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. 2000 May;11(5):1673–1685. doi: 10.1091/mbc.11.5.1673

Figure 7.

Figure 7

Figure 7

Mutagenesis analysis of Cdc6 DNA-binding and Abf1-stimulation activities. (A) EMSA was performed to examine the DNA-binding activity of wild-type and mutated GST fusions. Ten, 20, 30, and 40 ng of GST-AG6 (lanes 1–4), GST-P21S (lanes 5–8), GST-K46A (lanes 9–12), and GST-NP6 (lanes 12–16) were used. In lane 17, 100 ng of GST was used as a negative control. (B) The stimulation of Abf1 DNA-binding activities was described previously (Feng et al., 1998). EMSA was performed as in the DNA-binding assay, except that 10 ng of purified Abf1 protein was added in lanes 1–16. Lane 17 shows a no-protein control, and 10, 20, and 50 ng of Abf1 protein alone (lanes 18–20) were used as positive controls. The 283-bp ARS1 was used as the probe. Various amount of GST fusions (0.5, 1, 2, and 3 ng) were added in the reaction mixture: GST-NP6, lanes 1–4; GST-cdc6(K46A), lanes 5–8; GST-cdc6(P21S), lanes 9–12; and GST-AG6, lanes 13–16.