Northern blots

[Ed Stephenson]

We've been having trouble with Northern blots that I think we've traced to 
the RNA not staying on the filter. After transfer and baking, there's 
plenty of RNA on the filter as determined by methylene blue staining, but 
none on duplicate filters after hybridization and washing (methylene 
blue staining as per Sambrook et al). We have used Nytran and Hybond N 
nylon filters, and have the same problem whether the filters have been baked 
at 80 degrees or UV cross-linked after blotting. Has anybody had this 
problem? Does anyone know whether we should expect to still see RNA by 
methylene blue staining after hybridization? That is can anybody 
confirm being able to see RNA on the filter after hybridization? Thanks for 
your help. My news server has been very unreliable lately - e-mail would be 
best.

Ed Stephenson
Dept of Biological Sciences
Univ of Alabama
Tuscaloosa AL
estephen at biology.as.ua.edu