Experimental and Computational Studies on DNA Electrophoresis in Lyotropic Polymer Liquid Crystals
Wei, Ling (author)
Van Winkle, David H. (professor directing dissertation)
Shanbhag, Sachin (university representative)
Xiong, Peng (committee member)
Rikvold, Per Arne (committee member)
Wahl, Horst D. (committee member)
Florida State University (degree granting institution)
College of Arts and Sciences (degree granting college)
Department of Physics (degree granting department)
Electrophoresis as an analytical technique has made considerable contributions to the separations and analysis of macromolecules in biology-related research. Pluronic gels, which are composed of orderly packed spherical micelles assembled by tri-block copolymers, have been developed as novel sieving media to separate oligonucleotides, duplex DNA molecules and proteins, providing ease of manipulations due to their thermo-reversibility and higher resolution in comparison with other polymer gels. Electrophoretic mobility of short double-stranded DNA molecules in pluronic F127 is reported to have a non-monotonic dependence on DNA length, which is not observed in other polymer-based sieving media or explained by any well-developed theories. In this dissertation, the unusual DNA-length dependence of electrophoretic mobility is experimentally investigated in several different pluronic gels, and the DNA dynamics in pluronic liquid crystals is systematically studied by coarse-grained Brownian dynamics simulations. The crystal structures and micelle dimensions of pluronics P105, P123 and F127 are characterized by atomic force microscopy, small-angle x-ray scattering, small-angle neutron scattering and dynamic light scattering. Two-dimensional gel electrophoresis is performed and the electrophoretic mobility of DNA molecules in the size range of 20-500 bp is measured in pluronics P105, P123 and F127. The unusual DNA length-dependent mobility is consistently obtained in three pluronic gels, where the mobility of very short DNA molecules increases with increasing DNA length, and the mobility of long DNA molecules monotonically decreases with DNA length. Superposed on the rising and falling trends are the subtle oscillations of mobility with DNA length in the intermediate regime. Brownian dynamics simulations are implemented to numerically calculate the DNA mobility in pluronic lattices, by including the short-ranged intra-molecular hydrodynamic interactions, and modeling the interactions between DNA molecules and pluronic micelles via a repulsive force and entanglement effect. The rise, fall and oscillations of mobility with DNA length, as obtained in experimental measurements, are reproduced by the Brownian dynamics simulations, and essential physics that dominates the unusual features of mobility is extracted from the simulations. In addition, electric field-dependent mobility of DNA molecules in pluronic lattices is studied by Brownian dynamics simulations, and the conceptual connection between high-field simulations along specific field directions and low-field experiments in bulk gels is established, and the Brownian dynamic simulations are proven to be an appropriate approach to interpret the DNA electrophoretic dynamics in pluronic matrices. Moreover, electrophoretic mobility of duplex DNA flanked by single-stranded overhangs is measured in pluronic gels, and it is shown that the mobility of DNA with overhangs is higher than the corresponding blunt-ended DNA molecules. Brownian dynamics simulations are carried out, and the enhancement of mobility for DNA with overhangs is captured by the simulations. By integrating numerical simulations with experimental measurements, the fundamental physical quantities and interactions that manipulate the DNA electrophoretic migration in pluronic liquid crystals are revealed. Understanding the unusual DNA length-dependent mobility in pluronic gels potentially provides profound insights in designing and optimizing high-performance sieving matrices for size-based separation purposes.
Brownian dynamics simulation, DNA, electrophoresis, electrophoretic mobility, hydrodynamic interaction, pluronic
October 24, 2016.
A Dissertation submitted to the Department of Physics in partial fulfillment of the requirements for the degree of Doctor of Philosophy.
Includes bibliographical references.
David H. Van Winkle, Professor Directing Dissertation; Sachin Shanbhag, University Representative; Peng Xiong, Committee Member; Per Arne Rikvold, Committee Member; Horst Wahl, Committee Member.
Florida State University
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