Measuring Bending of Intrinsically Bent DNA

Intrinsically bent DNA sequences influence nucleosome positioning, which, in turn, helps regulate gene expression. The iconic example is a run of four or more contiguous adenine bases, known as an A-tract. While details of A-tract structure are well-studied, little is known about the mechanical properties. We seek to measure A-tract stiffness from the angular fluctuations of a short (~ 60 bp) DNA sequence with zero, one, or more A-tracts (repeated in phase with the helix screw). Our approach leverages DNA nanotechnology to embed the A-tract-laden sequence between a pair of microns-long rigid rods, which mechanically magnify the relative orientation of the sequence ends. The angular fluctuations are captured by video fluorescence microscopy, analyzed using circular statistics and compared with published results.