Fluorescent observation of individual DNA molecules on phospholipid bilayers and its application to analysis of DNA-protein interactions

Abstract

The advantage of single-molecule experiments on analysis of DNA-protein interactions is that it can manipulate the physical form of individual DNA molecules. In these experiments, surface treatments for anchoring individual DNA molecules are important. In this study, we carried out fluorescent observations of individual DNA molecules on phospholipid bilayers. First, we constructed a PDMS microchannel and neutravidin was immobilized on the glass surface to tether individual DNA molecules. And then, DOPC liposomes were applied to the microchannel and the lipid bilayers were constructed. Biotinylated DNA molecules labeled with fluorescent dyes were then injected to the channel and they were tethered to the glass surface. As a result, we obtained clear images of the DNA molecules. Intriguingly, in addition, we found that if the fluorescent observation was carried out with the phospholipid bilayers, photobleaching of the fluorescently-labeled DNA was slower by comparison with that without the bilayers. Moreover, we applied this method to DNA-protein interactions at a single-molecule level.