Does unautoclaved DEPC inhibit northern blotting?
Posted 23 August 2010 - 12:55 PM
Posted 23 August 2010 - 02:50 PM
Posted 26 August 2010 - 06:43 PM
I use DEPC treated H2O for all of the buffers from electrophoresis to immunodetection during northern blotting. If I stop autoclabing DEPC treated H2O or buffers made from it, will it have a negative impact on the blots? I know carboxymethylation of purines by DEPC inhibits translation but what about migration in the gel, RNA:RNA or DNA:RNA hybridization or immunodetection? It's not that I am lazy but we've all of the sudden have a very limited access to an autoclave.
You need to autoclave it to inactivate the trace DEPC. DEPC will react with anything that has a -N, -O, or -S, so it may screw up your RNA bases. I would autoclave to be safe, especially if you're using it to store RNA in.