Figure 3. DNA binding activity of RUNX2 or RUNX2^R131G.

HA tagged RUNX2 or RUNX2^R131G proteins were expressed in HeLa cells. Nuclear extracts from transfected HeLa cells were incubated with biotinylated wild type (Wt) or mutant (Mu) RUNX2 binding oligonucleotides for 1h at RT. Streptavidin coated Sepharose CL4B beads were added to nuclear protein/DNA complexes and incubated at RT for 1 hr with rotation. Reacted (A) or non-reacted (B) nuclear extracts were separated in 10% SDS-polyacrylamide gels, transferred, and immunoblotted with antibodies to RUNX2. Lamin B antibody was used as internal loading control.