Using recombinant Flag-tagged Dcr-2 and His-tagged protein X, pull-down assays were performed to determine whether protein X and Dcr-2 interact directly. The recombinant proteins (either alone or in combination) in binding buffer (150 mM NaCl, 1 mM EDTA, 50 mM Tris pH 7.5, 1% Triton X-100) were pulled-down by Flag antibodies coupled to Sepharose beads. Following extensive washing, the proteins were eluted from the beads with SDS buffer, and duplicate sets of samples were analyzed by SDS PAGE. Samples were also taken of the proteins before they were used in the pull-down assay and these were run on the same gel (input lanes in Figure 2B). One gel was stained with Coomassie blue dye (Figure 2A), and the other was transferred to nitrocellulose for Western Blotting. The membrane was probed with a His-antibody (Figure 2B).
The question asks: Do Dcr‐2 and protein X interact directly?
My reasoning is that they DO interact directly. In Fig 2B: the intense bands on LHS shows that Dcr-2 interacts with protein X-that is bound to anti-His. In Fig 2A: the protein X interacts with the Dcr-2 bound to antibody FLAG. I feel I'm not explaining this very though...Could someone please explain their reasoning? Thank you