesn't sit around rehydrating.
>But I think the vast majority of the urea is sucked into or through the paper
>anyway, so rehydration is not as much of a problem as one might think.
>It would be interesting to hear from people in REALLY humid places about
>how this works (anybody NOT fix their gels in Baltimore?)
>
>The idea that different techniques work in different places is intriguing;
>I knew a guy who was a sequencing wizard in Minnesota, but he went off to do
>a postdoc in France, and (after much frustration) he had to adopt a different
>set of procedures, particularly with regard to preparing the plates.
>Something in the water, he thought.
>
>Anyway, ... if it ain't broke, don't fix it. (Somebody had to say that B^)  ).
>
>
>Bob Horton (Ph.D.!)   /\ "Crash programs fail because of the theory that
>U. of Minnesota, CBS  || with nine women pregnant you get a baby a month" 
>1479 Gortner Ave.    /||\   -Werner von Braun.  Disclaimer:"Bob who?"
>St. Paul, MN 55108    ^^   horton at molbio.cbs.umn.edu/(612) 624-3790
>

I've been drying my sequencing gels without fixing for several years and
even in Louisiana, where the humidity lingers around 95%, there isn't a
problem.  What I've done, when leaving the saran wrap on would present a
problem, is spray a thin coat of silicone over the gel. You can pick up an
aresol can of silicone at your local hardware store.  The silicone does not
seem to block the signal.



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