Two Sequencing questions:
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)
I would claim to speak for my university but no one is likely to
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