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Nanofabricated quartz cylinders for angular trapping: DNA supercoiling torque detection

Department of Physics, Laboratory of Atomic and Solid State Physics, Cornell University, Ithaca, New York 14853, USA.
Nature Methods (Impact Factor: 25.95). 04/2007; 4(3):223-5. DOI: 10.1038/nmeth1013
Source: PubMed

ABSTRACT We designed and created nanofabricated quartz cylinders well suited for torque application and detection in an angular optical trap. We made the cylinder axis perpendicular to the extraordinary axis of the quartz crystal and chemically functionalized only one end of each cylinder for attachment to a DNA molecule. We directly measured the torque on a single DNA molecule as it underwent a phase transition from B-form to supercoiled P-form.

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Available from: Christopher Deufel, Aug 18, 2014
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    • "One focus has been on modelling DNA. For example, in experiments [1] [2] [3] single molecules of twist-storing polymers such as double stranded DNA can be held torsionally constrained and under constant stretching force. These experiments show abrupt phase transitions known as buckling, and the formation of conformational structures known as plectonemes. "
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    ABSTRACT: We consider a simple lattice model of a topological phase transition in open polymers. To be precise, we study a model of self-avoiding walks on the simple cubic lattice tethered to a surface and weighted by an appropriately defined writhe. We also consider the effect of pulling the untethered end of the polymer from the surface. Regardless of the force we find a first-order phase transition which we argue is a consequence of increased knotting in the lattice polymer, rather than due to other effects such as the formation of plectonemes.
    Journal of Physics A Mathematical and Theoretical 10/2014; 48(6). DOI:10.1088/1751-8113/48/6/065002 · 1.69 Impact Factor
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    • "The particles are designed to have their extraordinary optical axis perpendicular to the axis of cylindrical symmetry, and are chemically functionalized at only one of the bases (Fig. 6). Our protocol was developed from a previous implementation (Deufel et al., 2007), but modified to comply with restrictions imposed by the Stanford Nanofabrication Facility, which precluded placing anti-reflective coatings on the back sides of wafers. "
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    ABSTRACT: We present details of the design, construction, and testing of a single-beam optical tweezers apparatus capable of measuring and exerting torque, as well as force, on microfabricated, optically anisotropic particles (an "optical torque wrench"). The control of angular orientation is achieved by rotating the linear polarization of a trapping laser with an electro-optic modulator (EOM), which affords improved performance over previous designs. The torque imparted to the trapped particle is assessed by measuring the difference between left- and right-circular components of the transmitted light, and constant torque is maintained by feeding this difference signal back into a custom-designed electronic servo loop. The limited angular range of the EOM (+/-180 degrees ) is extended by rapidly reversing the polarization once a threshold angle is reached, enabling the torque clamp to function over unlimited, continuous rotations at high bandwidth. In addition, we developed particles suitable for rotation in this apparatus using microfabrication techniques. Altogether, the system allows for the simultaneous application of forces (approximately 0.1-100 pN) and torques (approximately 1-10,000 pN nm) in the study of biomolecules. As a proof of principle, we demonstrate how our instrument can be used to study the supercoiling of single DNA molecules.
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    • "The development of magnetic tweezers [31] [35] or the use of quartz cylinders in optical tweezers [9] allow researchers to investigate the in-vitro response of DNA molecules to torsional stress. Studies of the behaviour of this twist storing polymer are not just a game for physicists as it has been clearly established that, e.g. the assembly of RecA could be stalled by torsional constraints [41], or that the rate of formation and the stability of the complex formed by promoter DNA and RNA polymerase depends on the torque present in the DNA molecule [25]. "
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    ABSTRACT: The linking and writhing numbers are key quantities when characterizing the structure of a piece of supercoiled DNA. Defined as double integrals over the shape of the double helix, these numbers are not always straightforward to compute, though a simplified formula was established in a theorem by Fuller [Proc. Natl. Acad. Sci. U.S.A. 75, 3557 (1978)]. We examine the range of applicability of this widely used simplified formula, and show that it cannot be employed for plectonemic DNA. We show that inapplicability is due to a hypothesis of Fuller theorem that is not met. The hypothesis seems to have been overlooked in many works.
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