| 1. |
- Alizadehheidari, Mohammadreza, 1987, et al.
(författare)
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Nanoconfined Circular and Linear DNA: Equilibrium Conformations and Unfolding Kinetics
- 2015
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Ingår i: Macromolecules. - : American Chemical Society (ACS). - 0024-9297 .- 1520-5835. ; 48:3, s. 871-878
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Tidskriftsartikel (refereegranskat)abstract
- Studies of circular DNA confined to nanofluidic channels are relevant both from a fundamental polymer-physics perspective and due to the importance of circular DNA molecules in vivo. We here observe the unfolding of confined DNA from the circular to linear configuration as a light-induced double-strand break occurs, characterize the dynamics, and compare the equilibrium conformational statistics of linear and circular configurations. This is important because it allows us to determine to what extent existing statistical theories describe the extension of confined circular DNA. We find that the ratio of the extensions of confined linear and circular DNA configurations increases as the buffer concentration decreases. The experimental results fall between theoretical predictions for the extended de Gennes regime at weaker confinement and the Odijk regime at stronger confinement. We show that it is possible to directly distinguish between circular and linear DNA molecules by measuring the emission intensity from the DNA. Finally, we determine the rate of unfolding and show that this rate is larger for more confined DNA, possibly reflecting the corresponding larger difference in entropy between the circular and linear configurations.
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| 2. |
- Noble, Charleston, et al.
(författare)
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A fast and scalable kymograph alignment algorithm for nanochannel-based optical DNA mappings.
- 2015
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Ingår i: PLoS ONE. - : Public Library of Science (PLoS). - 1932-6203. ; 10:4
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Tidskriftsartikel (refereegranskat)abstract
- Optical mapping by direct visualization of individual DNA molecules, stretched in nanochannels with sequence-specific fluorescent labeling, represents a promising tool for disease diagnostics and genomics. An important challenge for this technique is thermal motion of the DNA as it undergoes imaging; this blurs fluorescent patterns along the DNA and results in information loss. Correcting for this effect (a process referred to as kymograph alignment) is a common preprocessing step in nanochannel-based optical mapping workflows, and we present here a highly efficient algorithm to accomplish this via pattern recognition. We compare our method with the one previous approach, and we find that our method is orders of magnitude faster while producing data of similar quality. We demonstrate proof of principle of our approach on experimental data consisting of melt mapped bacteriophage DNA.
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| 3. |
- Kesarimangalam, Sriram, 1983, et al.
(författare)
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Fluorescence Microscopy of Nanochannel-Confined DNA
- 2024
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Ingår i: Methods in Molecular Biology. - 1940-6029 .- 1064-3745. - 9781071633779 - 9781071633762 ; , s. 175-202
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Bokkapitel (övrigt vetenskapligt/konstnärligt)abstract
- Stretching of DNA in nanoscale confinement allows for several important studies. The genetic contents of the DNA can be visualized on the single DNA molecule level, and the polymer physics of confined DNA and also DNA/protein and other DNA/DNA-binding molecule interactions can be explored. This chapter describes the basic steps to fabricate the nanostructures, perform the experiments, and analyze the data.
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| 4. |
- Nilsson, Adam, et al.
(författare)
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Competitive binding-based optical DNA mapping for fast identification of bacteria - multi-ligand transfer matrix theory and experimental applications on Escherichia coli.
- 2014
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Ingår i: Nucleic Acids Research. - : Oxford University Press (OUP). - 1362-4962 .- 0305-1048. ; 42:15, s. 118-118
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Tidskriftsartikel (refereegranskat)abstract
- We demonstrate a single DNA molecule optical mapping assay able to resolve a specific Escherichia coli strain from other strains. The assay is based on competitive binding of the fluorescent dye YOYO-1 and the AT-specific antibiotic netropsin. The optical map is visualized by stretching the DNA molecules in nanofluidic channels. We optimize the experimental conditions to obtain reproducible barcodes containing as much information as possible. We implement a multi-ligand transfer matrix method for calculating theoretical barcodes from known DNA sequences. Our method extends previous theoretical approaches for competitive binding of two types of ligands to many types of ligands and introduces a recursive approach that allows long barcodes to be calculated with standard computer floating point formats. The identification of a specific E. coli strain (CCUG 10979) is based on mapping of 50-160 kilobasepair experimental DNA fragments onto the theoretical genome using the developed theory. Our identification protocol introduces two theoretical constructs: a P-value for a best experiment-theory match and an information score threshold. The developed methods provide a novel optical mapping toolbox for identification of bacterial species and strains. The protocol does not require cultivation of bacteria or DNA amplification, which allows for ultra-fast identification of bacterial pathogens.
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| 5. |
- Beech, Jason, et al.
(författare)
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Sorting cells by size, shape and deformability
- 2012
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Ingår i: Lab on a Chip. - : Royal Society of Chemistry (RSC). - 1473-0197 .- 1473-0189. ; 12, s. 1048-1051
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Tidskriftsartikel (refereegranskat)abstract
- While size has been widely used as a parameter in cellular separations, in this communication we show how shape and deformability, a mainly untapped source of specificity in preparative and analytical microfluidic devices can be measured and used to separate cells. © 2012 The Royal Society of Chemistry.
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| 6. |
- Fritzsche, Michael, et al.
(författare)
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A Highly UV-transparent Fused Silica Biochip for Sensitive Hepatotoxicity Testing by Autofluorescence
- 2014
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Ingår i: Biochip Journal. - : Springer Science and Business Media LLC. - 2092-7843 .- 1976-0280. ; 8:2, s. 115-121
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Tidskriftsartikel (refereegranskat)abstract
- Fabrication and application of a non-fluorescent and UV-transparent microfluidic biochip in fused silica that allows sensitive autofluorescence detection are described. The biochip is particularly useful in cell-based assays where the most informative autofluorescence signals from the cells reside in the ultraviolet spectral range and where plastic labware materials commonly used in cell culture work severely disturb such measurements. In this study the fused silica biochip was used for measuring intrinsic autofluorescence from liver cells in order to assess hepatotoxic effects of drugs. The assessment assay was carried out with the human liver cell line HepG2 under perfusion conditions in the microfluidics of the biochip. The autofluorescence from the.liver cells exposed to quinidine was readily recorded without background disturbance and correlated well with reference toxicity data.
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| 7. |
- Ling, TGI, et al.
(författare)
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Fa-brication and characterization of a molecular adhesive layer for micro- and nanofabricated electrochemical electrodes
- 2003
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Ingår i: Microelectronic Engineering. - 1873-5568. ; 67-8, s. 887-892
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Tidskriftsartikel (refereegranskat)abstract
- In the microelectronics industry, gold electrodes on silicon dioxide are produced using an adhesive layer of chromium or titanium. This method is applicable whenever a dry environment is present. However, when such an electrode is exposed to an electrolyte a galvanic cell is formed and the less noble metal will dissolve and eventually destroy the electrode. We describe here how a monolayer of mercaptopropyl-trimethoxysilane (MPTS) can be used as an adhesive. The layer can be structured lithographically making gold electrodes in the micro- and nano-range applicable for use in electrochemistry. The properties of the MPTS layer were investigated with photoelectron microscopy, atomic force microscopy and the electrochemical performance of the gold electrodes was determined by cyclic voltammetry. (C) 2003 Elsevier Science B.V. All rights reserved.
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| 8. |
- Persson, Henrik, et al.
(författare)
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Vertical oxide nanotubes connected by subsurface microchannels
- 2012
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Ingår i: Nano Reseach. - : Springer Science and Business Media LLC. - 1998-0124 .- 1998-0000. ; 5:3, s. 190-198
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Tidskriftsartikel (refereegranskat)abstract
- We describe the fabrication of arrays of oxide nanotubes using a combination of bottom up and top down nanofabrication. The nanotubes are made from epitaxially grown semiconductor nanowires that are covered with an oxide layer using atomic layer deposition. The tips of the oxide-covered nanowires are removed by argon sputtering and the exposed semiconductor core is then selectively etched, leaving a hollow oxide tube. We show that it is possible to create fluidic connections to the nanotubes by a combination of electron beam lithography to precisely define the nanotube positions and controlled wet under-etching. DNA transport is demonstrated in the microchannel. Cells were successfully cultured on the nanotube arrays, demonstrating compatibility with cell-biological applications. Our device opens up the possibility of injecting molecules into cells with both spatial and temporal control.
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| 9. |
- Werner, Erik, et al.
(författare)
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Monomer distributions and intrachain collisions of a polymer confined to a channel
- 2013
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Ingår i: Macromolecules. - : American Chemical Society (ACS). - 0024-9297 .- 1520-5835. ; 46:16, s. 6644-6650
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Tidskriftsartikel (refereegranskat)abstract
- We study the conformations of a self-avoiding polymer confined to a channel by computing the cross-sectional distributions of the positions of its monomers. By means of Monte Carlo simulations for a self-avoiding, freely jointed chain, we determine how the cross-sectional distribution for a given monomer depends on its location in the polymer and how strongly this distribution is affected by self-avoidance. To this end we analyze how the frequency of intrachain collisions between monomers depends on their spatial position in the channel and on their location within the polymer. We show that most collisions occur between closely neighboring monomers. As a consequence, the collision probability depends only weakly on the spatial position of the monomers. Our results explain why the effect of self-avoidance on the monomer distributions is weaker than predicted by mean-field theory. We discuss the relevance of our results for studies of DNA conformations in nanofluidic channels.
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| 10. |
- Werner, Erik, et al.
(författare)
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Orientational correlations in confined DNA
- 2012
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Ingår i: Physical Review E. - 1539-3755 .- 2470-0045 .- 2470-0053. ; 86:4
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Tidskriftsartikel (refereegranskat)abstract
- We study how the orientational correlations of DNA confined to nanochannels depend on the channel diameter D by means of Monte Carlo simulations and a mean-field theory. This theory describes DNA conformations in the experimentally relevant regime where Flory-de Gennes theory does not apply. We show how local correlations determine the dependence of the end-to-end distance of the DNA molecule upon D. Tapered nanochannels provide the necessary resolution in D to study experimentally how the extension of confined DNA molecules depends upon D. Our experimental and theoretical results are in qualitative agreement.
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