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help with gel shift assay - (May/04/2010 )

Hi all,

I am doing the gel shift assay (EMSA, protein-DNA interaction) for one of my proteins. I labeled my probe with biotin at 5'. I saw strong shift in protein + hot probe sample. However, when I tried to compete the binding with cold probe. I never see signal decreasing, instead I saw increasing binding. The binding reaction buffer I used is 15mM HEPES-KOH (pH7.5), 7.5mM KCl, 2mM DTT, 0.5mM EDTA, 6% Glycerol and 50ng/ul PolydI.dC. Anyone has the similar experience with me? Can you give me some explanation? Thank you so much! :lol:
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me 222222, same sin happened to me :angry:


Hi, I am experiencing exactly the same problem now.