Polymer support for exonucleolytic sequencing

Hinz, Michael; Gura, Sigalit; Nitzan, B.; Margel, Shlomo; Seliger, H.

doi:10.1016/s0168-1656(00)00419-3

Cited by 12 publications

(6 citation statements)

References 8 publications

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“…New and developing technologies for single molecule manipulation and analysis of DNA in micron and nanometer scale devices [1][2][3][4][5][6] have fueled considerable interest in the structure and dynamics of solutions of DNA in confined geometries. Predictive methods capable of describing the conformation and motion of polymer chains in micro-and nanofluidic geometries would be of considerable significance for the conception and design of such devices.…”

Section: Introductionmentioning

confidence: 99%

Shear-induced migration in flowing polymer solutions: Simulation of long-chain DNA in microchannels

Jendrejack

Schwartz

Pablo

et al. 2004

The Journal of Chemical Physics

232

209

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We simulate dilute solution dynamics of long flexible polymer molecules in pressure driven flow in channels with widths of roughly 0.1-10 times the polymer bulk radius of gyration. This is done using a self-consistent coarse-grained Langevin description of the polymer dynamics and a numerical simulation of the flow in the confined geometry that is generated by the motions of polymer segments. Results are presented for a model of DNA molecules of ϳ10-100 m contour length in micron-scale channels. During flow, the chains migrate toward the channel centerline, in agreement with well-known experimental observations. The thickness of the resulting hydrodynamic depletion layer increases with molecular weight at constant flow strength; higher molecular weight chains therefore move with a higher average axial velocity than lower molecular weight chains. In contrast, if the hydrodynamic effects of the confining geometry are neglected, depletion of concentration is observed in the center of the channel rather than at the walls, contradicting experimental observations. The mechanisms for migration are illustrated using a simple kinetic theory dumbbell model of a confined flexible polymer. The simple theory correctly predicts the trends observed in the detailed simulations. We also examine the steady-state stretch of DNA chains as a function of channel width and flow strength. The flow strength needed to stretch a highly confined chain away from its equilibrium length is shown to increase with decreasing channel width, independent of molecular weight; this is fairly well explained using a simple blob picture.

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Section: Introductionmentioning

confidence: 99%

Shear-induced migration in flowing polymer solutions: Simulation of long-chain DNA in microchannels

Jendrejack

Schwartz

Pablo

et al. 2004

The Journal of Chemical Physics

232

209

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“…To clarify inconsistent reports, we investigated the immobilization process in nL and μL drops, as well as in liquid solution. The latter plays an important role in many current technologies and applications, including the immobilization of oligonucleotides on nanoparticles [38-40], microspheres [41], magnetic microparticles [42], polymer [43,44] and glass beads [45] for microarraying [42] and flow cytometry [46], surface plasmon resonance [47], and next generation sequencing (Roche 454, ABI SOLiD, Ion Torrent and Illumina).…”

Section: Introductionmentioning

confidence: 99%

Drop drying on surfaces determines chemical reactivity - the specific case of immobilization of oligonucleotides on microarrays

et al. 2013

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BackgroundDrop drying is a key factor in a wide range of technical applications, including spotted microarrays. The applied nL liquid volume provides specific reaction conditions for the immobilization of probe molecules to a chemically modified surface.ResultsWe investigated the influence of nL and μL liquid drop volumes on the process of probe immobilization and compare the results obtained to the situation in liquid solution. In our data, we observe a strong relationship between drop drying effects on immobilization and surface chemistry. In this work, we present results on the immobilization of dye labeled 20mer oligonucleotides with and without an activating 5′-aminoheptyl linker onto a 2D epoxysilane and a 3D NHS activated hydrogel surface.ConclusionsOur experiments identified two basic processes determining immobilization. First, the rate of drop drying that depends on the drop volume and the ambient relative humidity. Oligonucleotides in a dried spot react unspecifically with the surface and long reaction times are needed. 3D hydrogel surfaces allow for immobilization in a liquid environment under diffusive conditions. Here, oligonucleotide immobilization is much faster and a specific reaction with the reactive linker group is observed. Second, the effect of increasing probe concentration as a result of drop drying. On a 3D hydrogel, the increasing concentration of probe molecules in nL spotting volumes accelerates immobilization dramatically. In case of μL volumes, immobilization depends on whether the drop is allowed to dry completely. At non-drying conditions, very limited immobilization is observed due to the low oligonucleotide concentration used in microarray spotting solutions. The results of our study provide a general guideline for microarray assay development. They allow for the initial definition and further optimization of reaction conditions for the immobilization of oligonucleotides and other probe molecule classes to different surfaces in dependence of the applied spotting and reaction volume.

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“…[1][2][3][4][5][6][7] For instance, in certain implementations of exonucleolytic sequence analysis it is desirable to link an individual long DNA strand to a surface ͑e.g., a bead͒ without manual intervention. 6,7 The ability to engineer such tasks will be greatly improved by predictive computational tools for polymer chains in microfluidic geometries. In this work, we present a general method for dynamic simulations of macromolecules in confined geometries.…”

Section: Introductionmentioning

confidence: 99%

Effect of confinement on DNA dynamics in microfluidic devices

Jendrejack

Schwartz

Graham

et al. 2003

The Journal of Chemical Physics

166

193

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The dynamics of dissolved long-chain macromolecules are different in highly confined environments than in bulk solution. A computational method is presented here for detailed prediction of these dynamics, and applied to the behavior of ϳ1-100 m DNA in micron-scale channels. The method is comprised of a self-consistent coarse-grained Langevin description of the polymer dynamics and a numerical solution of the flow generated by the motion of polymer segments. Diffusivity and longest relaxation time show a broad crossover from free-solution to confined behavior centered about the point HϷ10S b , where H is the channel width and S b is the free-solution chain radius of gyration. In large channels, the diffusivity is similar to that of a sphere diffusing along the centerline of a pore. For highly confined chains (H/S b Ӷ1), Rouse-type molecular weight scaling is observed for both translational diffusivity and longest relaxation time. In the highly confined region, the scaling of equilibrium length and relaxation time with H/S b are in good agreement with scaling theories. In agreement with the results of Harden and Doi ͓J. Phys. Chem. 96, 4046 ͑1992͔͒, we find that the diffusivity of highly confined chains does not follow the scaling relation predicted by Brochard and de Gennes ͓J. Chem. Phys. 67, 52 ͑1977͔͒; that relationship does not account for the interaction between chain and wall.

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Polymer support for exonucleolytic sequencing

Cited by 12 publications

References 8 publications

Shear-induced migration in flowing polymer solutions: Simulation of long-chain DNA in microchannels

Shear-induced migration in flowing polymer solutions: Simulation of long-chain DNA in microchannels

Drop drying on surfaces determines chemical reactivity - the specific case of immobilization of oligonucleotides on microarrays

Effect of confinement on DNA dynamics in microfluidic devices

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