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Project 11

Thermal switching and operation of hybrid DNA-origami plasmonic structures and nanopores

ESR: Aleksei Overchenko

State of the Art prior to DYNAMO (2022)

Solid-state and DNA-origami hybrid nanopores

Before the start of the DYNAMO project, nanopore technologies and DNA nanotechnology had already demonstrated strong potential for single-molecule sensing and sequencing applications[cite: 14]. Solid-state nanopores were widely investigated because of their stability, tunable geometry, and compatibility with electrical detection techniques [1,2][cite: 14]. At the same time, DNA origami emerged as a powerful platform for constructing programmable nanoscale architectures with controllable geometries and functionalities [3,4][cite: 14]. Early studies demonstrated that DNA-origami nanopores could be integrated with solid-state membranes to create hybrid systems with improved sensitivity and controllable molecular transport [5,6][cite: 14]. Bell et al. (2012) first showed that DNA-origami structures could be docked into and ejected from solid-state nanopores under applied voltage, enabling the resistive-pulse detection of λ-DNA through hybrid pores [5][cite: 14]. Subsequent work extended the concept to graphene membranes and to programmable DNA nanoswitches read out by ionic current [6,7][cite: 14]. However, recurring limitations were reported, including residual leakage currents through the DNA scaffold, elevated and variable ionic noise compared with bare solid-state pores, and only partial control over the orientation and stability of the inserted origami [2][cite: 14].

Electroosmotic flow in nanopores and nanocapillaries

Electroosmotic flow (EOF) had also already been identified as an important mechanism governing transport inside nanopores and nanocapillaries [8,9][cite: 14]. Previous studies showed that EOF could influence translocation dynamics, trapping efficiency, and ionic transport near charged surfaces [9,10][cite: 14]. Asandei et al. (2016) demonstrated that EOF could be used as an electroosmotic trap against the electrophoretic force to capture peptides at a protein nanopore [10][cite: 14], and similar trapping concepts were extended to whole proteins inside engineered biological pores [11][cite: 14]. In glass nanocapillaries, Laohakunakorn and Keyser established that EOF profiles depend strongly on pore geometry and ionic strength, with flow reversal observed outside conical pores when the salt concentration is lowered [9,12][cite: 14]. However, most experimental investigations were performed under relatively limited conditions, often focusing on a single salt type (typically KCl), narrow concentration ranges, or comparatively large capillary geometries with limited spatial resolution of the flow field[cite: 14]. As a result, the detailed dependence of EOF on ion identity, concentration, and nanoscale confinement remained insufficiently understood, and three-dimensional mapping of the flow field outside a nanopore had only just been demonstrated as a proof of concept [13][cite: 14].

Thermally responsive DNA nanostructures and hybridization physics

Thermally responsive DNA nanostructures and reversible DNA melting had already demonstrated the possibility of controllable structural switching at the nanoscale [14,15][cite: 14]. DNA origami nanocapsules and lattices reconfigurable by pH, temperature, or strand displacement had been reported, often relying on toehold-mediated strand exchange or triplex-forming pH latches [14,15,16][cite: 14]. Nevertheless, most earlier studies primarily focused on proof-of-concept demonstrations of structural reconfiguration, while the underlying physics of DNA hybridization remained only partially understood, especially under confined nanopore conditions and at the single-molecule level[cite: 14]. Theoretical and coarse-grained simulation work had outlined a picture of hybridization proceeding through non-specific contact, stochastic nucleation, and zipping [17,18][cite: 14], but direct experimental access to these elementary steps for short, surface-tethered strands embedded in nanofluidic environments was still largely missing[cite: 14].

References

  • [1] Dekker, C. Solid-state nanopores. Nat. Nanotechnol. 2, 209–215 (2007)[cite: 14].
  • [2] Hernández-Ainsa, S. & Keyser, U. F. DNA-origami nanopores: developments, challenges and perspectives. Nanoscale 6, 14121–14132 (2014)[cite: 14].
  • [3] Rothemund, P. W. K. Folding DNA to create nanoscale shapes and patterns. Nature 440, 297–302 (2006)[cite: 14].
  • [4] Dietz, H., Douglas, S. M. & Shih, W. M. Folding DNA into twisted and curved nanoscale shapes. Science 325, 725–730 (2009)[cite: 14].
  • [5] Bell, N. A. W. et al. DNA origami nanopores. Nano Lett. 12, 512–517 (2012)[cite: 14].
  • [6] Barati Farimani, A. et al. DNA origami–graphene hybrid nanopore for DNA detection. ACS Appl. Mater. Interfaces 9, 92–100 (2017)[cite: 14].
  • [7] Bell, N. A. W. & Keyser, U. F. Specific protein detection using designed DNA carriers and nanopores. J. Am. Chem. Soc. 137, 2035–2041 (2015)[cite: 14].
  • [8] Firnkes, M. et al. Electrically facilitated translocations of proteins through silicon nitride nanopores: conjoint and competitive action of diffusion, electrophoresis, and electroosmosis. Nano Lett. 10, 2162–2167 (2010)[cite: 14].
  • [9] Laohakunakorn, N. et al. Electroosmotic flow reversal outside glass nanopores. Nano Lett. 15, 695–702 (2015)[cite: 14].
  • [10] Asandei, A. et al. Electroosmotic trap against the electrophoretic force near a protein nanopore reveals peptide dynamics during capture and translocation. ACS Appl. Mater. Interfaces 8, 13166–13179 (2016)[cite: 14].
  • [11] Schmid, S. et al. Nanopore electro-osmotic trap for the label-free study of single proteins and their conformations. Nat. Nanotechnol. 16, 1244–1250 (2021)[cite: 14].
  • [12] Mc Hugh, J., Andresen, K. & Keyser, U. F. Cation-dependent electroosmotic flow in glass nanopores. Appl. Phys. Lett. 115, 113702 (2019)[cite: 14].
  • [13] Mc Hugh, J., Thorneywork, A. L., Andresen, K. & Keyser, U. F. 3D flow field measurements outside nanopores. Rev. Sci. Instrum. 93, 053703 (2022)[cite: 14].
  • [14] Ijäs, H. et al. Reconfigurable DNA origami nanocapsule for pH-controlled encapsulation and display of cargo. ACS Nano 13, 5959–5967 (2019)[cite: 14].
  • [15] Kosuri, P., Altheimer, B. D., Dai, M., Yin, P. & Zhuang, X. Rotation tracking of genome-processing enzymes using DNA origami rotors. Nature 572, 136–140 (2019)[cite: 14].
  • [16] Marras, A. E., Zhou, L., Su, H.-J. & Castro, C. E. Programmable motion of DNA origami mechanisms. Proc. Natl. Acad. Sci. USA 112, 713–718 (2015)[cite: 14].
  • [17] Ouldridge, T. E. et al. DNA hybridization kinetics: zippering, internal displacement and sequence dependence. Nucleic Acids Res. 41, 8886–8895 (2013)[cite: 14].
  • [18] SantaLucia, J. & Hicks, D. The thermodynamics of DNA structural motifs. Annu. Rev. Biophys. Biomol. Struct. 33, 415–440 (2004)[cite: 14].

Current State of the Art within DYNAMO (2026)

Coupled nanoscale transport and improved EOF resolution

Since the beginning of DYNAMO, the field has progressed toward a deeper understanding of coupled nanoscale transport phenomena[cite: 13]. Recent work has focused on improving the spatial resolution of electroosmotic flow measurements and investigating EOF behavior under different ionic environments and salt concentrations [13,19][cite: 13]. Three-dimensional, optical-tweezer-based flow-field mapping outside glass nanopores has been refined to capture the full out-of-plane structure of voltage-driven flow, revealing polarity-dependent flow fields consistent with competing inner- and outer-wall contributions [13][cite: 13]. Joint experimental and computational studies have further dissected the role of buffer salt concentration and catton identity in setting the magnitude and sign of EOF inside both biological and solid-state pores, showing strongly non-monotonic dependencies that classical Poisson–Nernst–Planck descriptions only partly capture [19,20][cite: 13]. These studies aim to better understand how ion-specific effects and nanoscale confinement influence flow dynamics at solid–liquid interfaces[cite: 13]. At the same time, the use of dark-field optical imaging and high-speed scattering microscopy has enabled more detailed investigation of transport processes with higher spatial and temporal resolution, including single-particle tracking near the pore mouth and inside nanocapillaries [21,22][cite: 13].

Single-molecule DNA hybridization and reconfigurable DNA-origami nanopores

In parallel, recent studies have increasingly focused on the physical mechanisms governing DNA hybridization and melting dynamics rather than only demonstrating structural switching [17,23][cite: 13]. In particular, nucleation and zipping models describing DNA binding and melting are now being investigated experimentally at the single-molecule level through kinetic measurements [23,24][cite: 13]. Hertel et al. (2022) quantified how the stability and number of nucleating interactions determine hybridization rates in the absence of secondary structure [23][cite: 13], while single-molecule fluorescence and DNA-PAINT approaches have provided base-pair-resolved access to association and dissociation kinetics [24][cite: 13]. On the DNA-origami side, fully reversible, mechanically compliant origami nanopores (MechanoPores) that switch between closed, intermediate, and open states by strand displacement have recently been demonstrated and inserted into lipid membranes, achieving size-selective transmembrane transport [25][cite: 13]. These studies provide deeper insight into reversible DNA-origami reconfiguration and may become important for future adaptive nanopore systems with controllable trapping and molecular-transport capabilities[cite: 13].

Outlook and remaining challenges

Current research focuses on understanding the fundamental physics governing nanoscale transport, reversible DNA reconfiguration, and electrohydrodynamic interactions under confined conditions[cite: 13]. Overall, the combination of DNA origami, electroosmotic flow, thermally responsive nanosystems, and single-molecule transport studies represents a promising direction toward adaptive nanopore sequencing technologies[cite: 13]. Coupling EOF and DNA-origami inserts in a single device introduces additional complexity, since the deformability and surface charge of the origami modify the local flow field and ionic environment in ways that are not yet fully predictable [2,25][cite: 13]. Substantial scientific and technological challenges therefore remain — in particular regarding low-noise integration, long-term stability of origami inserts in solid-state membranes, and quantitative modelling of coupled electrohydrodynamic and thermal effects — before fully integrated and scalable sequencing devices can be realized[cite: 13].

Selected Output

  • [13] Mc Hugh, J., Thorneywork, A. L., Andresen, K. & Keyser, U. F. 3D flow field measurements outside nanopores. Rev. Sci. Instrum. 93, 053703 (2022)[cite: 13].
  • [17] Ouldridge, T. E. et al. DNA hybridization kinetics: zippering, internal displacement and sequence dependence. Nucleic Acids Res. 41, 8886–8895 (2013)[cite: 13].
  • [19] Bhattacharya, S. et al. Changes in salt concentration modify the translocation of neutral molecules through a ΔCymA nanopore in a non-monotonic manner. ACS Nano 16, 9355–9366 (2022)[cite: 13].
  • [20] Brown, W. et al. Higher ion selectivity with lower energy usage promoted by electro-osmotic flow in the transport through conical nanopores. J. Phys. Chem. C 125, 3269–3276 (2021)[cite: 13].
  • [21] Teahan, J. et al. Scanning ion conductance microscopy: surface charge effects on electroosmotic flow delivery from a nanopipette. Anal. Chem. 93, 12281–12288 (2021)[cite: 13].
  • [22] Verpillat, F., Joud, F., Desbiolles, P. & Gross, M. Dark-field digital holographic microscopy for 3D-tracking of gold nanoparticles. Opt. Express 19, 26044–26055 (2011)[cite: 13].
  • [23] Hertel, S. et al. The stability and number of nucleating interactions determine DNA hybridization rates in the absence of secondary structure. Nucleic Acids Res. 50, 7829–7841 (2022)[cite: 13].
  • [24] Jungmann, R. et al. Multiplexed 3D cellular super-resolution imaging with DNA-PAINT and Exchange-PAINT. Nat. Methods 11, 313–318 (2014)[cite: 13].
  • [25] Yu, L. et al. Compliant DNA origami nanoactuators as size-selective nanopores. Adv. Mater. 36, 2405104 (2024)[cite: 13].
State of the art