Local conformation of confined DNA studied using emission polarization anisotropy

When confined in nanochannels with dimensions smaller than the DNA radius of gyration, DNA will extend along the channel. We investigate long DNA confined in nanochannels, using fluorescence microscopy and intercalated dyes. Studies of the dynamics and statics of DNA in such nanoscale confinements as a function of e.g. degree of confinement and ionic strength have yielded new insights into the physical properties of DNA with relevance for applications in genomics as well as fundamental understanding of DNA packaging in vivo. Our work extends the field by not only studying the location of the emitting dyes along a confined DNA molecule but also monitoring the polarization of the emitted light. By measuring the emission polarized parallel and perpendicular to the extension axis of the stretched DNA, information on the local spatial distribution of the DNA backbone can be obtained. Comparing polarizations in two directions for DNA confined in channels of effective diameters of 85 nm and 170 nm reveals a striking difference. Whereas the DNA in the larger channels shows an isotropic polarization of the emitted light, the light is to a large extent polarized perpendicular to the elongation of the DNA in the smaller channels. We expect this technique to have a large impact on the studies of changes in DNA conformation induced by protein binding or during DNA compactation as well as in fundamental polymer physics studies of DNA in confined environments, for example in bacterial spores and viruses