Session J50: Focus Session: Translocation through Nanopores - Novel Devices and Computational Approaches

Integrated Nanopore Detectors in a Standard Complementary Metal-Oxide-Semiconductor Process

High-bandwidth and low-noise nanopore sensor and detection electronics are crucial in achieving single-DNA base resolution. A potential way to accomplish this goal is to integrate solid-state nanopores within a CMOS platform, in close proximity to the biasing electrodes and custom-designed amplifier electronics. Here we report the development of solid-state nanopore devices in a commercial CMOS potentiostat chip implemented in On-Semiconductor's 0.5 micron technology. By using post-CMOS micromachining, a free-standing oxide membrane and electrodes are fabricated utilizing the N+ polysilicon/oxide/N+ polysilicon capacitor structure available in the aforementioned process. Nanopores with sub-5 nm diameter are drilled in the membrane using a Transmission Electron Microscope. The integrity of pores is validated by measuring current-voltage and noise characteristics. DNA translocation experiments are also performed utilizing these on-chip pores. In addition, electrical tests performed on the CMOS potentiostat circuitry show that the post-CMOS micromachining process does not have any detrimental effect on the CMOS circuitry.   

Graphene Gating of Solid State Nanopores for DNA Translocation

We report on ionic current measurements through gated solid state nanopores. Devices consist of Si$_{3}$N$_{4}$ membranes covered with a graphene sheet connected off-membrane to a gold contact pad. The graphene is insulated from solution with a TiO$_{2}$ layer deposited by atomic layer deposition, and the gold is exposed by an SF$_{6}$ etch. An electron-beam sculpted nanopore below 10 nm in diameter is drilled through the silicon nitride, graphene, and titania. Applying a voltage to the graphene modulates the ionic current through the pore. We measured the current-voltage characteristics for different gate potentials for our devices. We characterized the leakage current from the graphene as well as the ionic current noise in these pores to complement our current-voltage measurements. These results can lead to measurements of the influence on DNA translocation of a potential at the pore and set the groundwork for characterization of graphene-based sensing of DNA at a nanopore.   

Nanochannel Device with Embedded Nanopore: a New Approach for Single-Molecule DNA Analysis and Manipulation

Nanopore and nanochannel based devices are robust methods for biomolecular sensing and single DNA manipulation. Nanopore-based DNA sensing has attractive features that make it a leading candidate as a single-molecule DNA sequencing technology. Nanochannel based extension of DNA, combined with enzymatic or denaturation-based barcoding schemes, is already a powerful approach for genome analysis. We believe that there is revolutionary potential in devices that combine nanochannels with nanpore detectors. In particular, due to the fast translocation of a DNA molecule through a standard nanopore configuration, there is an unfavorable trade-off between signal and sequence resolution. With a combined nanochannel-nanopore device, based on embedding a nanopore inside a nanochannel, we can in principle gain independent control over both DNA translocation speed and sensing signal, solving the key draw-back of the standard nanopore configuration. We will discuss our recent progress on device fabrication and characterization. In particular, we demonstrate that we can detect - using fluorescent microscopy - successful translocation of DNA from the nanochannel out through the nanopore, a possible method to 'select' a given barcode for further analysis. In particular, we show that in equilibrium DNA will not escape through an embedded sub-persistence length nanopore, suggesting that the embedded pore could be used as a nanoscale window through which to interrogate a nanochannel extended DNA molecule.   

fd Virus as a Model Stiff Polymer for Translocation Experiments with Solid-State Nanopores

We report preliminary experimental results of the translocation of the filamentous virus fd through a solid-state nanopore. fd virus is suitable for translocation and detection in a voltage-biased nanopore because it is highly charged, 880 nm long, and 6.6 nm in diameter. Importantly, fd has a persistence length of $\sim $2 $\mu $m, a forty-fold increase over dsDNA, making fd a model stiff polymer for testing theories of polymer translocation dynamics. fd cannot coil in solution, therefore the dispersion of fd translocation times can test a model by Lu et al. that ascribes DNA translocation velocity fluctuations to the distribution of initial conformations of the DNA coil. That picture is in contrast with an alternative model by Li et al., which attributes the spread of DNA translocation times to thermal velocity fluctuations. The physics of fd capture by a nanopore also differs significantly from DNA since the ends of the virus cannot diffusively search for the pore independently of the middle. As a result, the rate of fd capture from solution may not increase monotonically with the applied voltage across the pore; it is possible for fd to become kinetically trapped against the nanopore membrane by the electric field. We will compare the distribution of translocation times of fd virus to distributions for DNA and discuss the influence of the virus's orientation and interactions with the nanopore on the translocation speed and the measured current blockage. We will also examine the dependence of capture rate on the applied voltage.   

Studies of DNA Translocation Dynamics Using Asymmetrical Nanopores

Despite extensive studies of DNA translocations through voltage-biased solid-state nanopores, the influence of the DNA coil on the translocation dynamics remains poorly understood. We investigated this issue experimentally by controlling the separation between the DNA coil and the nanopore. We studied lambda DNA translocations through devices comprising a 400 nm-high, ~2500 nm-wide, disc-shaped cavity bounded from above by a 20 nm-thin silicon nitride membrane with a ~10 nm wide nanopore in the center, and from below by at 400 nm-thick silicon nitride membrane with a 300 nm-wide opening in the center. The asymmetric nanopore-cavity structure introduced an 800 nm gap between the initial DNA coil and the nanopore when a molecule translocated from below, but no gap when it translocated from above. Translocation times were longer and the integrated charge deficit was larger for molecules translocating from below. These results are explained by the viscous drag on the DNA outside the pore, whose importance relative to the drag inside the pore we quantify. We outline a consistent model of DNA translocation speeds that depends on the initial configuration of the DNA coil, similar to the velocity fluctuation model of Lu et al.   

First-passage-time analysis of DNA translocation in solid-state nanopores

We report a DNA translocation experiment using solid-state nanopores and 48 kb lambda DNA samples. As reported previously, the DNA translocation dynamics in such standard solid-state nanopore experiments appear to be complex and multiple folded translocation pathways are observable. We use the translocation events with little or no detectable folded structures to construct a distribution function for the DNA translocation times. We find that the translocation time distribution can be fitted using the first-passage-time probability density function derived by Schrodinger for 1-D Brownian motion with a drift. The voltage dependence of the extracted DNA drift velocity shows excellent agreement with the Stokes' law at high voltages. Deviation from the Stokes' law is found at low voltages, but can be attributed to a systematic error in how different types of folded DNA translocations are sorted.   

Some aspects of polymer translocation dynamics through nanopore: comparison of recent the theories with simulation results

Translocation of a flexible poymer chain through a narrow pore has still remained an active field of research. Earlier theoretical studies of Sung and Park,\footnote{W. Sung and P.~J. Park, Phys. Rev. Lett. {\bf 77}, 783 (1996).} Muthukumar,\footnote{M. Muthukumar, J. Chem. Phys. {\bf 111}, 10371 (1999).} Chuang, Kantor and Kardar, Kantor and Kardar\footnote{J. Chuang, Y. Kantor and M. Kardar, Phys. Rev. E {\bf 65}, 011802 (2001); Y. Kantor and M. Kardar, \textit{ibid.} {\bf 69}, 021806 (2004).} for a flexible chain have been complemented by more recent theories of Sakaue\footnote{T. Sakaue, Phys. Rev. E {\bf 76}, 021803 (2007); \textit{ibid.} \textbf{81}, 041808 (2010).} where tension propagation(TP) along the chain backbone at the $cis$ side resulting in a nonuniform stretching of the chain has been proposed to be a key input for theoretical studies. Recently these elements of the TP theory has been incorporated in to a Brownian dynamics (BDTP) scheme and numerical studies of the equations of motion are in excellent agreement with prior simulation studies.\footnote{T. Ikonen, A. Bhattacharya, T. Ala-Nissila and W. Sung (submitted).} A driven translocating chain is essentially \textit{out-of-equilibrium}\footnote{A. Bhattacharya and Kurt Binder, Phys. Rev. E. \textbf{81}, 041804 (2010); A. Bhattacharya \textit{et al.}, Eur. Phys. J. E \textbf{29}, 423 (2009).} which results in \textit{cis-trans} asymmetries both in ocnformations and in dynamics. Therefore, results from theoretical studies should capture these features. In this talk first I will first present results from Langevin dynamics simulation citing several cases where how this \textit{cis-trans} asymmetry affects the chain conformations and the translocation dynamics. Then I will dicuss relevance of these results in the context of exisiting theories.   

DNA translocation measurements in solid-state nanopores fabricated using helium-ion microscope

We report high-quality DNA translocation measurements in solid-state nanopores drilled in free-standing SiN membranes by using a helium-ion beam in a Zeiss helium-ion microscope (HIM). We show that the HIM nanopores have similar performance as the TEM-drilled pores.   

Dynamics of Simultaneous, Single Ion Transport through Two Single-Walled Carbon Nanotubes: Observation of a Three-State System

The ability to actively manipulate and transport single molecules in solution has the potential to revolutionize chemical synthesis and catalysis. In previous work, we developed a nanopore platform using the interior of a single-walled carbon nanotube (diameter = 1.5 nm) for the Coulter detection of single cations of Li+, K+, and Na+. We demonstrate that as a result of their fabrication, such systems have electrostatic barriers present at their ends that are generally asymmetric, allowing for the trapping of ions. We show that above this threshold bias, traversing the nanopore end is not rate-limiting and that the pore-blocking behavior of two parallel nanotubes follows an idealized Markov process with the electrical potential. Such nanopores may allow for high-throughput linear processing of molecules as new catalysts and separation devices.   

Polymer Translocation through a Nanopore Modulated by a Sticky Site

We examine the translocation of a long polymer that has one ``sticky site'' where a single monomer experiences an attraction to the pore. Using a quasistatic model for the translocation dynamics and employing numerically exact methods to generate results, high precision values for the translocation times are obtained across a wide range of driving forces and sticky site well depths. It is found that the interplay between the sticky site well depth, the driving force, and entropic effects can lead to unexpected results such as a non-monotonic variation of the critical well depth with the driving force and abnormally long translocation times due to the generation of a metastable state at a critical driving force. The dynamics are also found to be strongly dependent on the location of the sticky site: a site near the head of the polymer increases the probability of successful translocation while a site in the middle acts merely as a ``speedbump.'' Treating the sticky site as a perturbation to an otherwise diffusive process (low driving forces) or driven process (high driving forces) yields good agreement with the numerical results.   

Translocation dynamics of a semi-flexible chain through a nano-pore

We study translocation dynamics of a semi-flexible chain through a nano-pore in two dimensions (2D) using Langevin dynamics simulation. Specifically, we show how the mean first passage time (MFPT) and the probability distribution of the MFPT are both influenced by the bending rigidity of the chain. Furthermore, we monitor the chain conformations both at the cis and the trans sides and relate these results with recent theories and experiments for a translocating chain through a nano-pore.   

Characterization of Idealized Helical Repeat Proteins in Silicon Nitride Nanopores

In this work, we report the measurement of consensus tetratricopeptide repeat (CTPR) proteins with single silicon nitride nanopores. The CTPR proteins were measured in KCl solution at pH below and above its isoelectric point (pI), as well as with and without denaturing agent, Guanidine HCl. When a CTPR protein molecule transits through a nanopore driven by an applied voltage, it partially blocks the ions (K$^{+}$ and Cl$^{-})$ flow in the nanopore and generates a characteristic electric current blockage signal. The current blockage signal reveals information about the size, conformation, and primary sequence of the CTPR protein molecule. Previous translocation studies carried out with DNA have established that higher bias voltages result in shorter duration current blockages indicating that DNA translocates faster at a stronger electric field. However, our CTPR translocation studies show that longer duration current blockades were observed at higher bias voltages. We discuss how the inhomogeneous distribution of the primary charge sequence of the CTPR proteins predicts translocation barriers that are proportional to the bias voltage. Larger barriers at higher bias voltages will result in longer translocation times, consistent with our experimental results.