Background problem with a northern blot
William Driscoll
driscoll at rci.rutgers.edu
Thu Aug 14 16:42:50 EST 1997
I am having a background problem with my northern blots. I am getting an
large amount of non-specific binding. I am using a biotin labled 40mer
DNA probe. My membrane is a nylon positively charged membrane(Boehringer
mannheim). We have narrowed the problem down to the hybridization
blocking step. The hybridization conditions are the following: \
Temp: 65 degrees C
Solution:7% SDS, 37g/L EDTA, 35.5g/L Sodium phosphate dibasic
I would appreciate and help or advice about what to do to reduce this
non-specificity problem. I was wondering what others have added to
the reduce the background when using a biotin labled probe. I was thinking
of adding herring sperm DNA but I don't know where to obtain this. (I've
tried pharmacia, sigma, Fisher, and BioRad.) I also was thinking of
adding Denhardt's Solution.
I don't get to check news as often as I'd like so please email me (driscoll at rciu.rutgers.edu) instead of just posting (I might miss the post) ... Thank you
Thanks in advance
Bill
driscoll at rci.rutgers.edu
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