FIGURE 5.

Gel mobility shift assay to examine the DNA-binding activity of RFPL3. Substrate DNA (0.5 kb, 250 pm) was incubated with two different concentrations (25 nm, left panel; 100 nm, right panel) of wheat germ cell-free-produced GST-RFPL3 (lanes 2 and 6), GST-DHFR (control protein, lanes 3 and 7) and E. coli-produced BAF (lanes 4 and 8) for 1 h at 30 °C. The mixtures were separated by agarose gel electrophoresis, and then Southern blotting analysis was carried out to detect substrate DNA using an alkaline phosphatase-labeled DNA probe. Lanes 1 and 5 are reaction of substrate DNA without proteins.