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.

No abstract

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.

Long DNA molecules can self-entangle into knots. Experimental techniques to observe such DNA knots (primarily gel electrophoresis) are limited to bulk methods and circular molecules below 10 kbp in length. Here we show that solid-state nanopores can be used to directly observe individual DNA knots in both linear and circular single molecules of arbitrary length. DNA knots are observed as short spikes in the nanopore current traces of traversing DNA molecules. The observation of knots is dependent on sufficiently high measurement resolution, which can be achieved using high-concentration LiCl buffers. We study the percentage of DNA molecules with knots for different DNA molecules, up to 166 kbp in length. We find that the knotting probability rises strongly with length, and compare our experimental data to 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 small sizes below 100 nm. In the case of linear molecules, we observe that knots are able to slide out upon applying high driving forces (voltage). Our results demonstrate that the solid-state nanopore technique can provide a wealth of information about the position and the size of knots, including the number of DNA strands inside DNA knots. 831-Pos Board B611Enhanced Electrostatic Force Microscopy Imaging Reveals Mechanism of TRF2 Mediated DNA Compaction Parminder kaur. Physics, North Carolina State University, Raleigh, NC, USA. T-loop formation at telomeres is proposed to play an important role in telomere protection through the sequestration of the 3' single-stranded overhang. Previous studies indicated that shelterin protein TRF2 modulates telomere structure by promoting DNA compaction and T-loop formation. To further understand the mechanism underlying TRF2-mediated DNA compaction, we applied Dual-Resonance-frequency-Enhanced Electrostatic force Microscopy (DREEM), a recently developed technique capable of high-resolution imaging of weak electrostatic potentials. DREEM images of nucleosomes clearly reveal DNA strands wrapped around histone proteins in nucleosomal arrays. In contrast, DREEM imaging shows DNA compacted inside TRF2 complexes through a 3-dimensional stacking of TRF2 dimers mediated by collective actions of multiple copies of TRF2 proteins. TRF2-mediated DNA compaction leads to electric potential gradients across the complexes. Surprisingly, while DNA wrapped around histones displays similar electrostatic potential signals compared to bare DNA, TRF2 DNA compaction leads to significant differences in the electrical potential signals inside multioligomeric TRF2 complexes compared to protein or DNA alone. These results clearly demonstrate the electrostatic changes in TRF2-DNA complexes upon oligomerizaton of proteins and DNA compaction, and underscore the importance of developing new electrostatic force microscopy imaging techniques for studying biological systems. 832-Pos Board B612Grab & Watch: Correlative Optical Tweezers-Flu...

simultaneously label by means of fluorescence the genetic locus and the synthesized mRNA using the EGFP-labeled MS2 coat protein [1]. Our method, previously applied to the tracking of gene arrays in cultured cells [2], has a temporal resolution of 10-100ms, and additionally records the 3D position of the genetic locus by moving along a circular orbit the focused laser beam. Distinct regions of active transcription display a well defined spatial organization, corralling the denser part of the genetic locus. In most cases each region maintains a defined angle in the reference system of the orbit, and the transcriptional activities of different regions are not cross-correlated. The fluorescence time traces of each of these regions highlight the existence of slow (10-100s) transitions between distinct intensity values, corresponding to the timescale of a single mRNA dwell on the gene or to that of a transcription burst. We observe autocorrelation of the fluorescence intensity on timescales smaller than 1s. We relate these fast fluctuations to the faster kinetics of mRNA transcription, down to individual MS2-EGFP molecules binding to the newly transcribed mRNAs. Measurements of the size and shape of the genetic array by calculating the modulation of the first and second harmonic of the fluorescence along each orbit suggest that the gene's decondensation is not a necessary condition for transcription to occur.