Native gel or Agarose for RNA EMSA?
 

Hi there,

I am going to do a biotinylated RNA EMSA. I would like to know what is the difference and how does it matter if I go for a native gel or an agarose gel.

Also, do you guys make individual components of a buffer (EDTA, DTT, APS) in DEPC treated water?

Re: Native gel or Agarose for RNA EMSA?
 

What percentage gel shall I make? My RNA is 2.4kb and protein is 68kD

Re: Native gel or Agarose for RNA EMSA?
 

For EMSA you need to run either a native polyacrylamide or a native agarose gel. People often use 0.5x TBE as a buffer.

It's probably not absolutely necessary to use DEPC water, Milli-Q should be sufficiently pure (assuming your glassware isn't contaminated w/ RNaseA).

I think you will have trouble detecting a mobility shift for a 2.4 kb RNA on a polyacrylamide gel, so you may need to go for an agarose gel. The alternative is to use a fragment of the RNA that you think contains the binding site.