Hybridization Thermodynamics of DNA Oligonucleotides during Microchip Capillary Electrophoresis

Abstract

Capillary electrophoresis (CE) is a powerful analytical tool for performing separations and characterizing properties of charged species. For reacting species during a CE separation, local concentrations change leading to nonequilibrium conditions. Interpreting experimental data with such nonequilibrium reactive species is nontrivial due to the large number of variables involved in the system. In this work we develop a COMSOL multiphysics-based numerical model to simulate the electrokinetic mass transport of short interacting ssDNAs in microchip capillary electrophoresis. We probe the importance of the dissociation constant, K(D), and the concentration of DNA on the resulting observed mobility of the dsDNA peak, μ(w), by using a full sweep of parametric simulations. We find that the observed mobility is strongly dependent on the DNA concentration and K(D), as well as ssDNA concentration, and develop a relation with which to understand this dependence. Furthermore, we present experimental microchip capillary electrophoresis measurements of interacting 10 base ssDNA and its complement with changes in buffer ionic strength, DNA concentration, and DNA sequence to vary the system equilibria. We then compare our results to thermodynamically calculated K(D) values.

Confinement effects on DNA hybridization in electrokinetic micro‐ and nanofluidic systems

“…For the dispensing step, the electric field applied to the north channel leg is reversed, and an electric field of 49.3 kV/m guides the sample plug from the loading step past a detection region 20 mm from the channel cross. The 100 nm channels require a larger injection sample plug to remain detectable 20 mm from the cross, so we add an additional 1 s gating step to these injections . For each channel depth, ten experimental trials are collected.…”

“…al. recently presented a method to simulate NECEEM for DNA hybridization systems and calculate K D in a microfluidic channel using equations for mass transport, electromigration, and reaction kinetics . As this model most closely represents our system, we employ the same method to estimate our disassociation constants.…”

“…The simulation solves for the electrophoretic transport of a finite‐sized plug of three reacting species in a straight channel, assuming that the advective velocity equals 0 . This assumption is made because the area‐averaged electroosmotic flow in microchannels falls within 95% of the Helmholtz‐Smoluchowski velocity, allowing us to neglect non‐uniform electroosmotic flow .…”

Kinetic Study of Chiral Intermolecular Interactions by Moment Analysis Based on Affinity Capillary Electrophoresis

“…In this work, commercially available COMSOL Multiphysics version 5.2b with microfluidics module was used to perform the simulation. This software has been utilized to simulate the behavior of capillary electrophoresis, , which we think is appropriate for the simulation, as long as the data points and time step are appropriate.…”