Help Northern Blocking!

Mark Watson watson at dogwood.botany.uga.edu
Fri Jun 14 08:44:26 EST 1996


Hello Net People,
I was wondering if any of you folks have had or seen this problem.  It has 
arisen suddenly and I can't seem to figure out what is going on.  I have had 
horrible non-specific background on my last couple of northerns.  It seems the 
probe is binding to the filter and not the RNA.  I remove the unicorporated 
nucleotides from my random primer labelled DNA probe.  I blocked overnight 
with 5X SSC + 5X Denhardts + 1% SDS and 100 ug/ml sheered/denatured Herring 
Sperm DNA + 50% formamide (biorad Mol. Biol. Grade) at 42oC.  I have been 
using one of those rotary hyb ovens.  The nonspecific background can be as bad 
as to show the RNA as a negative image.  Please help before I go crazy!
Thanks
Mark

ps  I have been blotting to biorad zeta probe nylon with 10X ssc


*******************************
 Mark Watson
 watson at dogwood.botany.uga.edu
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