Parallel multiplex thermodynamic analysis of coaxial base stacking in DNA duplexes by oligodeoxyribonucleotide microchips

Abstract

Parallel thermodynamic analysis of the coaxial stacking effect of two bases localized in one strand of DNA duplexes has been performed. Oligonucleotides were immobilized in an array of three-dimensional polyacrylamide gel pads of microchips (MAGIChips'). The stacking effect was studied for all combinations of two bases and assessed by measuring the increase in melting temperature and in the free energy of duplexes formed by 5mers stacked to microchip-immobilized 10mers. For any given interface, the effect was studied for perfectly paired bases, as well as terminal mismatches, single base overlaps, single and double gaps, and modified terminal bases. Thermodynamic parameters of contiguous stacking determined by using microchips closely correlated with data obtained in solution. The extension of immobilized oligonucleotides with 5,6-dihydroxyuridine, a urea derivative of deoxyribose, or by phosphate, decreased the stacking effect moderately, while extension with FITC or Texas Red virtually eliminated stacking. The extension of the immobilized oligonucleotides with either acridine or 5-nitroindole increased stacking to mispaired bases and in some GC-rich interfaces. The measurements of stacking parameters were performed in different melting buffers. Although melting temperatures of AT- and GC-rich oligonucleotides in 5 M tetramethylammonium chloride were equalized, the energy of stacking interaction was significantly diminished.

Reducing Facet Nucleation during Algorithmic Self-Assembly

“…For example, ∆G°1 could be different for each sticky end sequence and ∆G°2 might not be a simple sum if steric effects come into play. Furthermore, it is possible that the favorable energy of blunt end stacking 29 between sT1 and sT2 needs to be taken into account; our model neglects it. There may also be small but nonzero energies between nonmatching sticky ends as well as stoichiometry imbalance among the tile types.…”

One Dimensional Boundaries for DNA Tile Self-Assembly

“…Are the parameters used in the simulations reasonable? Based on generic nearest-neighbor parameters for base stacking within a duplex [10], coaxial base stacking at nick sites [13], and dangling ends [2], we estimate 5 rule tile sticky ends to bind with K eq between 2 and 200 µM, and boundary tile sticky ends to bind with K eq between 10 and 100 nM at 25 • C. The multi-kTAM simulations used 6 µM and 34 pM respectively, while the reversible aggregation simulations used 1 nM for the boundary tiles. Thermodynamic parameters for our DNA tiles should be measured experimentally.…”

Modelling DNA origami self-assembly at the domain level

“…where ∆G shape s,s represents the contributions from the entropic costs of scaffold loop formation [26,29,37], and ∆G stack s,s is a contribution from the coaxial stacking of duplex sections [38][39][40][41]. We discuss our approximations for the various contributions to ∆G 0 s,s in Section II C. The value of ∆G 0 s,s associated with hybridization of a domain determines the ratio between binding and unbinding transition rate constants, but does not determine their absolute values.…”

“…It is well known that coaxial stacking of bases across a nick in the DNA backbone can stabilize the flanking duplexes [38][39][40][41]. We add a coaxial stacking contribution ∆G stack (T ) to the free energy of each state wherever two adjacent scaffold domains are both hybridized to staples.…”

“…[29]. ∆G stack (T ), which has been considered by SantaLucia and others [38][39][40][41], is much more weakly constrained than other aspects of the nearest-neighbour model. We thus treat n as an adjustable parameter of the model, and explore the consequences of varying n in Section III.…”