Single-molecule DNA sequencing based on measuring the physical properties of bases as they pass through a nanopore1,2 eliminates the need for the enzymes and reagents used in other approaches. Theoretical calculations indicate that electron tunneling could identify bases in single-stranded DNA, yielding long reads and eliminating enzymatic processing.3–5 It was shown recently that tunneling can sense individual nucleotides6 and nucleosides.7 Here, we show that tunneling electrodes functionalized with recognition reagents can identify a single base flanked by other bases in a short DNA oligomer. The residence time of a single base in a recognition junction is on the order of a second, but pulling the DNA through the junction with a force of tens of piconewtons would yield reading speeds of tens of bases per second.

Nucleosides diffusing through a 2 nm electron-tunneling junction generate current spikes of sub-millisecond duration with a broad distribution of peak currents. This distribution narrows 10-fold when one of the electrodes is functionalized with a reagent that traps nucleosides in a specific orientation with hydrogen bonds. Functionalizing the second electrode reduces contact resistance to the nucleosides, allowing them to be identified via their peak currents according to deoxyadenosine > deoxycytidine > deoxyguanosine > thymidine, in agreement with the order predicted by a density functional calculation.

Hydrogen bonding has a ubiquitous role in electron transport1,2 and in molecular recognition, with DNA base-pairing being the best known example.3 Scanning tunneling microscope (STM) images4 and measurements of the decay of tunnel-current as a molecular junction is pulled apart by the STM tip, 5 are sensitive to hydrogen-bonded interactions. Here we show that these tunnel-decay signals can be used to measure the strength of hydrogen bonding in DNA basepairs. Junctions that are held together by three hydrogen bonds per basepair (e.g., guanine-cytosine interactions) are stiffer than junctions held together by two hydrogen bonds per basepair (e.g., adenine-thymine interactions). Similar, but less-pronounced, effects are observed on the approach of the tunneling probe, implying that hydrogen-bond dependent attractive forces also have a role in determining the rise of current. These effects provide new mechanisms for making sensors that transduce a molecular recognition event into an electronic signal.

No abstract

Single molecules in a tunnel junction can now be interrogated reliably using chemically-functionalized electrodes. Monitoring stochastic bonding fluctuations between a ligand bound to one electrode and its target bound to a second electrode (“tethered molecule-pair” configuration) gives insight into the nature of the intermolecular bonding at a single molecule-pair level, and defines the requirements for reproducible tunneling data. Simulations show that there is an instability in the tunnel gap at large currents, and this results in a multiplicity of contacts with a corresponding spread in the measured currents. At small currents (i.e. large gaps) the gap is stable, and functionalizing a pair of electrodes with recognition reagents (the “free analyte” configuration) can generate a distinct tunneling signal when an analyte molecule is trapped in the gap. This opens up a new interface between chemistry and electronics with immediate implications for rapid sequencing of single DNA molecules.