Thermodynamics and Kinetics of DNA Origami Cross-Tile Array Formation

Abstract

The final reading approval of the thesis was granted by Elton Graugnard, Ph.D., Chair of the Supervisory Committee. The thesis was approved by the Graduate College. However, these structures are far from ideal and contain a number of defects that limit the adoption of this approach for manufacturing. In order to create large defect-free DNA arrays, further study is needed into the fundamental mechanisms governing array formation. Toward this goal, the thermodynamics and kinetics of DNA array formation were investigated using a DNA origami cross-tile that assembles into arrays through DNA hybridization. The assembly of dimers, quadramers, and unbound arrays in solution from monomers with complementary dye and quencher labeled hybridization interfaces was monitored by observing the change in fluorescence of the solution as a function of temperature and over time under varying buffer conditions and temperatures. The melting temperature of each structure was measured and generally increased with an increasing number of active sticky-ends per monomer. Values for standard thermodynamic parameters were determined for each array design. The reaction kinetics data were fit with a second order reaction model, and the effective reaction rate increased with viii increasing buffer magnesium concentrations and increasing temperatures. Finally, it was determined that large, unbounded 2D DNA origami cross-tile arrays sediment out of solution in only a few hours. The findings of this study provide insight into the mechanisms of DNA array formation and establish practical ranges for key processing parameters.