Low stringency Southern trouble

KELLY THOME 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

Kelly Thome			kmorris1 at opal.tufts.edu
Department of Pathology
Brigham and Women's Hospital
Boston, MA 
	All thoughts expressed here are my own!



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