DNA in nanochannels: theory and applications

Abstract

Nanofluidic structures have over the last two decades emerged as a powerful platform for detailed analysis of DNA on the kilobase pair length scale. When DNA is confined to a nanochannel, the combination of excluded volume and DNA stiffness leads to the DNA being stretched to near its full contour length. Importantly, this stretching takes place at equilibrium, without any chemical modifications to the DNA. As a result, any DNA can be analyzed, such as DNA extracted from cells or circular DNA, and it is straight-forward to study reactions on the ends of linear DNA. In this comprehensive review, we first give a thorough description of the current understanding of the polymer physics of DNA and how that leads to stretching in nanochannels. We then describe how the versatility of nanofabrication can be used to design devices specifically tailored for the problem at hand, either by controlling the degree of confinement or enabling facile exchange of reagents to measure DNA-protein reaction kinetics. The remainder of the review focuses on two important applications of confining DNA in nanochannels. The first is optical DNA mapping, which provides the genomic sequence of intact DNA molecules in excess of 100 kilobase pairs in size, with kilobase pair resolution, through labeling strategies that are suitable for fluorescence microscopy. In this section, we highlight solutions to the technical aspects of genomic mapping, including the use of enzyme-based labeling and affinitybased labeling to produce the genomic maps, rather than recent applications in human genetics. The second is DNA-protein interactions, and several recent examples of such studies on DNA compaction, filamentous protein complexes, and reactions with DNA ends are presented. Taken together, these two applications demonstrate the power of DNA confinement and nanofluidics in genomics, molecular biology, and biophysics.

Strain-level bacterial typing directly from patient samples using optical DNA mapping

“…We have developed an ODM assay based on the non-covalent competitive binding of the fluorescent YOYO-1 and the AT-specific, non-fluorescent netropsin to DNA that results in a sequence-specific emission intensity variation along the DNA molecule 10 . The emission intensity variation can be visualized using fluorescence microscopy whilst the DNA is stretched in nanofluidic channels 11 . Since the method relies solely on the interaction of YOYO-1 and netropsin with DNA, it is agnostic to the DNA in the sample, which means that any bacterium can be investigated without, for example, designing primers.…”

Polymer Brushes on Silica Nanostructures Prepared by Aminopropylsilatrane Click Chemistry: Superior Antifouling and Biofunctionality

“…To extend the range of applications for the APS-SMCC-PEG modification beyond planar surfaces, we applied the method to fluidic chips with micro- and nanosized channels in SiO 2 . Such chips are established for several bioanalytical purposes, in particular electrostatic trapping of biomolecules and stretching of DNA. The use of nanofluidic devices to study DNA-protein interactions at the single-molecule level has gained increasing interest and passivation of the channel surfaces is crucial to minimize adsorption of proteins .…”

“…Typically, there is a need to suppress interactions with the surface since the molecules to be studied or detected will be (or should be) in the solution phase. This is the case in, for example, electrostatic traps, 6 nanochannels for DNA analysis, 7 convex lens-induced confinement, 8 and solid-state nanopore sensors. 9 Notably, all these devices have oxidized silicon as the material forming the interface to the biological solution and preventing biomolecule adsorption is key to successful operation in all cases.…”

Probing physical properties of single amyloid fibrils using nanofluidic channels

“…The nanofluidic devices used in this work were fabricated using traditional semiconductor processing methods that are described in detail elsewhere. 22,35 Briefly, the design consists of 80 nanochannels, that run between two microchannels, each having two loading reservoirs. The nanochannels are 500 µm long and 300 nm deep.…”