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Single-molecule studies of DNA dynamics and intermolecular forces

  • Author(s): Robertson, Rae Marie
  • et al.
Abstract

DNA molecules were used as a model system to investigate fundamental problems in polymer physics; namely, how molecular length, topology and concentration influence the dynamical properties of polymers. A set of DNA molecules suitable for polymer studies was prepared using molecular biology techniques. Video fluorescence microscopy and single-molecule tracking were used to determine self- diffusion coefficients of DNA molecules. Optical tweezers were used to measure the intermolecular forces confining entangled DNA molecules. Scaling of diffusion with molecular length was in agreement with the Zimm model for dilute solutions of linear and circular DNA, indicating that excluded volume effects are appreciable for both topologies. Scaling of diffusion with concentration was also determined for the four possible topological combinations of linear and circular molecules: linear DNA diffusing in a solution of linear DNA, linear DNA in circular DNA, circular in circular, and circular in linear. For lower concentrations molecular topology had little effect and scaling was in agreement with that of the Rouse model. As concentration was increased topology played a much larger role and scaling crossed over to that of the reptation model, predicted to describe the dynamics of entangled polymers. The notable exception was the strongly hindered diffusion observed for a circular molecule diffusing in an entangled linear solution, suggesting the importance of constraint release. Using a new experimental approach with optical tweezers, a tube- like field confining a single entangled molecule was measured, in accord with the key assumption of the reptation model. A time-dependent harmonic potential opposed displacement transverse to the molecular contour, and the force relaxations following displacement were composed of three distinct modes. A characteristic tube radius of the entangled solution was also determined, close to the classically predicted value. The dependence of the above findings on molecular topology and concentration was also investigated. In particular, for an entangled solution of circular DNA of the same length and concentration, the confining tube radius was 25% smaller and the longest relaxation time was ̃3 times shorter than with linear DNA. For large displacements the confining force for circular DNA was also substantially lower and shorter range than that measured with linear DNA

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