Figure 5
Identification of the 24–10 aptamer region optimal for R8 binding. The binding of R8-GST to the full length 24–10 aptamer, as well as its variants appropriately shortened from the 5′ or 3′ end, was tested. (A) Pull down assay. Lanes: (1) full length 24–10, (2) 24–10 (5′/−10), (3) 24–10 (3′/−10), (4) 24–10 (5′/−10 and 3′/−10), (5) 24–10 (5′/−10 and 3′/−20), (6) 24–10 (5′/−20 and 3′/−10), (7) 24–10 (5′/−30 and 3′/−10) and (8) 24–10 (5′/−40 and 3′/−10) aptamer variants immobilized on streptavidin agarose beads were incubated with the analyzed protein. The bound protein was denatured. Next, one fortieth of the volume of each protein sample was separated on 12% SDS-PAGE followed by Coomassie staining. This is one of three independent experiments which is representative. (B) Densitometric analysis of data from three independent pull down assays was done using Multispectral Imaging System IMAGER with Launch VisionWorksLS. Results are the mean of three measurements. The error bars represent standard deviation. The results were normalized relative to signal from the protein sample bound by the full length aptamer (100%).
