Direct observation of DNA knots using a solid-state nanopore.

Long DNA molecules can self-entangle into knots. Experimental techniques for observing such DNA knots (primarily gel electrophoresis) are limited to bulk methods and circular molecules below 10 kilobase pairs in length. Here, we show that solid-state nanopores can be used to directly observe individual knots in both linear and circular single DNA molecules of arbitrary length. The DNA knots are observed as short spikes in the nanopore current traces of the traversing DNA molecules and their detection is dependent on a sufficiently high measurement resolution, which can be achieved using high-concentration LiCl buffers. We study the percentage of molecules with knots for DNA molecules of up to 166 kilobase pairs in length and find that the knotting occurrence rises with the length of the DNA molecule, consistent with a constant knotting probability per unit length. Our experimental data compare favourably with previous simulation-based predictions for long polymers. From the translocation time of the knot through the nanopore, we estimate that the majority of the DNA knots are tight, with remarkably small sizes below 100 nm. In the case of linear molecules, we also observe that knots are able to slide out on application of high driving forces (voltage).

Detection of Individual Proteins Bound along DNA Using Solid-State Nanopores.

DNA in cells is heavily covered with all types of proteins that regulate its genetic activity. Detection of DNA-bound proteins is a challenge that is well suited to solid-state nanopores as they provide a linear readout of the DNA and DNA-protein volume in the pore constriction along the entire length of a molecule. Here, we demonstrate that we can realize the detection of even individual DNA-bound proteins at the single-DNA-molecule level using solid-state nanopores. We introduce and use a new model system of anti-DNA antibodies bound to lambda phage DNA. This system provides several advantages since the antibodies bind individually, tolerate high salt concentrations, and will, because of their positive charge, not translocate through the pore unless bound to the DNA. Translocation of DNA-antibody samples reveals the presence of short 12 µs current spikes within the DNA traces, with amplitudes that are about 4.5 times larger than that of dsDNA, which are associated with individual antibodies. We conclude that transient interactions between the pore and the antibodies are the primary mechanism by which bound antibodies are observed. This work provides a proof-of-concept for how nanopores could be used for future sensing applications.

Room-temperature gating of molecular junctions using few-layer graphene nanogap electrodes.

We report on a method to fabricate and measure gateable molecular junctions that are stable at room temperature. The devices are made by depositing molecules inside a few-layer graphene nanogap, formed by feedback controlled electroburning. The gaps have separations on the order of 1-2 nm as estimated from a Simmons model for tunneling. The molecular junctions display gateable I-V-characteristics at room temperature.