I am planning to perform EMSA with a supposedly DNA binding protein and a 22-mer oligo DNA. But I was wondering what's the difference between using purified protein fraction versus cell extract extracts + noncompetitve DNA such as dIdC?
My protein is transiently transfected to human cells using lipofectamine 2000. I can already have my protein of interest purified from 293T human cells using anti-FLAG beads, but I can only get this much (see attached picture) protein. It's also time consuming and costly. I already tried to use the purified protein to do EMSA several times and I get little to no binding.
I'm reluctant to use the cell lysate because the expression may not be enough. Is this much protein expression enough for whole cell EMSA?
Edited by qpwoei4756, 08 February 2013 - 05:44 PM.