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. 2008 Aug 29;283(35):23645–23655. doi: 10.1074/jbc.M801686200

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Copyright © 2008, The American Society for Biochemistry and Molecular Biology, Inc.

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FIGURE 6. — Estimation of the apparent K^DNA_d for REV1 to 24-mer/36-G-mer and 24-mer/36-N²-BPG-mer by electrophoretic mobility shift assay. A, 24-mer/36-G-mer; B, 24-mer/36-N²-BPG-mer. Reaction mixtures containing 0.5 nm of ³²P-labeled 24-mer/36-mer primer-template duplex DNA were incubated with increasing concentrations of human REV1 (2.5–320 nm) and resolved on a 4% nondenaturing polyacrylamide gel to separate the free DNA and the REV1-DNA complexes. C, the fractions of REV1-bound DNA were plotted against the concentrations of free human REV1. Data were fit to a single-site binding equation, as described under “Experimental Procedures.” The fitted values of the apparent K ^DNA_d are indicated in the figure. The following values of K_d were estimated: 24-mer/36-G-mer (▪), 180 ± 10 nm; 24-mer/36-N²-BPG-mer (▴), 59 ± 5 nm.