Northern blot: Wetting a membrane - oops... (Jun/18/2009 )

Hello everyone,

Yesterday I ran an PAGE gel to detect miRNA. All was going well, then I set-up the apparatus for the transfer. I did it as any capillary protocol says: dish witn 20x SSC, glass over support, wet paper over glass, gel over paper surrounded with parafilm, membrane over gel, wet paper over membrane, paper towels and a weight. However, prior to using the membrane I did not wet it (this is where the oops comes in). How important is it to wet the membrane? Would I get any transfer if I failed to do this?

As I type this I am preparing a second membrane, hopefully it will be OK.

I used a Millipore Immobilon Ny+, in case you were wondering.

The purpose of wetting the membrane is to minimizing trapping bubbles in the membrane. It may be OK. After transfer, you can look at your gel (if you have stained with EB) to see if RNA has been transferred.