DNA-Protein Interactions Studied Directly Using iSCAT Imaging of GNP-Tagged Proteins.

Abstract

Many protein interactions with DNA are reliant on the presence of specific DNA sequences, adducts, or structures. In bulk-phase experiments such DNA features are facile to include in a study. For single-molecule imaging this can be more difficult, because the constraints of the assay limit the variety of adducts that can be used. Surface-immobilized DNA provides an ideal compromise, and the use of interferometric scattering microscopy allows for high-speed imaging of these interactions. Furthermore, this technique offers the ability to identify binder stoichiometry and the composition of protein complexes. Its implementation is relatively simple; however data analysis and deconvolution are more challenging. In this chapter we examine how this technique is implemented and reveal software that can be used to deconvolute the images. Altogether, we hope to make this technique more accessible for studying specific DNA-protein interactions on tailored substrates.