I am doing EMSAs, and I am seeing a shift in a control lane where the cell have not been stimulated (Ie should not expect to see the wanted protiens in the nuclear extract.) the protein also should not be in a high concentration in the cytoplasm (NFKB)
The negative control (no nuclear extract added to the binding reaction) works fine (no shift-->no protien bound)
does this mean that I am having nonspecific protiens binding to my DNA?
Also, this control lane (no stimulation to cells) has a shift with the same size as the experimental lanes... so I don't even know if the experimental lanes are true based on this.
do you think its a problem with the binding reaction, or Nuclear extraction methods?