Bulletin of the American Physical Society
APS March Meeting 2014
Volume 59, Number 1
Monday–Friday, March 3–7, 2014; Denver, Colorado
Session T10: Focus Session: Confined Nucleic Acids II |
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Sponsoring Units: DBIO Chair: Kevin Dorfman, University of Minnesota Room: 201 |
Thursday, March 6, 2014 11:15AM - 11:27AM |
T10.00001: Conformation of nanoconfined DNA as a function of ATP, AMP, CTP, Mg$^{2+}$, and dye binding Maedeh Roushan, Robert Riehn DNA molecules stretch in nanochannels with a channel cross-section of 100x100 nm$^{2}$, thereby allowing analysis by observation of a fluorescent dye. The length and configuration of DNA can be directly observed, and the effect of different DNA-binding proteins on DNA configuration can be studied. Recently, we reported on the ability of T4 ligase to transiently manipulate DNA as a function of ATP and magnesium exposure. In this process we have extensively probed the interactions of dyes and enzyme co-factors with DNA under nanoconfinement. We find negligible effects if DNA is visualized using groove-binding dyes such as DAPI. However, if an intercalating dye (YOYO-1) is used, we find a significant shortening of the DNA in the presence of ATP that we attribute to an interaction of dye and ATP (as well as AMP and CTP). We did not record a noticeable effect due to Mg$^{2+}$. [Preview Abstract] |
Thursday, March 6, 2014 11:27AM - 11:39AM |
T10.00002: Nanochannel Platform for Single-DNA Studies; From DNA-Protein Interaction to Large Scale Genome Sequencing Johan van der Maarel, Jeroen van Kan, Ce Zhang The study of nanochannel-confined DNA molecules is of importance from both biotechnological and biophysical points of view. We produce nanochannels in elastomer-based biochips with soft lithography using proton beam writing technology. The cross-sectional diameter of the channels is in the range of 50 to 300 nm. Single DNA molecules confined inside these channels can be visualized with fluorescence microscopy. Two related issues concerning DNA confined in such a nanospace will be discussed. For the first issue, the dynamic effects of nucleoid associated proteins (H-NS and HU) and protamine on the conformation and condensation of DNA will be presented. We use a novel, cross-channel device, which allows the monitoring of the conformational response after a change in environmental solution conditions in situ. The second issue concerns bottlebrush-coated DNA. The bottlebrush has an increased bending rigidity and thickness, which results in an amplified stretch once it is confined inside a nanochannel. It will be demonstrated that large-scale genomic organization can be sequenced using single DNA molecules on an array of elastomeric nanochannels with cross-sectional diameters of 200 nm. Overall, our results show that the effects of proteins on the conformation and folding of DNA are not only related to protein binding, osmolarity, and charge, but that the interplay with confinement in a nanospace is of paramount importance. [Preview Abstract] |
Thursday, March 6, 2014 11:39AM - 11:51AM |
T10.00003: Targeted manipulation of single large DNA molecules Zubair Azad, Robert Riehn We have developed a technique to manipulate one or more strands of DNA independently inside junctions of nanochannels. The work extends the concept of controlling DNA configurations through confinement by adding deliberate real-time control. The technique is based on independent control of fluid flow or voltages in the channels leading to the nanochannel junctions. By mismatching the flows into each of the channels, we create flow gradients at the channel junctions that manipulate the DNA configuration. Specific examples of DNA configuration control are the folding single DNA molecules, and the colocation of two independent molecules in the same channel segment. We believe that these manipulation techniques aid the study of DNA-DNA interactions. [Preview Abstract] |
Thursday, March 6, 2014 11:51AM - 12:27PM |
T10.00004: Entropy-driven DNA tug-of-war and confinement-induced reptation in 2D slitlike channels Invited Speaker: Chia-Fu Chou Given its simplicity in geometry and fabrication, nanofluidic confinement, in the form of nanoslits, nevertheless offers unique platforms for the study of molecular biophysics and single molecule analysis [1-3]. Here, we established an entropy-driven single DNA tug-of-war (TOW) system composed of two micro-to-nanofluidic interfaces bridged by a nanoslit. This surprisingly simple system enables us to study polymer TOW dynamics and the conformation recovery through entropic recoiling, without using sophisticated external force apparatus such as optical tweezers, magnetic tweezers, and atomic force microscopy [4]. By changing the slit length and depth, we determined the scaling behavior of the entropic recoiling force ($f_{rec})$ on the nanoconfinement ($h)$ to be $f_{rec}$ $\sim$ 1/$h$ for $h=$ 40-110 nm. This observation is also supported by our scaling analysis [5]. Further, we observed unexpected reptation of single DNA molecules in nanoslits of 25 nm height or less. The reptation behavior is quantitatively characterized using orientation correlation and transverse fluctuation analysis. We propose that tube-like polymer motion arises for a tense polymer under strong uniaxial confinement and the interaction with the surface-passivation polymers. \\[4pt] [1] L.J. Guo, X. Cheng, and C.F. Chou, \textit{Nano Lett}. \textbf{4}, 69 (2004).\\[0pt] [2] J. Gu, R. Gupta, C.F. Chou, Q. Wei, and F. Zenhausern, \textit{Lab Chip} \textbf{7}, 1198 (2007).\\[0pt] [3] T. Le\"{\i}chl\'{e}, Y.L. Lin, P.C. Chiang, K.T. Liao, S.M. Hu, and C.F. Chou, \textit{Sens. Actuators B} \textbf{161}, 805 (2012).\\[0pt] [4] J.W. Yeh, A. Taloni, Y.L. Chen, and C.F. Chou, \textit{Nano Lett}. \textbf{12}, 1597 (2012). (\textit{Nature} \textbf{482}, 442 (2012))\\[0pt] [5] A. Taloni, J.W. Yeh, and C.F. Chou, \textit{Macromolecules} \textbf{46}, 7989 (2013). [Preview Abstract] |
Thursday, March 6, 2014 12:27PM - 12:39PM |
T10.00005: Fluidic Switching in Nanochannels for the Control of a Synthetic DNA-based Motor C.S. Niman, M. Balaz, J.O. Tegenfeldt, P.M.G. Curmi, N.R. Forde, M. Zuckermann, Heiner Linke Processive molecular motors are thought to utilize a ``power stroke'' whereby chemical changes are converted into conformational changes, facilitating forward motion. We have developed a concept for a synthetic molecular motor, the Inchworm (IW), which harnesses salt-induced changes in DNA conformation$^{1}$ to achieve power strokes. To implement IW we must switch between four solutions (of varied salt concentration) surrounding DNA confined in a nanochannel (NC) while monitoring its response. We have developed NCs of radii 100-400 nm, with 10-20 nm wide top-slits for buffer exchange via diffusion from adjacent microfluidic channels$^{2}$. NCs are made in SiO$_{2}$ to allow for imaging through the substrate. To cycle through four buffers specifically designed microchannels are used$^{3}$. We measure changes in intensity when fluids containing fluorescent molecules are switched, with and without a pressure difference over the NCs. By fitting this data we extract effective diffusivity of molecules and determine fluid velocities, information that is crucial for evaluating IW performance. \\[4pt] [1] W.Reisner et al., PRL 2007, 99, 058302;\\[0pt] [2] M.Graczyk et al., J. Vac. Sci. {\&} Technol. B 2012, 30, 6;\\[0pt] [3] C.S.Niman et al., Lab Chip 2013, 13, 2389 [Preview Abstract] |
Thursday, March 6, 2014 12:39PM - 12:51PM |
T10.00006: Nanobiodevices for fast DNA separation and detection toward nanopore-based DNA sequencing Noritada Kaji, Takao Yasui, Yoshinobu Baba There is an increasing demand for using micro- and nanofabricated structures as tools for separation, manipulation, detection and analysis of biomolecules such as DNA and proteins. So far, we have developed fabrication techniques for constructing several types of nanostructures on quartz substrate for biomolecules separation, e.g., nanopillar and nanowall array structures, and demonstrated their analytical performances. Some important findings were that the nanopillar array pattern could control the DNA separation mode and electroosmotic flows in the nanopillar array structures were reduced according to the nanopillar spacing. Since these small confined nanospaces are suitable for manipulating biomolecules at a single molecule level, several approaches have been tried to analyze DNA denaturation and DNA-protein interactions in parallel. However, it is difficult to say that the observed phenomena reflect an intrinsic DNA property or DNA-protein interaction manner because all these approaches requires fluorescently labeled DNA molecules for observation. To address these issues, we are trying to develop a novel nanostructure-based and label-free detection system to integrate a biomolecule separation media and a detection system on a single chip. [Preview Abstract] |
Thursday, March 6, 2014 12:51PM - 1:03PM |
T10.00007: Quantum-Sequencing: Fast electronic single DNA molecule sequencing Josep Casamada Ribot, Anushree Chatterjee, Prashant Nagpal A major goal of third-generation sequencing technologies is to develop a fast, reliable, enzyme-free, high-throughput and cost-effective, single-molecule sequencing method. Here, we present the first demonstration of unique ``electronic fingerprint'' of all nucleotides (A, G, T, C), with single-molecule DNA sequencing, using Quantum-tunneling Sequencing (Q-Seq) at room temperature. We show that the electronic state of the nucleobases shift depending on the pH, with most distinct states identified at acidic pH. We also demonstrate identification of single nucleotide modifications (methylation here). Using these unique electronic fingerprints (or tunneling data), we report a partial sequence of beta lactamase (bla) gene, which encodes resistance to beta-lactam antibiotics, with over 95{\%} success rate. These results highlight the potential of Q-Seq as a robust technique for next-generation sequencing. [Preview Abstract] |
Thursday, March 6, 2014 1:03PM - 1:15PM |
T10.00008: Tunneling Currents through DNA Bases Tightly Constrained in a Fluid Channel Luke A Somers, Manuel Schottdorf, Meni Wanunu, Eva Y. Andrei Directing single-stranded DNA through a tunnel-junction is a strategy for rapid DNA sequencing. The main limiting factor in the viability of this method is coercing the ssDNA strand to pass only directly between the tunneling tips. This both ensures sequencing completely in order and minimizes the geometric effects on tunneling. We present such a device and results of tunneling through different bases. This device, employing graphene as a super-thin electrode, lies entirely in-plane rather than acting through a membrane. This geometry enables dense packing of devices with a minimum of fabrication. [Preview Abstract] |
Thursday, March 6, 2014 1:15PM - 1:27PM |
T10.00009: A Two Nanopore System for Controlling DNA Motion Tamas Szalay, Daniel Branton, Jene Golovchenko By positioning two nanopores with independently controllable bias voltages sufficiently close to capture a single molecule of DNA, the net motion and ionic current can be decoupled, enabling new studies of capture and stretching dynamics. We report on our two-nanopore system, including a novel chip geometry we have developed in order to optically monitor the position of the nanopores. [Preview Abstract] |
Thursday, March 6, 2014 1:27PM - 1:39PM |
T10.00010: Nanofluidic Device with Embedded Nanopore Yuning Zhang, Walter Reisner Nanofluidic based devices are robust methods for biomolecular sensing and single DNA manipulation. Nanopore-based DNA sensing has attractive features that make it a leading candidate as a single-molecule DNA sequencing technology. Nanochannel based extension of DNA, combined with enzymatic or denaturation-based barcoding schemes, is already a powerful approach for genome analysis. We believe that there is revolutionary potential in devices that combine nanochannels with nanpore detectors. In particular, due to the fast translocation of a DNA molecule through a standard nanopore configuration, there is an unfavorable trade-off between signal and sequence resolution. With a combined nanochannel-nanopore device, based on embedding a nanopore inside a nanochannel, we can in principle gain independent control over both DNA translocation speed and sensing signal, solving the key draw-back of the standard nanopore configuration. We demonstrate that we can detect - using fluorescent microscopy - successful translocation of DNA from the nanochannel out through the nanopore, a possible method to 'select' a given barcode for further analysis. We also show that in equilibrium DNA will not escape through an embedded sub-persistence length nanopore until a certain voltage bias is added. [Preview Abstract] |
Thursday, March 6, 2014 1:39PM - 1:51PM |
T10.00011: Pre-stretching a Polymer to Reduce the Variance on Mean Translocation Times David Sean, Hendrick de Haan, Gary Slater Recent theoretical developments in driven polymer translocation highlight the importance of the polymer conformation before translocation. The rate of the propagation of tension arising from the application of a driving force is highly dependent upon the initial position of the monomers due to the separation of time scales between the polymer relaxation time and the translocation time. In this high P\'eclet number limit, we use Langevin Dynamics computer simulations and Tension-Propagation theory to investigate how pre-stretching the polymer controls translocation time distributions. Motivated by the influence of monomer crowding on the trans-side, we explore the contrast between applying a driving force inside the pore and applying a pulling force on the polymer end. [Preview Abstract] |
Thursday, March 6, 2014 1:51PM - 2:03PM |
T10.00012: Optical Nanofluidic Piston: Assay for Dynamic Force-Compression of Single Confined Polymer Chains Ahmed Khorshid, Philip Zimny, Patrick Macos, Geremia Massarelli, David T\'etreault-La Roche, Walter Reisner While single-molecule approaches now have a long-history in polymer physics, past methodology has a key limitation \textit{: it is not currently possible to apply well-defined forces to a precise number of chains in a well-defined volume. To this end, }we have developed a nanofluidic assay for the study of DNA compression in vitro, the \textit{optical nanofluidic piston.} The optical nanofluidic piston is a nanofluidic analog of a macroscopic piston-cylinder apparatus based on a nanosphere (``the piston'') optically trapped inside a 200-400nm nanochannel with embedded barrier (the ``cylinder''). The nanofluidic piston enables quantification of force required to compress single or multiple chains within a defined volume. We present combined fluorescence and force-measurements for the compression of T4 DNA under a variety of compression rates. Surprisingly, we find that compression occurs on a force-scale roughly 100x higher than that predicted by equilibrium theories, suggesting that the DNA is present in highly entangled states during the compression. Moreover, we observe that compression at high rates induces a ``shock-wave'' of high-polymer concentration near the bead, suggesting that our setup can quantitatively access novel non-equilibrium polymer phenomena. [Preview Abstract] |
Thursday, March 6, 2014 2:03PM - 2:15PM |
T10.00013: Nano-funnels as electro-osmotic ``tweezers and pistons'' Yanqian Wang, Sergey Panyukov, Jinsheng Zhou, Laurent D. Menard, J. Michael Ramsey, Michael Rubinstien An electric field is used to force a DNA molecule into a nano-channel by compensating the free energy penalty that results from the reduced conformational entropy of the confined macromolecule. Narrow nano-channels require high critical electric fields to achieve DNA translocation, leading to short dwell times of DNA in these channels. We demonstrate that nano-funnels integrated with nano-channels reduce the free energy barrier and lower the critical electric field required for DNA translocation. A focused electric field within the funnel increases the electric force on the DNA, compresses the molecule, and increases the osmotic pressure at the nano-channel entrance. This ``electro-osmotic piston'' forces the molecule into the nano-channel at lower electric fields than those observed without the funnel. Appropirately designed nano-funnels can also function as tweezers that allow manipulation of the position of the DNA molecule. The predictions of our theory describing double-stranded DNA behavior in nano-funnel -- nano-channel devices are consistent with experimental results. [Preview Abstract] |
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