Low stringency Southern trouble
kmorris1 at opal.tufts.edu
Wed Feb 14 15:08:38 EST 1996
I'm hoping someone can help me with remedial Southern analysis technique :-).
I'mtrying, without much success, to do low stringency Southerns (ie, hunting for
family members in one species/looking for homologs in other species). My hyb
and washing conditions for high stringency blotting are pretty standard, I
guess... hyb is 6X SSC/10X Dextran Sulfate/ 0.5% SDS/ 50micrograms/ml ssDNA at
65 degrees, washing is 1X SSC/0.5% SDS at 65 degrees. I tried lowering the
SDS to 0.1% and hybing at 42, but then I got NO signal, not even the signal I
see at high stringency. I'm getting sort of frustrated at blank blots and
would appreciate advice from someone with experience in this sort of thing.
Thanks in advance!
Kelly Thome kmorris1 at opal.tufts.edu
Department of Pathology
Brigham and Women's Hospital
All thoughts expressed here are my own!
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