415905 Synthesis, Characterization, and Single Molecule Dynamics of Branched DNA Polymers

Tuesday, November 10, 2015: 9:30 AM
251B (Salt Palace Convention Center)
Danielle J. Mai1, Amanda B. Marciel2 and Charles M. Schroeder1, (1)Chemical and Biomolecular Engineering, University of Illinois at Urbana-Champaign, Urbana, IL, (2)Center for Biophysics and Computational Biology, University of Illinois Urbana-Champaign, Urbana, IL

We report the synthesis, characterization, and single molecule dynamics of graft-onto branched DNA polymers. In this work, we utilize a two-step scheme to graft polymer branches to template DNA backbones, thereby producing precise star, H, and comb-shaped homopolymers (DNA only) and copolymers (poly(ethylene glycol) branches on DNA). Branched DNA components are synthesized via polymerase chain reaction with chemically modified deoxyribonucleotides and primers: branches include a terminal azide group, and template backbones contain internal dibenzylcyclooctyne (DBCO) groups. In this way, we utilize strain-promoted [3+2] alkyne-azide cycloaddition "click" chemistry for facile grafting of azide-terminated branches at DBCO sites along backbones. Copper-free click reactions are bio-orthogonal and nearly quantitative when carried out under mild conditions. Structurally defined branched polymers are characterized via polyacrylamide gel electrophoresis, denaturing high performance liquid chromatography, and matrix assisted laser desorption/ionization mass spectrometry.

We also extend this two-step scheme to generate macromolecular DNA branched polymers with "short," branches (1-10 kilobase pairs) and "long" backbones (10-30 kilobase pairs). In some cases, branches contain internal Cy5 labels (red fluorescence). In the presence of a DNA stain (e.g., SYTOX Green®), we independently observe DNA branches and backbones via dual-color single molecule fluorescence microscopy (SMFM). Our imaging approach allows for characterization of these materials at the single molecule level (e.g., quantification of polymer contour length and branch distributions for varying synthetic conditions). We also apply SMFM to single polymer rheology experiments on branched polymers. Specifically, we utilize microfluidic devices to study branched DNA in tethered shear and planar extensional flow fields. We study conformational relaxation dynamics by characterizing backbone relaxation from high stretch, and we find branched polymer relaxation depends on the number of branches and position of branch points along the main chain backbone. In future work, we seek to probe the impact of branching on steady-state extension and dynamics in dilute, semi-dilute, and concentrated solutions. In this way, our work will contribute to the overall understanding of topologically complex polymer melts and solutions.

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See more of this Session: Excellence in Graduate Polymer Research
See more of this Group/Topical: Materials Engineering and Sciences Division