Self-Diffusion of Entangled Linear and Circular DNA Molecules:  Dependence on Length and Concentration

Robertson, Rae M.; Smith, Douglas E.

doi:10.1021/ma070051h

Cited by 88 publications

(117 citation statements)

References 35 publications

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“…In the case of DNA, we recently measured diffusion coefficients for entangled linear and relaxed circular molecules by tracking the Brownian motion of single molecules with fluorescence microscopy. 11 We found that the scaling of the diffusion coefficient with concentration and length was consistent with the reptation model for both topologies, but relaxed circular DNA diffused up to 7 times faster than linear DNA of the same length and concentration (45 kbp at 1 mg/mL). Thus, relaxed circular DNA appears to be less effective than linear DNA at producing restrictive entanglements.…”

Section: Introductionsupporting

confidence: 68%

Direct Measurement of the Confining Forces Imposed on a Single Molecule in a Concentrated Solution of Circular Polymers

Robertson

Smith

2007

Macromolecules

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We measure the forces confining the displacement of a single DNA molecule embedded within a concentrated solution of long relaxed circular DNA molecules (115 kbp at 1 mg/mL) using optical tweezers. We compare these measurements with our previous measurements of forces imposed by entangled linear DNA molecules of the same length and concentration. A tube-like confining field and three relaxation modes were observed, but the tube radius was 25% smaller (=0.6 µm) and the longest relaxation time ∼3 times shorter (=11 s) than with linear molecules, consistent with recent theoretical predictions by Iyer, Lele, and Juvekar. For displacements greater than the tube radius, the confining force imposed by circular polymers was substantially lower and shorter range than that measured with linear polymers.

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Section: Introductionsupporting

confidence: 68%

Direct Measurement of the Confining Forces Imposed on a Single Molecule in a Concentrated Solution of Circular Polymers

Robertson

Smith

2007

Macromolecules

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“…The DNA concentration within phi29, like many other doublestranded DNA bacteriophages, is very high (∼0.5 g/mL), and one would expect excluded volume and chain entanglements to strongly restrict molecular motion (39,47,48). The effect of entanglements on the dynamics of polymers in melts and concentrated solutions has been successfully predicted by reptation models, in which polymer motion is restricted to a tube-shaped region parallel to the chain contour (39,49).…”

Section: Discussionmentioning

confidence: 99%

Nonequilibrium dynamics and ultraslow relaxation of confined DNA during viral packaging

Berndsen

Keller²,

Grimes

et al. 2014

Proc. Natl. Acad. Sci. U.S.A.

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Many viruses use molecular motors that generate large forces to package DNA to near-crystalline densities inside preformed viral proheads. Besides being a key step in viral assembly, this process is of interest as a model for understanding the physics of charged polymers under tight 3D confinement. A large number of theoretical studies have modeled DNA packaging, and the nature of the molecular dynamics and the forces resisting the tight confinement is a subject of wide debate. Here, we directly measure the packaging of single DNA molecules in bacteriophage phi29 with optical tweezers. Using a new technique in which we stall the motor and restart it after increasing waiting periods, we show that the DNA undergoes nonequilibrium conformational dynamics during packaging. We show that the relaxation time of the confined DNA is >10 min, which is longer than the time to package the viral genome and 60,000 times longer than that of the unconfined DNA in solution. Thus, the confined DNA molecule becomes kinetically constrained on the timescale of packaging, exhibiting glassy dynamics, which slows the motor, causes significant heterogeneity in packaging rates of individual viruses, and explains the frequent pausing observed in DNA translocation. These results support several recent hypotheses proposed based on polymer dynamics simulations and show that packaging cannot be fully understood by quasistatic thermodynamic models.soft matter | virology | DNA condensation D NA packaging is both a critical step in viral assembly and a unique model for understanding the physics of polymers under strong confinement. Before packaging, the DNA (∼6-60 μm long) forms a loose random coil of diameter ∼1-3 μm. After translocation into the viral prohead (∼50-100 nm in diameter), a ∼10,000-fold volume compaction is achieved. Packaging is driven by a powerful molecular motor that must work against the large forces resisting confinement arising from DNA bending, repulsion between DNA segments, and entropy loss (1-8).DNA packaging in bacteriophages phi29, lambda, and T4 has been directly measured via single-molecule manipulation with optical tweezers and the packaging motors have been shown to generate forces of >60 pN, among the highest known for biomotors, while translocating DNA at rates ranging from ∼100 bp (for phage phi29, which packages a 19.3-kbp genome into a 42 × 54-nm prohead shell) up to as high as ∼2,000 bp/s (for phage T4, which packages a 171-kbp genome into a 120 × 86-nm prohead) (9-15). The force resisting packaging rises steeply with prohead filling and has been proposed to play an important role in driving viral DNA ejection (16).Recently, a variety of theoretical models for viral DNA packaging have been proposed (3)(4)(5)(17)(18)(19)(20)(21). The simplest treat DNA as an elastic rod with repulsive self-interactions and assume that packaging is a quasistatic thermodynamic process, i.e., that the DNA is able to continuously relax to a free-energy minimum state (3)(4)(5)(19)(20)(21). The DNA arrangement is generally assumed ...

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“…In particular, the conformation and relaxation of large polymer chains [62,63], as well as interaction with other chains [64], have been investigated in great detail by single-molecule techniques. Further, as cyclic and star-branched DNA are relatively easily obtainable, topology eff ects on polymer dynamics have also been investigated [65,66]. Th e study of large DNA is facilitated by its size-a fully labeled DNA helix is easily imaged by fl uorescence microscopy and changes in position and shape can be easily and precisely followed.…”

Section: Structure/conformation and Physics Of Non-conjugated Polymersmentioning

confidence: 99%

Conformation and physics of polymer chains: a single-molecule perspective

Vácha

Habuchi

2010

NPG Asia Mater

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N anotechnology has been one of the driving forces of science and technology over the past 20 years. Scaling down the size of an object to nanometer dimensions can result in dramatic changes to its physical properties. With recent progress in the nanoscience and nanotechnology of polymeric materials, new methods are now required for the characterization of polymer structures and properties. Despite the great success of electron microscopy and scanning probe microscope techniques, there remains a need for a non-invasive method that could provide information not only on the structure but also on the physical phenomena occurring on the nanoscale level. One method that has recently emerged as an excellent tool for nanoscale characterization is single-molecule optical detection and spectroscopy. Single-molecule spectroscopy relies on the detection and measurement of the fl uorescence signal and spectra from single isolated molecules or polymer chains (Figure 1). Th ese can either be immobilized in a solid matrix or be diff using in a solution or melt. In all cases, the concentration of the emitting molecules must be suffi ciently low to ensure that the spatial separation between individual molecules is larger than the optical resolution of the setup, typically on the order of micrometers. As the fl uorescence signal from a molecule is strongly infl uenced by the nanoscale environment, measuring many molecules one by one provides the distribution of physical properties of the respective nano-environments. Moreover, monitoring a molecule over a period of time reveals dynamic changes in its environment. Th ese static and dynamic heterogeneities of nanoscale properties are averaged and hidden in the usual ensemble experiments. Th e strength of fl uorescence detection is in its sensitivity and low background level -emissions of just a few hundred photons per second can be readily detected. Th e technique also off ers remarkable dynamic range, from sub-nanosecond timescales to seconds and longer, and is non-invasive, making it possible to detect emissions from molecules buried deep in a sample.Single-molecule spectroscopy has evolved since the beginning of the 1990s from low-temperature, high-resolution optical spectroscopic

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Self-Diffusion of Entangled Linear and Circular DNA Molecules: Dependence on Length and Concentration

Cited by 88 publications

References 35 publications

Direct Measurement of the Confining Forces Imposed on a Single Molecule in a Concentrated Solution of Circular Polymers

Direct Measurement of the Confining Forces Imposed on a Single Molecule in a Concentrated Solution of Circular Polymers

Nonequilibrium dynamics and ultraslow relaxation of confined DNA during viral packaging

Conformation and physics of polymer chains: a single-molecule perspective

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