We use a double-stranded DNA probe, biotinylated at each 50 end. Addition of a molar excess of an identical double-stranded DNA that is not biotinylated competes for binding with the biotinylated probe, demonstrating specificity (Irarrazabal et al., 2004). Supershift of the complex by a specific antibody is used to test whether a given protein is part ofthe complex bound to the ORE element. Controls include absence of antibody or cellular extract. The protocol is a modified LightShift chemiluminescent electrophoretic mobility-shift assay (LS EMSA) (Pierce).
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