Session P30: Nanopores and Related Structures for DNA Detection

The statistics of a single DNA capture by a solid-state nanopore

We have investigated the statistics of DNA threading through solid-state nanopores that are approximately 10 nm in diameter. Intense electric fields are generated in the vicinity of the pore when a voltage is applied across it in ionic solution. The electric forces experienced by a negatively charged DNA molecule are sufficient to pull it through in a folded, ``hairpin'' configuration. The ionic current blockade signal that results offers information about where along the 16.5 micrometer-long DNA molecule the fold was induced. We have analyzed the results of translocation experiments to build a probability distribution for the DNA capture location. We propose a simple polymer scaling theory to explain the results. Our model is based on the equilibrium distribution of polymer conformations in solution, and it predicts the observed bias for capturing molecules near the ends.   

Fabrication of a CMOS compatible nanopore detector for DNA

Nanopore based DNA sequencers require integration of miniaturized electrodes and amplifier electronics in close proximity to the nanopores in a CMOS platform. This will facilitate portability, enable faster analysis, and improve sensing performance. Here we report for the first time the fabrication of a DNA nanopore detector compatible with a standard CMOS process. Our nanopore devices are made using an N+ polysilicon/gate oxide/N+ polysilicon stack on an oxidized silicon substrate identical to the AMI 0.5$\mu $ process. The nanopores are created in the gate oxide membrane (36 nm) while doped polysilicon layers (250 and 370 nm) act as electrodes to apply bias across pores. Five lithography masks are used to pattern the oxide membrane and the electrodes. The nanopores are defined by etching the membrane using electron beam lithography patterned holes in a resist mask. Using this method we have directly fabricated pores with diameters as small as 11 nm, without applying conventional pore shrinkage techniques. This is enhanced by cold development of the e-beam exposed resist resulting in sub-10 nm pores. DNA experiments are currently underway utilizing our nanopores.   

Controlled Reverse Translocation of DNA Through a Solid-State Nanopore

Reverse DNA translocation [1] is a process in which a DNA is pulled out of a nanopore against the forward electric force. In this process, the effective diffusion constant of the DNA is significantly reduced, thereby suppressing thermal smearing effect due to diffusion in the positional measurements of DNA sequences, e.g. in the hybridization-assisted nanopore sequencing platform. We describe a new experimental setup which will provide dramatic improvement upon the previous experiment [1]. \\[4pt] [1] Hongbo Peng and X.S. Ling, ``Reverse DNA translocation through a solid-state nanopore by magnetic tweezers,'' Nanotechnology, \underline {20}, 185101(2009).   

Electro-fluidic gating in solid-state nanopores with a chemically reactive surface

We are exploring the use of electrically functionalized solid-state nanopores for controlling the transport of ions and single DNA molecules in solution. Modulating the surface charge density in a fluidic device differs crucially from gating in a semiconductor because real, chemically reactive surfaces obtain a charge density spontaneously in contact with solution. An applied gate field can shift the chemical equilibrium of the ionizable surface groups. Here we present an electrochemical model of electro-fluidic gating that captures the influence of pH, salt concentration, and specific chemical surface groups. We have also tested ion transport in electrostatically actuated nanopore devices in which the applied gate potential influenced the ionic conductance through the pore. Our experiments reveal how applied electric fields can influence the density of mobile counter-ions inside the pore, but that capacitive charging and the chemistry of the surface both play important roles.   

Characterize and Control the Motion of DNA in a Solid-State Nanopore

Controlling the motion of a single-stranded DNA (ssDNA) at a single-base resolution is critical to all nanopore based DNA sequencing technologies. Experimental studies till now demonstrated that the overall translocation speed of DNA driven by a biasing electric field could be affected by ion concentration, solvent viscosity or temperature. Although the DNA translocation could be slowed down, the instant motion of DNA is too diffusive to allow each DNA base to be measured. Using extensive all-atom molecular dynamics simulations, we studied the diffusion constant, friction coefficient, electrophoretic mobility, and effective charge of ssDNA in a solid state nanopore. Simulation results showed that the spatial fluctuation of ssDNA in one nano-second is comparable to the spacing between neighboring DNA bases, which makes the sensing of a DNA base very difficult. The recently proposed DNA transistor (Appl. Phys. Lett. 91, 153103 (2007)) could potentially solve this problem by electrically trapping ssDNA inside a nanopore. Our simulations demonstrated that the DNA transistor could achieve base-by-base ratcheting of ssDNA when ssDNA is either pulled by an optical tweezer or driven by a biasing electric field. Using the Fokker-Planck equation, simulated motion of ssDNA in the DNA transistor was theoretically characterized.   

Functionalizing a nanopore with nano-electrodes for control of the translocation of DNA with single base resolution

Recently, application of nanopores to low-cost DNA sequencing has attracted great interest as there is great need to reduce the cost of sequencing a whole human genome to \$1000. A key issue in the field of nanopore DNA sequencing is to control the DNA translocation. Here we will report the development of what we call a DNA transistor: a nanopore-based electrical device for controlling the translocation of DNA with single base resolution. The key part of this device is a free standing membrane, within which multiple layers of electrically addressable metal electrodes separated by dielectric layers are embedded. A 1-5 nanometer size pore is made through the membrane. We demonstrated that such a device is electrically viable for the electrode layer or the spacing dielectric layer as thin as 3 nm. Confirming the basic function of the device, induced electrical signals on the nano-electrodes by the translocating DNA, as well as the modulation of DNA translocation speed by the voltage bias applied on the nano- electrodes are also observed. Our ongoing experiments test if the modulated electrical field can trap or translocate DNA at a single base resolution.   

Solid-state nanopore studies of hybridized DNA oligomers

Hybridization-assisted nanopore sequencing (HANS) uses short oligomers of DNA bound to a long single-stranded DNA in order to obtain the positional information of the bases that make up the long DNA molecule. To test the feasibility of the HANS approach, we carried out experiments to detect 12-base hybridizations in a tri-mer complex consisting of three single-stranded oligos hybridized at their ends sequenctially. These DNA complexes are connected to polystyrene beads through biotin-streptavidin bonds to enable their detection by nanopores. The experiment is to measure the time dependence of the nanopore ionic current at fixed voltage when the \textit{cis} side is filled with the oligo-attached beads. Computer simulations are used as guides in the identification of translocation dynamics. Distinct features are found that can be attributed to tri-mers, dimmers, and monomers attached to the beads. The measured mean-first-passage time between two hybridization segments is extracted and is found to be consistent with theoretical estimates.   

An integrated nanopore-nanochannel system for biodetection: longitudinally-displaced transverse nanoelectrodes along a nanochannel

In this talk, I'll describe a novel device concept for a biodetection system with combined characteristics of nanopores and nanochannels. Solid-state nanopores drilled in a thin membrane have a short channel length of the order of 20nm or smaller, capable of detecting short features. Nanochannels have their own useful property of providing a 1D comfinement for a DNA. To combine these two devices in a single system is of great interest to DNA sequencing and other biodetection applications. Here I describe a new device concept capable of this goal. The device consists of a pair of longitudinally-displaced transverse nanoelectrodes as voltage probes along a nanochannel. The longitudinal displacement should be of 20nm or smaller, thereby effectively making a nanopore on a segment of the nanochannel. I'll describe a feasibility study using a commercial software COMSOL. The sensitivity issues will be discussed.   

Memory effects in the ion dynamics in nanopores

We have performed molecular dynamics simulations of ion dynamics across solid-state nanopores subject to a periodic electric field. We have analyzed both the ion build-up at the surfaces of the pores as well as the ion conductivity. Our results show that ion dynamics exhibits memory effects due to both the finite relaxation times of ions in solution as well as the water polarization/depolarization time-scales. These memory effects are reminiscent of the recently proposed memcapacitance (capacitance with memory) features of certain non-linear circuit elements [1], and may have important implications in DNA dynamics and sequencing [2]. [1] M. Di Ventra, Y.V. Pershin and L.O. Chua, Proc. IEEE 97, 1717 (2009). [2] M. Zwolak and M. Di Ventra, Rev. Mod. Phys. 80, 141 (2008).   

Quantized ionic conductance in nanopores

Ionic transport in nanopores is a fundamentally and technologically important problem in view of its occurrence in biological processes and its impact on novel DNA sequencing applications. Using molecular dynamics simulations we show that ion transport may exhibit strong nonlinearities as a function of the pore radius reminiscent of the conductance quantization steps as a function of the transverse cross section of quantum point contacts. In the present case, however, conductance steps originate from the break up of the hydration layers that form around ions in aqueous solution. We discuss this phenomenon and the conditions under which it should be experimentally observable.   

Detection of DNA using longitudinally-displaced microelectrodes along a microfluidic channel as voltage probes

In this talk, we describe a micron-scale prototype of a novel device concept [1] of longitudinally-displaced nanoelectrodes along a nanoscale fluidic channel as voltage probes for DNA detection. Preliminary results of simultaneous fluorescence imaging and voltage sensing will be presented. \\[4pt] [1] X. S. Ling, abstract in this meeting.   

Probing electrostatic interactions between DNA and the walls of slit-like nanofluidic channels

Recent progress in nanofluidic technology suggests the possibility of controlling single DNA molecules using purely electrostatic forces, which are influenced by the ionic strength. We studied the ionic dependence of DNA conformations in slit-like nanochannels of varying height using fluorescence video microscopy. By applying polymer scaling theory to the measured radius of gyration of individual molecules, we inferred the electrostatic interaction between the negatively charged DNA and the negatively charged channel walls. With decreasing ionic strength, the excluded region near the channel wall showed an abrupt saturation around $\sim $1mM that was not anticipated by existing theories. We can explain our observations by considering the DNA segments as multivalent ions which strongly influence the local ionic strength, and hence the electrostatic screening length. These results allow us to predict the magnitude and range of the electrostatic forces that can be exerted on confined DNA molecules using the electro-fluidic field effect.   

Fundamental interaction between Au quantum dots and DNA

Semiconductor quantum dots (QDs) and metal nanoparticles (NPs) have attracted a great deal of attention in biology community due to their application as fluorescent labels and sensors. The optical properties of QDs and NPs allow them to be effective imaging agents. However, QDs have the potential to be used as more than just sensors and labels. Their biological sensing abilities include identifying target DNA through a linker followed by color change and electrical signaling. If this property can be combined with the direct binding of QDs with DNA, many other applications in bio-nanotechnological field are possible. In this paper, we investigate the interaction between colloidal Au QDs and 30-base sequence single strand DNA. Our preliminary results indicate that the DNA strand tend to form different structures in the presence of Au QDs. Furthermore, small as well as large agglomerated Au particles appear to be linked along the DNA strand.   

A Low Temperature STM Manipulation of a Molecular Rotor

4Fe3Set molecules consist of ferrocene arms mounted on a ruthenium atomic bearing atop a molecular stand and thus allowing rotation of arms when the molecule is absorbed on a metal surface with its base. Here, we present a study of molecule rotor (4Fe3Set) deposited on Au(111) substrate using Scanning Tunneling Microscope (STM) at 4.6K and 75K, respectively. At 4.6 K, molecules form stable configurations without rotation on the surface due to a lower thermal energy at this temperature. In contrast, the spinning rotors were routinely detected at 75 K substrate temperature. Step-wise rotation of the rotator part of the molecule is realized by inelastic tunneling electron excitation using the STM-tip. Rotation rate was found to vary with tunneling current, and molecule preferred some quantized angles to rotate with.