Figure 4
From: Selective inhibition of c-Myb DNA-binding by RNA polymers
Order-of-addition analysis. A mixture of 40 fmol NR1R2R3 and 20 fmol R2R3 was incubated with 30 fmol MRE-containing DNA probe and 1 ng poly(G) (lane 4–6), 10 ng poly(G) (lane 7–9) and 100 ng poly(A) (lane 10–12). In the cases marked R (for "RNA first", lane 4, 7 and 10), the proteins were incubated with homoribopolymers for 15 minutes at 25°C before addition of DNA-probe and further incubation for 15 minutes at 25°C. The cases marked S (for ``simultaneous addition'', lane 5, 8 and 11) represents reactions where homoribopolymers and DNA probe were mixed before addition of protein and 15 minutes of incubation. Finally, the cases marked D (for ``DNA first'', lane 6, 9 and 12) show reactions where the protein were incubated with DNA first and then homoribopolymers were added. The reactions in lane 1–3 contained no homoribopolymers, NR1R2R3 protein alone in lane 1 and R2R3 protein alone in lane 2. The binding reactions were subsequently analyzed by EMSA and phosphorimaging.