crackling sequencing gels

A.F. Simpson AFS7 at le.ac.uk
Fri May 26 18:07:27 EST 2000


T. Max M. Soegaard wrote:
> 
> thin 0.4mm sequencing
> gels in a size of app 50cmx30 cm. Do you think it would hurt to
> transfer them to filter paper first and then fix and wash them.? Im
> thinking that it will be some very delicate handling steps with a big,
> thin, gel..(but I dont know)

I didn't realise they weren't being fixed!  That'll teach me to make
assumptions.  Yes, they _definately_ need to be fixed.

As for handling the gel:

The filter papaer would fall to pieces in the fix, and if it didn't,
you'd have a piece of filter paper soaked in fix to dry out as well as
the gel.

The way I used to handle fixing sequencing gels was to soak for 1 hour
in 10% acetic adic/10% methanol (as per the Sequenase 2 kit booklet). 
However, this was for 1.2mm-0.4mm gels, so a shorter time for a 0.4mm
gel should be sufficient.  I find pouring the fix into the tray and then
lowering the gel in SLOWLY is much better than putting the gel in the
tank then pouring in the fix.

To transfer to the paper I lift the gel out of the tank rather than
pouring off the fix.  Again, SLOWLY is the key; if the gel starts to
slide it can usually be rescued by tilting the plate back down.  Then
transfer to a double thickness of filter paper.  The sheet in contact
with the gel gets quit wet and therefore fragile, so a double thicknes
makes it easier to handle.

(BTW, you _are_ posting in HTML.  Luckily my browser can read it and
quote back properly, but some people's won't and they probably won't
bother reading your posts.  I don't know anything about Mozilla, but
there muct be an option somewhere to turn the HTML off.)

> -Max






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