Northern Blot - RNase-free

Simone Marker via methods%40net.bio.net (by marker from rhrk.uni-kl.de)
Tue Jul 31 06:28:16 EST 2007


Hi,

In general, when I prepare a denaturing agarose gel (with formaldehyde) for 
separation of RNA do I have to make the solutions and the apparatus 
RNase-free?
Is it the same for denaturing polyacrylamid gels with urea as denaturing 
reagent? (I want to detect small RNA (siRNA) by northern)

And when I continue with blotting for a northern, which steps need to be 
RNase-free? Is it really necessary to prepare RNase-free Whatman-paper, 
membrane, vacuum blotter and so on? What is the easiest way to do this? I 
heard of wiping the equipment with H2O2, but I'm not sure about the 
concentration. And how can I get Whatman paper RNase-free?
Or do you think that working quickly might suffice to reduce degradation by 
RNases?

Thank you for your advice,
Simone 




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