Two Sequencing questions:

Michael Holloway ca566 at cleveland.Freenet.Edu
Mon Aug 17 19:03:21 EST 1992


In a previous article, rrumpf at magnus.acs.ohio-state.edu (Robert Rumpf) says:

>I have two questions regarding sequencing protocols:
>
>1.  Our 35-S gels are still sticky after drying; the saran wrap sticks to them 
>and is difficult to remove.  Why?  We also noticed that removing the saran wrap
>and placing film directly on the gels apparently causes some damage to the 
>film from the moisture or urea in the gel...
>
>2.  Anyone have any experience with 33-P yet?  We would like to know if the 
>energy from this isotope is sufficient to pass through saran wrap so we can 
>expose films without taking the wrap off.
>
>Any help will be greatly appreciated...

Don't know how much help I can offer but I can offer some sympathy.  I've
recently tried 33P in dideoxy sequencing.  I don't "fix" the gel and I take
the saran off after vacuum drying.  I did notice that the gel is definitely
sticky but I was able to remove the wrap and the gel does not stick to the
film.  I attributed the stickyness to the urea left in the gel.  I didn't
usually observe much stickyness when I eluted the gel prior to drying while
using 35S.  I'm running wedge gels and dry them at 80 deg C for one hour. 
The stickyness is definitely worse at the thicker end.  I had thought that
I might try drying for even longer but it hasn't been a problem for me yet
and if I start drying for 2 hours I might just as well elute out the urea. 

Please let me know if you try exposing with the film on or if someone else
mailed you their experiences.
-- 
E-mail: mhollowa at ccmail.sunysb.edu (mail to freenet is forwarded)
phone:  (516)444-3612
I would claim to speak for my university but no one is likely to
believe it.



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