DNA Origami as a Platform for Hybrid Nanostructures

By 2022, DNA origami had matured into a versatile three-dimensional construction platform with sub-nanometre positional precision. At Leipzig, Seidel and co-workers had pioneered DNA origami molds for shape-controlled metallic nanoparticle synthesis — casting gold nanorods inside tubular cavities (Helmi et al., 2014) and developing a modular construction kit for complex metal nanostructures. The platform had been extended to palladium nanostructures (Ye et al., 2021). DNA origami nanopores had been demonstrated in solid-state platforms and lipid bilayers. However, the integration of thermal control with these nanostructures — to reversibly gate molecular access or dynamically reconfigure sensing elements — remained unrealised.

DNA Hybridization Kinetics

Equilibrium thermodynamics of DNA duplex formation were well described by the nearest-neighbour model (SantaLucia, 1998), but kinetic parameters — association and dissociation rates — were poorly characterised. Single-molecule techniques (smFRET, DNA-PAINT) provided dwell-time information for specific systems, but systematic temperature-dependent measurements across diverse sequences were absent. Whether association rates increased, decreased, or remained constant with temperature was actively debated. No high-throughput platform existed to measure sequence-specific kon and koff across temperatures simultaneously.

Strand Displacement Dynamics

Toehold-mediated strand displacement (Zhang & Winfree, 2009) had become the standard mechanism for dynamic DNA nanotechnology. While displacement kinetics had been characterised in bulk at room temperature (Srinivas et al., 2013; Irmisch et al., 2020), the temperature dependence of displacement rate constants and the activation barriers governing strand invasion and branch migration were not systematically characterised. This gap was critical for designing thermally switchable DNA devices that must operate across a range of temperatures.

Plasmonic Sensing and SERS

Surface-enhanced Raman scattering (SERS) in plasmonic nanostructures had achieved single-molecule sensitivity. DNA origami provided scaffolds for assembling nanoparticle dimers with defined gap sizes (~2–10 nm), creating electromagnetic hotspots with enhancement factors exceeding 108. However, systematic methods for reproducible analyte positioning and orientation control within hotspots were lacking. The DNA origami mould technology offered a promising route for casting metal nanostructures with programmable gap geometries, but quantitative integration with thermally switchable DNA elements had not been demonstrated.

Temperature Measurement at the Nanoscale

Fluorescence thermometry using Rhodamine B and other temperature-sensitive dyes had been demonstrated in microfluidic contexts, but integration with TIRF microscopy for surface-temperature calibration during single-molecule experiments was not established. Sub-diffraction temperature mapping — essential for characterising thermal gradients around plasmonic heaters — remained an unmet challenge.

Selected Output

• N. Kumar, F. Ricci, R. Seidel, P. Irmisch. "Engineering the temperature response of DNA strand displacement." Nature Communications [Under Review]
• N. Kumar, A. Overchenko, A. Sivaraman, C. Bastiaanssen, P. Irmisch, F. Cichos, R. Seidel, C. Joo. "Temperature- and sequence-dependent DNA hybridization kinetics from high-throughput single-molecule measurements." [In Preparation — Nature Chemistry]
• N. Kumar et al. "Probing sequence-specific DNA hybridization dynamics using ThermoSPARXS." Biophysical Journal, 125(4), 287a (2026).
• S. Banerjee, C. Hadlich, M. Scherf, N. Kumar, R. Seidel, J. Kneipp. "SERS detects site-specific labeling of DNA nanocages via hexynyl modifications." [In Preparation]
• N. Kumar, A. Sivaraman, P. Irmisch, D. Renger, C. Joo, R. Seidel. "DNA Origami-Enabled Single-Molecule Ratiometric Thermometry with Nanoscale Spatial Resolution." [In Preparation]