Does purifying PCR probes for EMSA from EtBr gel interfere with binding? - (Nov/18/2009 )
I have a non-radioactive labeled (DIG) PCR product as a probe for EMSA. The PCR solution purified using qiagen spin columns have 2 bands in the probe only control in EMSA, a strong dark band at the bottom, and a much fainter band on top. I am thinking its single stranded. When i perform the PCR without the labeled DIG-dUTP, i only get 1 band of the right size. But the labeling somehow introduces another band twice the size.
So, im thinking of purifying the labeled probe from a gel to isolate the right band.. i'm worried the EtBr in the gel may interfere with DNA binding, is that a valid concern?
I think you will probably be OK, but you might do a small test-run with just a control to check before you use up your extract.