Competitive gel mobility shift assay. (A) Schematic of the AAV-2 ITR demonstrating the positions of the Rep-binding site as well as the pertinent 30 meric oligonucleotides. The location of the consensus sequence for CCAAT/enhancer binding protein alpha (C/EBP) and human zinc finger 5 protein (ZF5) is also shown. (B) Gel mobility shift assay performed with equivalent amounts of nuclear protein derived from the liver of male and female mice and radiolabeled 30 meric single-stranded oligonucleotides encompassing the A region (oligo 7) include the rep binding site, C region (oligo 5), and the D— region (oligo D—) in the presence (+) or absence (–) of a 50-M excess of unlabeled cognate. Retarded nuclear DNA complex (C) and free radiolabeled probe (P) are indicated by arrows. (C) Gel mobility shift assay demonstrating the intensity of binding to radiolabeled oligonucleotide 7 with equivalent amounts of hepatocyte nuclear extracts derived from murine (MF indicates naive female; MF + DHT, female pretreated with DHT; MM, unmanipulated male; CMM, castrated male mouse) and human donors (HM indicates male; HF, female) in the presence (+) or absence (–) of a 50-M excess of unlabeled cognate. Retarded nuclear DNA complex (C) is indicated by the arrow.