Studying the affinity, kinetic stability, and specificity of RNA/protein interactions: SINE ncRNA/Pol II complexes as a model system.

The number of documented interactions between proteins and noncoding RNAs (ncRNA) of all types has grown rapidly in the past several years. A current challenge is to experimentally characterize these interactions to ultimately understand their biological roles at a mechanistic level, which will require a combination of multiple experimental techniques. One such category of techniques is biochemical assays that determine the affinity, kinetic stability, and specificity of ncRNA/protein complexes. Here we describe how to experimentally determine these important parameters using electrophoretic mobility shift assays (EMSAs). The interaction between mammalian SINE-encoded ncRNAs and human RNA polymerase II is presented as a model system; however, the experiments could be readily adapted to other ncRNA/protein complexes.