Simulations of the overshoot in the build-up of orientation of long DNA during gel electrophoresis based on a distribution of oscillation times.
The periodic extension-contraction motion observed for long DNA molecules undergoing agarose gel electrophoresis in a constant field is believed to be important for the separation mechanism in pulsed field gel electrophoresis. These oscillations give rise to an overshoot and an undershoot in the ensemble orientation of DNA in the beginning of a field pulse, when the molecules oscillate coherently. After approximately one oscillation cycle, the coherence between the molecules is lost, and a constant, cycle-averaged orientation is reached. In this paper we simulate this build-up of the ensemble orientation of DNA by using a distribution of oscillation times (the time between two consecutive compressed conformations) determined by fluorescence microscopy for YOYO-stained DNA. Six different orientation profiles, describing the orientation during one oscillation cycle, were used. The simulated orientation responses are compared with an orientation response measured by linear dichroism (LD) under the same experimental conditions as in the microscopy study. We found that the choice of orientation profile during the oscillation is important. Best agreement between the simulated and the experimental orientation response was obtained for an orientation profile based on a theoretical model by Schurr and Smith (Biopolymers 1990, 29, 1161-1165). The influence of the distribution of oscillation times and its standard deviation on the orientation response was also investigated. Furthermore, simulations at different field strengths and DNA sizes were performed and found to agree quite well with the experimentally obtained LD data.
Department of Physical Chemistry, Chalmers University of Technology, Gothenburg, Sweden.
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Carlsson
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Larsson
D003198:Computer Simulation; D004247:DNA / Q000737:chemistry*; D004587:Electrophoresis, Agar Gel; D008856:Microscopy, Fluorescence; D008956:Models, Chemical; D013057:Spectrum Analysis; D013997:Time Factors
D004247:DNA