Electrostatic Control of DNA Hydridization Kinetics Studied with the Single-Molecule Field-Effect Transistor

Single-molecule studies generally rely on fluorescence-based reporting with signal levels limited by photon emission from single optical reporters to effective current levels in optical detectors of much less than 1 fA. Bioelectronic detection with a point-functionalized carbon nanotube transistor, known as the single-molecule field-effect transistor (smFET), in contrast offers signal levels that are more than 106 times higher. In our case, point functionalization is achieved with a nano-confined diazonium attachment chemistry. We previously used smFETs to investigate DNA hybridization kinetics, yielding rate constants, melting curves and activation energies for different oligonucleotides. Temporal analysis of association and dissociation reaction rate constants with temperature allows both target cDNA concentrations and free energies for hybridization to be determined. Here we show that hybridization kinetics are strongly affected by bias between the smFET device and the surrounding electrolyte, allowing bias to act as a proxy for temperature. We identify various concentrations of 20-mer target sequences from the Ebola Zaire nucleoprotein gene through smFET detection. Electrostatic modulation enables the detection of single base mismatches due to significantly altered kinetics under applied potential.