Patent Application
20210190724
The disclosure relates to biosensors, and in particular to methods, apparatus and systems for single-molecule biosensors having a transition metal dichalcogenide or silicon nanobridge for sequencing, information storage and reading.
Analysis of biomolecules including DNAs and genomes has received an increasing amount of attention in recent years in various fields of precision medicine or nanotechnology. The seminal work of Maclyn McCarty and Oswald T. Avery in 1946 (see, “Studies On The Chemical Nature Of The Substance Inducing Transformation Of Pneumococcal Types II. Effect Of Desoxyribonuclease On The Biological Activity Of The Transforming Substance,” The Journal of Experimental Medicine 83(2), 89-96 (1946)), demonstrated that DNA was the material that determined traits of an organism. The molecular structure of DNA was then first described by James D. Watson and Francis HC Crick in 1953 (see a published article, “Molecular structure of nucleic acids.”, Nature 171,737-738 (1953)), for which they received the 1962 Nobel Prize in Medicine. This work made it clear that the sequence of chemical letters (bases) of the DNA molecules encode the fundamental biological information. Since this discovery, there has been a concerted effort to develop means to actually experimentally measure this sequence. The first method for systematically sequencing DNA was introduced by Sanger, et al in 1978, for which he received the 1980 Nobel Prize in Chemistry. See an article, Sanger, Frederick, et al., “The nucleotide sequence of bacteriophage φX174.” Journal of Molecular Biology 125, 225-246 (1978).
Sequencing techniques for genome analysis evolved into utilizing automated commercial instrument platform in the late 1980's, which ultimately enabled the sequencing of the first human genome in 2001. This was the result of a massive public and private effort taking over a decade, at a cost of billions of dollars, and relying on the output of thousands of dedicated DNA sequencing instruments. The success of this effort motivated the development of a number of “massively parallel” sequencing platforms with the goal of dramatically reducing the cost and time required to sequence a human genome. Such massively parallel sequencing platforms generally rely on processing millions to billions of sequencing reactions at the same time in highly miniaturized microfluidic formats. The first of these was invented and commercialized by Jonathan M. Rothberg's group in 2005 as the 454 platform, which achieved thousand fold reductions in cost and instrument time. See, an article by Marcel Margulies, et al., “Genome Sequencing in Open Microfabricated High Density Picoliter Reactors,” Nature 437, 376-380 (2005). However, the 454 platform still required approximately a million dollars and took over a month to sequence a genome.
The 454 platform was followed by a variety of other related techniques and commercial platforms. See, articles by M. L. Metzker, “Sequencing Technologies—the Next Generation,” Nature Reviews Genetics 11(1), 31-46 (2010), and by C. W. Fuller et. al, “The Challenges of Sequencing by Synthesis,” Nature Biotechnology 27(11), 1013-1023 (2009). This progress lead to the realization of the long-sought “$1,000 genome” in 2014, in which the cost of sequencing a human genome at a service lab was reduced to approximately $1,000, and could be performed in several days. However, the highly sophisticated instrument for this sequencing cost nearly one million dollars, and the data was in the form of billions of short reads of approximately 100 bases in length. The billions of short reads often further contained errors so the data required interpretation relative to a standard reference genome with each base being sequenced multiple times to assess a new individual genome.
Thus, further improvements in quality and accuracy of sequencing, as well as reductions in cost and time are still needed. This is especially true to make genome sequencing practical for widespread use in precision medicine (see the aforementioned article by Fuller et al), where it is desirable to sequence the genomes of millions of individuals with a clinical grade of quality.
While many DNA sequencing techniques utilize optical means with fluorescence reporters, such methods can be cumbersome, slow in detection speed, and difficult to mass produce to further reduce costs. Label-free DNA or genome sequencing approaches provide advantages of not having to use fluorescent type labeling processes and associated optical systems, especially when combined with electronic signal detection that can be achieved rapidly and in an inexpensive way.
In this regard, certain types of molecular electronic devices can detect single molecule, biomolecular analytes such as DNAs, RNAs, proteins, and nucleotides by measuring electronic signal changes when the analyte molecule is attached to a circuit, for example, a field effect transistor device. Such methods are label-free and thus avoid using complicated, bulky and expensive fluorescent type labeling apparatus. These methods can be useful for lower cost sequencing analysis of DNA, RNA and genome.
Certain types of molecular electronic devices can detect the biomolecular analytes such as DNAs, RNAs, proteins, and nucleotides by measuring electronic signal changes when the analyte molecule is attached to the circuit comprising a pair of conductive electrodes. Such methods are label-free and thus avoids using complicated, bulky and expensive fluorescent type labeling apparatus. One example of sequencing-by-synthesis approach is to utilize a single molecule polymerase with incorporated DNAs, the sequence of which is detected through a current pulse signal when each type of the nucleotides (A,T,C, G) is attached to the polymerase complex with a distinct electrical signal.
While current molecular electronic devices can electronically measure molecules for various applications, they lack the reproducibility as well as scalability and manufacturability needed for rapidly sensing many analytes at a scale of up to millions in a practical manner. Such highly scalable methods are particularly important for DNA sequencing applications, which often need to analyze millions to billions of independent DNA molecules. In addition, the manufacture of current molecular electronic devices is generally costly due to the high level of precision needed.
Disclosed herein are principles that provide new and improved sequencing apparatuses, device structures and methods using two-dimensional layer structured semiconductors, which can provide reliable DNA genome analysis performance and are amenable to scalable manufacturing. In various embodiments, the present disclosure provides nanofabrication of biomolecular sensing devices and fabrication of devices for analyzing DNA and related biomolecules. In various embodiments, the present disclosure provides DNA-based memory systems.
In various embodiments herein, biomolecular sensors comprise a nanobridge structure disposed over a nanogap, wherein the nanobridge comprises a transition metal dichalcogenide (TMD) material or a silicon material, e.g., pure crystalline silicon or various doped silicon semiconductor materials.
Two dimensional (2D) layered transition metal dichalcogenides (TMDs) materials and devices have attracted a great deal of interest due to their novel electronic, physical and chemical characteristics. One example is MoS2 which can be incorporated as a sensor device. MoS2 type 2D materials can be a single layered material or several layered material, and can be obtained by various techniques, such as e.g., by isolation of very thin MoS2 layer through mechanical exfoliation, physical or chemical vapor deposition, molecular beam epitaxy (MBE) type construction or sulfurization of transition metal layer such as Mo or W.
Transition metal dichalcogenide (TMD) monolayers are in general atomically thin semiconductors of the type MX2, which M a transition metal atom (notably Mo, W, or Ti, Zr, Hf, V, Nb, Ta, Tc, Re, Co, Rh, Ir, Ni, Pd, Pt) and X a chalcogen atom (S, Se, or Te). One layer of M atoms is sandwiched between two layers of X atoms. A MoS2 monolayer can be about 6.5 Å thick. TMD monolayers of e.g., MoS2, WS2, MoSe2, WSe2, MoTe2 have a direct band gap, and can be used in electronics as transistors or sensors. Either monolayer TMD or few layer TMD can be structurally modified, to be utilized as solid state DNA or genome sensors. Being an ultrathin direct bandgap semiconductor, a transition metal dichalcogenide such as single-layer MoS2 may have potential for widespread applications in nanoelectronics, optoelectronics, and energy harvesting.
The layered TMDs typically have a hexagonal type structure with space group P63/mmc. It should be noted that monolayers of TMD materials are not just one atom thick as graphene, but are made up with tri-atomic thick layers consisting of metal atoms (such as Mo or W) sandwiched between two layers of chalcogen atoms (such as S, Se, or Te). The atoms in-plane in MoS2 type 2D materials are put together and bonded by strong covalent bonds. The adjacent layers of TMD like MoS2 along the thickness direction are joined together by a weak van der Wall force binding. This force is strong enough to hold the layers together with mechanical integrity. The TMD materials provide interesting and unique possibilities to design electronic devices involving hetero structures. The direct band gap of TMD monolayers is tunable with the application of the mechanical strain.
Single nucleotide identification and DNA sequencing have already been demonstrated with biological nanopores or solid state nanopores such as those in graphene and MoS2 layers. A DNA type molecule is threaded through a nanopore under an applied electric field, so that the sequence of nucleotides is read by monitoring small changes in the ionic current flowing through the pore, which are induced by individual nucleotides temporarily residing within the pore during threading. However, the fragility of such pores, together with difficulties related to reproducible and low noise measurement of detection signals in nanopore sequencing methods in general are some of the current issues that need to be addressed.
The disclosed principles provide, among others, new biomolecular sensor devices and associated methods, employing transition metal dichalcogenide nanoribbons as a component of molecular bridge, which in turn comprises an attached, preferably single molecule polymerase to analyze DNA lines or fragments by step by step attachments of nucleotides or short DNA fragments.
Various embodiments are disclosed herein regarding specially processed, 2D layer-containing enzyme polymerase sensor device structures and methods of manufacture for a multitude of devices for use in electronic DNA, RNA or genome sequencing systems. Unique geometrical modifications are made so as to enable a construction of sensor device comprising only a single molecule polymerase enzyme for more accurate electronic analysis. Such label-free, single molecule based sequencing analysis systems utilize preferably a nanoscale dimension-controlled, transition metal dichalcogenide (TMD) micro-ribbon or nano-ribbon bridge. The electronic system may also be used in analyzing other types of biomolecules, such as proteins, depending on how the molecular sensors are functionalized to interact with biomolecule sensing targets. The TMD-based sequencing systems disclosed here can be assembled into a massively parallel configuration for rapid analysis of targets including nucleotides, in particular for applications to sequencing of a DNA molecule, or a collection of such molecules constituting an entire human genome. Such systems in the present disclosure can also be used for DNA-based information storage, for example, for archival storage of huge volume of information in human society.
In various embodiments, a single-molecule biosensor comprises: a conducting electrode pair disposed on a substrate, the electrode pair comprising a first electrode and a second electrode spaced-apart from the first electrode by a nanogap of fixed width; a TMD, Si, or doped Si-semiconductor layer disposed on the substrate electrically connecting the first and second electrodes; a dielectric layer disposed completely over the first and second electrodes to encase the first and second electrodes, and disposed partially over the TMD, Si, or doped Si-semiconductor layer so as to leave an exposed portion of the TMD, Si, or doped Si-semiconductor layer having a width less than the width of the nanogap; an enzyme molecule attached to the exposed portion of the TMD, Si, or doped Si-semiconductor layer; and a microfluidic system encasing the conducting electrode pair, the contiguous TMD layer and the enzyme molecule attached thereto.
In various embodiments, the TMD layer comprises at least one TMD having a structure MS(2−x), MS(2+x), MSe(2−x), MSe(2+x), MTe(2−x) or MTe(2+x), wherein x is 0-0.3, and M is Mo, W, Ti, Zr, Hf, V, Nb, Ta, Tc, Re, Co, Rh, Ir, Ni, Pd, or Pt.
In various embodiments, the TMD layer comprises a TMD selected from the group consisting of MoS2, WS2, TiS2, ZrS2, HfS2, VS2, NbS2, TaS2, TcS2, ReS2, CoS2, RhS2, IrS2, NiS2, PdS2, PtS2, MoSe2, WSe2, TiSe2, ZrSe2, HfSe2, VSe2, NbSe2, TaSe2, TcSe2, ReSe2, CoSe2, RhSe2, IrSe2, NiSe2, PdSe2, PtSe2, MoTe2, WTe2, TiTe2, ZrTe2, HfTe2, VTe2, NbTe2, TaTe2, TcTe2, ReTe2, CoTe2, RhTe2, IrTe2, NiTe2, PdTe2, PtTe2, and mixtures thereof.
In various embodiments, the TMD layer comprises a mixed TMD compound selected from the group consisting of Mo(SxSeyTez)2, W(SxSeyTez)2, Ti(SxSeyTez)2, Zr(SxSeyTez)2, Hf(SxSeyTez)2, V(SxSeyTez)2, Nb(SxSeyTez)2, Ta(SxSeyTez)2, Tc(SxSeyTez)2, Re(SxSeyTez)2, Co(SxSeyTez)2, Rh(SxSeyTez)2, Ir(SxSeyTez)2, Ni(SxSeyTez)2, Pd(SxSeyTez)2, Pt(SxSeyTez)2, and mixtures thereof, wherein (x+y+z) is 0.7-1.3.
In various embodiments, the TMD layer comprises at least one TMD compound of structure M(1−w)NyX(2−z)Yz, wherein M is Al, Si, Ga, Ge, In, Sn, Sb, Bi, Na, K, Ca, Mg, Sr, or Ba; X is S, Se, or Te; Y is Li, B, C, N, O, P, F, Cl, or I; w is 0-0.3; and z is 0-0.3.
In various embodiments, the substrate comprises Si, SiO2 on Si, or Al2O3 on Si.
In various embodiments, the pair of conducting electrodes comprise at least one of Au, Ag, Pd, Pt, Ru, Rh, Al, Cu, Ni, or alloys thereof.
In various embodiments, the Si, or doped Si-semiconductor layer comprises single crystalline silicon, n-type doped silicon, or p-type doped silicon.
In various embodiments, the width of the nanogap is from about 20 nm to about 100 nm, and wherein the width of the exposed portion of the TMD, Si, or doped Si-semiconductor layer is about 5 nm.
In various embodiments, the enzyme molecule comprises a DNA polymerase or an RNA polymerase.
In various embodiments, a DNA or RNA sequencing device comprises: a pair of electrodes disposed on a substrate, the pair comprising a first electrode and a second electrode spaced-apart from the first electrode by a nanogap having a width W1; a first nanopillar electrically attached to the first electrode and a second nanopillar electrically attached to the second electrode, wherein the two nanopillars are separated by a width W2 and wherein W2≥W1; a dielectric layer disposed over the electrode pair and over the nanogap to surround the nanopillars such that only a top surface of each nanopillar is exposed; a TMD, Si, or doped Si-semiconductor layer disposed on the dielectric layer and over the exposed top surface of each nanopillar, electrically connecting the first and second nanopillars; an enzyme molecule attached to a region of the TMD, Si, or doped Si-semiconductor layer between the nanopillars and directly over the nanogap; and a microfluidic system encasing the electrode pair, the TMD, Si, or doped Si-semiconductor layer and the enzyme molecule attached thereto.
In various embodiments, the TMD layer comprises at least one TMD having a structure MS(2−x), MS(2+x), MSe(2−x), MSe(2+x), MTe(2−x) or MTe(2+x), wherein x is 0-0.3, and M is Mo, W, Ti, Zr, Hf, V, Nb, Ta, Tc, Re, Co, Rh, Ir, Ni, Pd, or Pt.
In various embodiments, the TMD layer comprises a TMD selected from the group consisting of MoS2, WS2, TiS2, ZrS2, HfS2, VS2, NbS2, TaS2, TcS2, ReS2, CoS2, RhS2, IrS2, NiS2, PdS2, PtS2, MoSe2, WSe2, TiSe2, ZrSe2, HfSe2, VSe2, NbSe2, TaSe2, TcSe2, ReSe2, CoSe2, RhSe2, IrSe2, NiSe2, PdSe2, PtSe2, MoTe2, WTe2, TiTe2, ZrTe2, HfTe2, VTe2, NbTe2, TaTe2, TcTe2, ReTe2, CoTe2, RhTe2, IrTe2, NiTe2, PdTe2, PtTe2, and mixtures thereof.
In various embodiments, the TMD layer comprises a mixed TMD compound selected from the group consisting of Mo(SxSeyTez)2, W(SxSeyTez)2, Ti(SxSeyTez)2, Zr(SxSeyTez)2, Hf(SxSeyTez)2, V(SxSeyTez)2, Nb(SxSeyTez)2, Ta(SxSeyTez)2, Te(SxSeyTez)2, Re(SxSeyTez)2, Co(SxSeyTez)2, Rh(SxSeyTez)2, Ir(SxSeyTez)2, Ni(SxSeyTez)2, Pd(SxSeyTez)2, Pt(SxSeyTez)2, and mixtures thereof, wherein (x+y+z) is 0.7-1.3.
In various embodiments, the TMD layer comprises at least one TMD compound of structure M(1−w)NyX(2−z)Yz, wherein M is Al, Si, Ga, Ge, In, Sn, Sb, Bi, Na, K, Ca, Mg, Sr, or Ba; X is S, Se, or Te; Y is Li, B, C, N, O, P, F, Cl, or I; w is 0-0.3; and z is 0-0.3.
In various embodiments, the dielectric layer comprises PMMA, SiO2 or HSQ.
In various embodiments, the first and second electrodes comprise at least one of Au, Ag, Pd, Pt, Ru, Rh, Al, Cu, Ni, or alloys thereof, and wherein the first and second nanopillars comprise Au, Pd, Pt, Ru, or Rh.
In various embodiments, the Si, or doped Si-semiconductor layer comprises single crystalline silicon, n-type doped silicon, or p-type doped silicon.
In various embodiments, W1 is from about 5 nm to about 20 nm, and W2 is from about 5 nm to about 100 nm.
In various embodiments, the enzyme molecule comprises a DNA polymerase or an RNA polymerase.
The features and advantages of embodiments of the present disclosure will become more apparent from the detailed description set forth below when taken in conjunction with the drawings. The drawings and the associated descriptions are provided to illustrate embodiments of the disclosure and not to limit the scope of what is claimed.
Various aspects of the present disclosure generally provide sequencing apparatus, device structures, and methods for using two-dimensional, layer structured semiconductors usable to provide DNA and genome analysis performance. Various disclosed embodiments are amenable to scale-up processes in commercial manufacturing.
In various embodiments of the present disclosure, a biomolecular sensor comprises a nanobridge connected to spaced-apart electrodes and suspended over a gap between them. In various embodiments, the nanobridge comprises a transition metal dichalcogenide material or a silicon material. In various embodiments, a silicon material herein may include pure crystalline silicon or any type of doped silicon semiconductor material. Such materials may be obtained from silicon-on-insulator wafers.
Two dimensional (2D) layered materials such as transition metal dichalcogenides (TMDs) materials and devices have received much attention in recent years by virtue of their unique electronic, physical and chemical properties. One example is molybdenum dichalcogenide MoS2 which can be incorporated as a sensor device. MoS2 type 2D materials can be a single layered material or several layered material. The 2D layer materials such as MoS2 can be produced by various known techniques, e.g., by isolation of very thin MoS2 layer through mechanical exfoliation, physical or chemical vapor deposition, molecular beam epitaxy (MBE) type construction, or sulfurization of a transition metal layer such as Mo or W.
As used herein, the term “nucleotide” means either the native dNTPs like A, T, C, G (i.e., dATP, dTTP, dCTP and dGTP), or collectively refers to various types of modified dNTPs.
As used herein, the term “polymerase” means an enzyme that synthesizes long chains or polymers of nucleic acids. For example, DNA polymerase and RNA polymerase can copy a DNA or RNA template strand, respectively, using base-pairing interactions, thus assembling DNA and RNA molecules.
In various embodiments, a TMD layer is incorporated as a part of sensor bridge structure to attach an enzyme type biomolecule to attract various types of nucleotides for electronic detection signals.
Two dimensional transition metal dichalcogenide (TMD) monolayers are in general atomically thin semiconductors of the type MX2, with M a transition metal atom (notably including Mo, W, or Ti, Zr, Hf, V, Nb, Ta, Tc, Re, Co, Rh, Ir, Ni, Pd, Pt) and X a chalcogen atom (such as S, Se, or Te.). One layer of M atoms is sandwiched between two layers of X atoms. Both the transition metal and the chalcogenide element can be partly replaced (or doped) with other elements. Therefore, the two dimensional TMD layer incorporated into the molecular sensor bridge construction can have various modified or altered composition ranges, including the following:
(i) MoS2, WS2, or TiS2, ZrS2, HfS2, VS2, NbS2, TaS2, TcS2, ReS2, CoS2, RhS2, IrS2, NiS2, PdS2, PtS2 and their modifications or combinations, including modified stoichiometry of sulfur contents having MX(2−x) or MX(2+x) with the desired value of x in the range of 0-1.0, preferably in the range of 0-0.5, even more preferably 0-0.3. For various embodiments, the sulfur stoichiometry is intentionally altered so as to provide vacancy defects, interstitial defects, and aggregated defects so as to increase the surface energy and enhance the adhesion of biomolecule to the bridge sensor. Such defects can also increase the bandgap for stronger sensor signals;
(ii) MoSe2, WSe2, or TiSe2, ZrSe2, HfSe2, VSe2, NbSe2, TaSe2, TcSe2, ReSe2, CoSe2, RhSe2, IrSe2, NiSe2, PdSe2, PtSe2 and their modifications or combinations, including modified stoichiometry of selenium contents having MX(2−x) or MX(2+x) with the desired value of x in the range of 0-1.0, preferably in the range of 0-0.5, even more preferably 0-0.3. For various embodiments, the selenium stoichiometry is intentionally altered so as to provide vacancy defects, interstitial defects, and aggregated defects so as to increase the surface energy and enhance the adhesion of biomolecule to the bridge sensor. Such defects can also increase the bandgap for stronger sensor signals;
(iii) MoTe2, WTe2, or TiTe2, ZrTe2, HfTe2, VTe2, NbTe2, TaTe2, TcTe2, ReTe2, CoTe2, RhTe2, IrTe2, NiTe2, PdTe2, PtTe2 and their modifications or combinations, including modified stoichiometry of tellurium contents having MX(2−x) or MX(2+x) with the desired value of x in the range of 0-1.0, preferably in the range of 0-0.5, even more preferably 0-0.3. For various embodiments, the tellurium stoichiometry is intentionally altered so as to provide vacancy defects, interstitial defects, and aggregated defects so as to increase the surface energy and enhance the adhesion of biomolecule to the bridge sensor. Such defects can also increase the bandgap for stronger sensor signals;
(iv) Mixed TMD compounds in which the MX2 compound has mixed metals and/or mixed chalcogenide. For example Mo(SxSeyTez)2, W(SxSeyTez)2, or Ti(SxSeyTez)2, Zr(SxSeyTez)2, Hf(SxSeyTez)2, V(SxSeyTez)2, Nb(SxSeyTez)2, Ta(SxSeyTez)2, Tc(SxSeyTez)2, Re(SxSeyTez)2, Co(SxSeyTez)2, Rh(SxSeyTez)2, Ir(SxSeyTez)2, Ni(SxSeyTez)2, Pd(SxSeyTez)2, Pt(SxSeyTez)2 where the combined (x+y+z) is 1-3, preferably 0.5-1.5, even more preferably 0.7-1.3. Alternatively, two or more metals can be combined for sulfur containing, Se-containing or Te-containing TMD layers, e.g., (MoxWyCoz)S2, (HfxWyCoz)Te2 and so forth; or
(v) M(1−w)NyX(2−z)Yz structure in which the transition metal M is partially substituted with non-transition elements N, with a concentration of w and the N element selected from one or more of Al, Si, Ga, Ge, In, Sn, Sb, Bi, Al, Na, K, Ca, Mg, Sr, Ba, with the w value in the range of 0-0.3, and the chalcogenide element X partially substituted with a non-chalcogenide element Y, with the Y element selected from one or more of Li, B, C, N, O, P, F, Cl, I, with the z value in the range of 0-0.3.
In various embodiments, the thickness of a MoS2 monolayer can be about 6.5 Å. The TMD materials in their simplest monolayer structure, e.g., MoS2, WS2, MoSe2, WSe2, MoTe2, have a direct band gap, and hence can be used in electronics as transistors or sensors. Either monolayer TMD or few layer TMD can be structurally modified, to be utilized as solid state DNA or genome sensors, without labeling with optical capability. Being an ultrathin direct bandgap semiconductor, a transition metal dichalcogenide such as single-layer MoS2 has found some useful applications in nanoelectronics, optoelectronics, and energy harvesting. However, not many sensor applications have been attempted or demonstrated with proper characteristics, especially for DNA or genome sequencing purposes.
In various embodiments, a biosensor herein may comprise a nanobridge made of a silicon material rather than a TMD material. As used herein, “Si-material” or “Si-type” bridge, nanobridge or layer may comprise pure crystalline silicon or any type of doped silicon semiconductor material, as described herein.
In various embodiments, a silicon nanobridge may comprise a semiconductor wire or a ribbon structure. In various embodiments, a silicon nanobridge may comprise material derived from a silicon-on-insulator (SOI) wafer, and may take the form of a nanostructure or related semiconductor nanoribbon or nanowire structure.
A single nanobridge structure of crystalline silicon may be obtained from silicon-on-insulator (SOI) semiconductor substrate (or other related semiconductor layers), which can be made into a reliable nanobridge sensor structure since the material is already properly-doped semiconductor films with such a nanobridge material being less sensitive to nanopatterning-related damages. Therefore, a need for high temperature processing is minimized. Unique SOI-based fabrication of nanobridge structures, processing methods and applications of such a single-bridge SOI-based nanoribbon biosensors are disclosed in the drawing figures.
The thickness of a doped SOI Si thin film nanobridge is desirably thin, e.g., <30 nm, preferably <20 nm thick, more preferably <10 nm thick layer, and even more preferably <5 nm, with a width of <40 nm, preferably <20 nm, more preferably <10 nm, and even more preferably <3 nm. A smaller cross-section nanowire or nanoribbon bridge across two mating electrodes can provide higher signal to noise ratio during sequencing interactions.
Also, a smaller cross-section nanowire or nanoribbon bridge of SOI-based Si nanobridge or crystalline semiconductors in general (e.g., <10 nm width, preferably <5 nm width, more preferably 3 nm width), ensures that only a single (or at most a few) nanobridge structures will attach to form one or at most a few bridges, due to size exclusion, instead of multiple bridges, with the latter introducing multiple, undesirably complicated signals from the parallel bridges stuck together on the same electrode lead pair. A single and narrower sensor bridge will also prefer a single-molecule polymerase placement for each electrode pair.
Substantially narrowed SOI-Si structure for a single nanobridge can be prepared using highly advanced nanofabrication of Si and SiO2 structures. Another approach to further ensure a single bridge formation of SOI-Si, according to various aspects of the present disclosure, is to utilize a nano-mask array, for example, by placing an array of e.g., <5 nm wide graphene, carbon, ceramic or metallic nanoribbon mask so that the SOI-Si underneath can be patterned and shaped to become narrow nanobridges. Such pre-made narrowed graphene, carbon, ceramic or metallic nanoribbon mask can be prepared on another substrate followed by stamp transfer using a PMMA (Polymethyl methacrylate), PDMS (Polydimethylsiloxane) or other polymer type stamp materials. Such pre-made narrowed bridge can be used as a sensor bridge on which a single enzyme molecule such as polymerase can be attached.
In various embodiments, such SOI-Si type or other semiconductor ribbon strips can be split into two nanobridges between which another molecular bridge (such as a single DNA or peptide) can be attached for ease of polymerase bonding and nucleotide analysis. Such splitting of amorphous semiconductor into two separated parts with a nanogap in between (e.g., 20-100 nm) can be accomplished by e.g., focused laser beam slicing, focused ion beam cutting or patterning and etching. The ends of the split ribbons facing each other can desirably be sharpened to a pointed-tip geometry of e.g., 2-5 nm radius of curvature, so as to facilitate an attachment of a single DNA or a single peptide molecular bridge, using either electric field alignment, flow alignment in a microfluidic chamber, or stamp transfer of pre-aligned DNA or peptide. Stamp transfer of pre-aligned DNA or peptide array can be made using PMMA, PDMS or other polymer type soft stamps.
Disclosed herein are label-free DNA or RNA sequencing device structures utilizing a TMD- or Si-based frame with an enzyme polymerase for detection of electronic signals when an individual nucleotide is attached onto a nucleic acid template. In various embodiments, two dimensional semiconductors of processed, defective or nanoporous Transition Metal Dichalcogenide (TMD) layer material are employed so as to utilize altered bandgaps of the TMD layer and enhanced attachment of single biomolecules. In various embodiments, the TMD-based sequencing systems disclosed herein can be assembled into a massively parallel configuration for rapid analysis of targets including nucleotides, in particular for applications of sequencing of a DNA molecule, or a collection of such molecules constituting an entire genome. Such systems are also useful for DNA-based information storage, for which the writing is performed by encoding specific nucleotide-based arrangements or sequences and the reading is carried out by sequencing analysis using TMD-bridge based molecular sensor array.
A bridge-configured sensor structure comprising an elongated nano-dimension, crystalline semiconductor wire or ribbon, such as silicon or doped silicon, is another way of producing label-free molecular sensors for genome sequencing without introducing complicated fluorescence imaging methodologies. Such semiconductor nanowires of, e.g., made of Si derived from silicon-on-insulator (SOI) wafer, can be connected to a pair of electrodes (with optional gate structure) in high density electronic circuit assembly, also equipped with microfluidic environment comprising floating DNAs, nucleotides, enzyme polymerase molecules, etc. It is possible to attach a polymerase single molecule (or few molecules) to such a nanobridge or elongated biomolecule bridge, using, for example, functionalities and ligands such as biotin-streptavidin, antibody-antigen, pyrene or peptide complexes.
Inorganic nanobridges, either van der Waals force connected or metallization connected to the device electrode leads can offer much higher electrical conductivity and substantially higher electrical sensor signals than organic nanobridges as the use of undesirably high electrical resistance ligands or attaching functionalities can be minimized.
In various embodiments of the present disclosure, biosensor bridges connected between a pair of conducting electrodes (such as made of Au, Ag, Pd, Pt, Ru, Rh, Al, Cu, Ni, etc.), can be accomplished by an active attachment route using electrical field (electrophoretic alignment and attachment using various AC or DC mode of electric fields, often at a level of 0.2 to 10 volts). Alternatively, aligned nanobridge structures connecting the mating electrodes can also be achieved by stamp transfer of pre-aligned (or pre-patterned) inorganic nanobridge array from another substrate using PMMA, PDMS or other stamp materials. On the device substrate or temporary preparation substrate, the nanobridge array in parallelly aligned configuration is prepared either by nanopatterning by lithographic means, flow alignment in a microfluidic chamber, or by electrical field alignment (e.g., dielectrophoretic alignment using AC or DC electric field).
While a number of nanomaterials can be considered for such nanobridge structures, these nanomaterials need to be processed into nano-dimension wires or ribbons by nanolithography or other treatments, which tends to cause damage to crystallographic structures or disruption of atomic arrangements causing unintended changes or deterioration of physical or electronic properties. While a high temperature annealing process (e.g., 500° to 1000° C.) can sometimes repair or reduce such damages, the use of high temperature processing steps often damages electronic device structure and hence should be avoided for ease of device manufacturing and reliability.
For accurate signal detection on nucleotide attachment events (or other biomolecule attachment events, e.g., to polymerase, to enable electrical signal detection for genome sequencing), it is highly desirable to provide a single elongated bridge (made of inorganic or organic nanowires or nanoribbons) between mating electrodes (made of Au, Pd or other conducting lead wires). If multiple nanobridges are attached between the two mating electrodes, often clumped together, multiple signals may occur by the presence of parallel sensors, which makes the analysis of such complicated signals very difficult.
DNA data storage is a process of digital encoding and decoding binary data, to and from synthesized or duplicated DNA strands. For example, the binary code information storage of (00), (01), (10) and (11) can be replaced by various arrangements of oligonucleotides (A, C, G, T). DNA molecules are genetic blueprints for living organisms, and the information stored in DNA is known to last more than 10,000 years under certain environment. With its huge capacity (many orders of magnitude larger than what is possible with current technology) to store enormous amount of information in very small space, DNA storage could be the answer to a modern era problem of too much information that needs to stored, e.g., on the order of hundreds of zettabytes every year in the near future. Currently available information storage capability including magnetic disk, tape, optical or other related technologies can cover only a fraction of such a needed capacity.
While substantial progress has been made in DNA information storage in recent years, cost effective data storage techniques for practical applications are yet to be achieved. For efficient retrieval of stored information, the encoded DNA nucleotide arrangements need to be decoded by, e.g., reading (or sequence analysis). A fast, economic method of reading the encoded DNA information is essential for the success of DNA based data storage. This disclosure also provides new methods and device structure to enable such progress.
Biosensors having a TMD or silicon nanobridge provide reliable DNA genome analysis performance and are more easily amenable to scalable manufacturing, because the need for high temperature processing is minimized. The disclosed structures herein are also useful for DNA-based large-capacity information storage devices including archival or randomly-accessible-memory and logic devices.
In various embodiments, a sequencing device is provided comprising: (a) an electrode array of conducting electrode pairs, each pair of electrodes comprising a source and a drain electrode separated by a nanogap, said electrode array deposited and patterned on a dielectric substrate, and optionally comprising a third electrode as a gated system; (b) a single-layer or few-layer thick transition metal dichalcogenide (TMD) layer disposed on each pair of electrodes, connecting each source and drain electrode within each pair, and bridging each nanogap of each pair; (c) a dielectric masking layer disposed on the TMD layer and comprising size-limited openings that define exposed TMD regions, each opening sized so as to allow only a single enzyme biomolecule to fit therein and to attach onto the exposed TMD region defined by each opening; (d) an enzyme molecule attached to each exposed TMD region such that only one enzyme molecule is found within each opening; and (d) a microfluidic system encasing the electrode array, wherein attachment or detachment of a biomolecule selected from the group consisting of a nucleotide monomer, a protein, and a DNA segment, onto the enzyme molecule, one at a time, can be monitored as a uniquely identifiable electrical signal pulse to determine the specific nature of the biomolecule attaching or detaching.
In various embodiments, the dielectric substrate comprises SiO2. In various embodiments, the dielectric substrate comprises SiO2 or Al2O3.
In various embodiments, the TMD is selected from MoS2, WS2, TiS2, ZrS2, HfS2, VS2, NbS2, TaS2, TcS2, ReS2, CoS2, RhS2, IrS2, NiS2, PdS2, PtS2 and their modifications or combinations. In various embodiments, the TMD is MoS2. In various embodiments, the TMD is WS2.
In various embodiments, the TMD is selected from MoS2, WS2, TiS2, ZrS2, HfS2, VS2, NbS2, TaS2, TcS2, ReS2, CoS2, RhS2, IrS2, NiS2, PdS2, PtS2 and their modifications or combinations, including modified stoichiometry of sulfur contents having MX(2−x) or MX(2+x) wherein x is in the range of 0-1.0, preferably in the range of 0-0.5, even more preferably 0-0.3. In various embodiments, the TMD is selected from MoS2, WS2, TiS2, ZrS2, HfS2, VS2, NbS2, TaS2, TcS2, ReS2, CoS2, RhS2, IrS2, NiS2, PdS2, PtS2 and their modifications or combinations and the stoichiometry of sulfur is not modified.
In various embodiments, the TMD is selected from MoSe2, WSe2, or TiSe2, ZrSe2, HfSe2, VSe2, NbSe2, TaSe2, TcSe2, ReSe2, CoSe2, RhSe2, IrSe2, NiSe2, PdSe2, PtSe2 and their modifications or combinations, including modified stoichiometry of selenium contents having MX(2−x) or MX(2+x) with the desired value of x in the range of 0-1.0, preferably in the range of 0-0.5, even more preferably 0-0.3. In various embodiments, the TMD is selected from MoSe2, WSe2, or TiSe2, ZrSe2, HfSe2, VSe2, NbSe2, TaSe2, TcSe2, ReSe2, CoSe2, RhSe2, IrSe2, NiSe2, PdSe2, PtSe2 and their modifications or combinations and the stoichiometry of selenium is not modified.
In various embodiments, defects are artificially introduced into TMD. In various embodiments, the defects are introduced to increase bandgap. In various embodiments, the defects are introduced to provide active site edge locations for strong adhesion of bridge structures or biomolecules such as enzyme molecules.
In various embodiments, TMD is selected from MoTe2, WTe2, or TiTe2, ZrTe2, HfTe2, VTe2, NbTe2, TaTe2, TcTe2, ReTe2, CoTe2, RhTe2, IrTe2, NiTe2, PdTe2, PtTe2 and their modifications or combinations.
In various embodiments, TMD is selected from MoTe2, WTe2, or TiTe2, ZrTe2, Hffe2, VTe2, NbTe2, TaTe2, TcTe2, ReTe2, CoTe2, RhTe2, IrTe2, NiTe2, PdTe2, PtTe2 and their modifications or combinations, including modified stoichiometry of Tellurium contents having MX(2−x) or MX(2+x) with the desired value of x in the range of 0-1.0, preferably in the range of 0-0.5, even more preferably 0-0.3. In various embodiments, TMD is selected from MoTe2, WTe2, or TiTe2, ZrTe2, Hffe2, VTe2, NbTe2, TaTe2, TcTe2, ReTe2, CoTe2, RhTe2, IrTe2, NiTe2, PdTe2, PtTe2 and their modifications or combinations and the stoichiometry of tellurium is not modified.
In various embodiments, the tellurium stoichiometry is intentionally altered so as to provide vacancy defects, interstitial defects, and aggregated defects in order to increase the surface energy of the TMD layer and enhance the adhesion of biomolecule to the bridge sensor for stronger sensor signals.
In various embodiments, the TMD comprises a mixed TMD selected from TMD compounds in which the MX2 compound has mixed metals and/or mixed chalcogenide, selected from the group consisting of Mo(SxSeyTez)2, W(SxSeyTez)2, Ti(SxSeyTez)2, Zr(SxSeyTez)2, Hf(SxSeyTez)2, V(SxSeyTez)2, Nb(SxSeyTez)2, Ta(SxSeyTez)2, Tc(SxSeyTez)2, Re(SxSeyTez)2, Co(SxSeyTez)2, Rh(SxSeyTez)2, Ir(SxSeyTez)2, Ni(SxSeyTez)2, Pd(SxSeyTez)2, and Pt(SxSeyTez)2 wherein the combined (x+y+z) is 1-3, 0.5-1.5, or 0.7-1.3.
In various embodiments, two or more metals are combined for sulfur containing, Se-containing or Te-containing TMD layers.
In various embodiments, the TMD layer comprises (MoxWyCoz)S2 or (HfxWyCoz)Te2.
In various embodiments, the TMD comprises a M(1−w)NyX(2−z)Yz structure in which the transition metal M is partially substituted with non-transition elements N, with a concentration of w and the N element selected from one or more of Al, Si, Ga, Ge, In, Sn, Sb, Bi, Al, Na, K, Ca, Mg, Sr, Ba, with the w value in the range of 0-0.3, and the chalcogenide element X partially substituted with a non-chalcogenide element Y, with the Y element selected from one or more of Li, B, C, N, O, P, F, Cl, I, with the z value in the range of 0-0.3. In various embodiments, the w value is greater than 0.3, for example, greater than 0.4, greater than 0.5, greater than 0.6, greater than 0.7, greater than 0.8, greater than 0.9, or greater than 1.0. In various embodiments, the z value is greater than 0.3, for example, greater than 0.4, greater than 0.5, greater than 0.6, greater than 0.7, greater than 0.8, greater than 0.9, or greater than 1.0.
In various embodiments, a sequencing device is provided comprising: (a) an electrode array of conducting electrode pairs, each pair of electrodes comprising a source and a drain electrode separated by a nanogap, said electrode array deposited and patterned on a dielectric substrate, and optionally comprising a third electrode as a gated system; (b) a single-layer or few-layer thick silicon or doped silicon semiconductor layer disposed on each pair of electrodes, connecting each source and drain electrode within each pair, and bridging each nanogap of each pair; (c) a dielectric masking layer disposed on the silicon layer and comprising size-limited openings that define exposed silicon regions, each opening sized so as to allow only a single enzyme biomolecule to fit therein and to attach onto the exposed silicon region defined by each opening; (d) an enzyme molecule attached to each exposed silicon region such that only one enzyme molecule is found within each opening; and (d) a microfluidic system encasing the electrode array, wherein attachment or detachment of a biomolecule selected from the group consisting of a nucleotide monomer, a protein, and a DNA segment, onto the enzyme molecule, one at a time, can be monitored as a uniquely identifiable electrical signal pulse to determine the specific nature of the biomolecule attaching or detaching.
In various embodiments, the electrodes in each electrode pair may comprise Au, Ag, Pd, Pt, Ru, Rh, Al, Cu, Ni, etc.
In various embodiments, the dielectric substrate comprises SiO2. In various embodiments, the dielectric substrate comprises SiO2 or Al2O3.
In various embodiments, the silicon material forming the nanobridge comprises pure crystalline silicon, such as single crystalline silicon.
In various embodiments, the silicon material forming the nanobridge comprises an n- or p-doped silicon semiconductor material. Dopants for n-type or p-type doped silicon semiconductor may comprise acceptors from Group III elements or donors from Group V elements. Dopants for n- or p-doped silicon semiconductor include, but are not limited to As, B, P, Sb, Ga, Zn, and Fe.
In various embodiments, a silicon nanobridge may be obtained from a SOI wafer manufactured with the desired silicon material (e.g., single crystalline silicon, n-type silicon semiconductor, or p-type silicon semiconductor).
Various aspects of the present disclosure provide biosensor structures, materials and geometries, as well as fabrication methods and application methods, such as described below in reference to the various drawing figures.
In various embodiments, the metallic conducting electrode pair is selected from Au, Ag, Pd, Pt, Ru, Rh, Al, Cu, Ni, or their alloys. In various embodiments, the conducting electrode pair 3 may comprise Al, Cu, Ru, Pt or Pd, whereas the pillars 6 may comprise Ru, Pt, or Pd.
In various embodiments, the nanogap 1 is from about 5 nm to about 20 nm. In various embodiments, the nanogap is less than 5 nm, for example less than 3 nm, or less than 1.0 nm. In various embodiments, the nanogap is greater than 20 nm, for example, greater than 25 nm, greater than 30 nm, greater than 35 nm, greater than 40 nm, greater than 45 nm, or greater than 50 nm.
In various embodiments, the size-limiting openings 5 are preferably less than 30 nm average equivalent diameter each, more preferably less than 20 nm equivalent diameter, even more preferably less than 10 nm equivalent diameter. The openings 5 can be fashioned by lithographic or nanofabrication defined coverage of a dielectric material layer, polymer or ceramic in all the areas outside a specific, size-limited region intended for attaching only a single molecule. The term “average equivalent diameter” is used when the openings 5 are not circular, but where the calculated surface area can be expressed as though the openings are circular. Polymerase molecules, as well as streptavidin-type linked molecules, often have a steric size on the order of ˜5 nm regime. With the openings 5 thus configured, only one biomolecule fits within each opening to attach to the exposed TMD or Si-material inside the opening.
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In various embodiments, a single-molecule biosensor comprises: a conducting electrode pair disposed on a substrate, the electrode pair comprising a first electrode and a second electrode spaced-apart from the first electrode by a nanogap; a contiguous transition metal dichalcogenide (TMD) layer disposed on the first electrode and on the second electrode, the TMD layer configured as a bridge suspended over the nanogap; an enzyme molecule attached to a region of the TMD layer suspended over the nanogap; and a microfluidic system encasing the conducting electrode pair, the contiguous TMD layer and the enzyme molecule attached thereto.
In various embodiments, the TMD layer comprises a TMD selected from the group consisting of MoS2, WS2, TiS2, ZrS2, HfS2, VS2, NbS2, TaS2, TcS2, ReS2, CoS2, RhS2, IrS2, NiS2, PdS2, PtS2, and mixtures thereof.
In various embodiments, the TMD layer comprises a TMD selected from the group consisting of MoSe2, WSe2, TiSe2, ZrSe2, HfSe2, VSe2, NbSe2, TaSe2, TcSe2, ReSe2, CoSe2, RhSe2, IrSe2, NiSe2, PdSe2, PtSe2, and mixtures thereof.
In various embodiments, the TMD layer comprises at least one TMD of structure MSe(2−x) or MSe(2+x), wherein x is 0-0.3, and M is Mo, W, Ti, Zr, Hf, V, Nb, Ta, Tc, Re, Co, Rh, Ir, Ni, Pd, or Pt.
In various embodiments, the TMD layer comprises a TMD selected from the group consisting of MoTe2, WTe2, TiTe2, ZrTe2, HfTe2, VTe2, NbTe2, TaTe2, TcTe2, ReTe2, CoTe2, RhTe2, IrTe2, NiTe2, PdTe2, PtTe2, and mixtures thereof.
In various embodiments, the TMD layer comprises a mixed TMD compound selected from the group consisting of Mo(SxSeyTez)2, W(SxSeyTez)2, Ti(SxSeyTez)2, Zr(SxSeyTez)2, Hf(SxSeyTez)2, V(SxSeyTez)2, Nb(SxSeyTez)2, Ta(SxSeyTez)2, Tc(SxSeyTez)2, Re(SxSeyTez)2, Co(SxSeyTez)2, Rh(SxSeyTez)2, Ir(SxSeyTez)2, Ni(SxSeyTez)2, Pd(SxSeyTez)2, Pt(SxSeyTez)2, and mixtures thereof, wherein (x+y+z) is 0.7-1.3.
In various embodiments, the TMD layer comprises at least one TMD compound of structure M(1−w)NyX(2−z)Yz, wherein M is Al, Si, Ga, Ge, In, Sn, Sb, Bi, Na, K, Ca, Mg, Sr, or Ba; X is S, Se, or Te; Y is Li, B, C, N, O, P, F, Cl, or I; w is 0-0.3; and z is 0-0.3.
In various embodiments, the substrate comprises SiO2 or Al2O3.
In various embodiments, the pair of conducting electrodes comprise at least one of Au, Pt, Ag, Pd, Rh, Ru, or alloys thereof.
In various embodiments, the nanogap is from about 1 nm to about 50 nm.
In various embodiments, the single-molecule biosensor further comprises a dielectric, ceramic or polymer coating layer disposed on the TMD layer on a side opposite the first and second electrodes, wherein the dielectric, ceramic or polymer coating layer includes an opening on the region of the TMD layer suspended over the nanogap, the opening leaving an exposed portion of the TMD layer therein, the opening dimensioned to accommodate only one enzyme molecule.
In various embodiments, the dielectric, ceramic or polymer layer comprises PMMA or SiO2.
In various embodiments, the enzyme molecule comprises a DNA polymerase or an RNA polymerase.
In various embodiments, the enzyme molecule is attached to the TMD layer via a biotin-streptavidin linkage.
In various embodiments, the TMD layer includes as least one of vacancy defects, interstitial defects, and aggregated defects.
In various embodiments of the present disclosure, a DNA or RNA sequencing device comprises: an electrode array of conducting electrode pairs disposed on a substrate, each pair of electrodes comprising a source electrode and a drain electrode spaced-apart from the source electrode by a nanogap; a contiguous transition metal dichalcogenide (TMD) layer disposed on the first electrode and on the second electrode, the TMD layer configured as a bridge suspended over the nanogap; a polymerase enzyme molecule attached to a region of the TMD layer over the nanogap; and a microfluidic system encasing the conducting electrode pair, the contiguous TMD layer and the enzyme molecule attached thereto.
In various embodiments, the TMD layer comprises at least one TMD of structure MX(2−x) or MX(2+x), wherein X is S, Se or Te; x is 0-0.3; and M is Mo, W, Ti, Zr, Hf, V, Nb, Ta, Tc, Re, Co, Rh, Ir, Ni, Pd, or Pt.
In various embodiments, a single-molecule biosensor comprises: a conducting electrode pair disposed on a substrate, the electrode pair comprising a first electrode and a second electrode spaced-apart from the first electrode by a nanogap; a contiguous Si-material layer disposed on the first electrode and on the second electrode, the Si-material layer configured as a bridge suspended over the nanogap; an enzyme molecule attached to a region of the Si-material layer suspended over the nanogap; and a microfluidic system encasing the conducting electrode pair, the contiguous Si-material layer and the enzyme molecule attached thereto.
In various embodiments, the Si-material layer comprises a crystalline silicon, a p-type doped silicon semiconductor material or an n-type doped silicon semiconductor material.
In various embodiments, the substrate comprises SiO2 or Al2O3.
In various embodiments, the pair of conducting electrodes comprise at least one of Au, Pt, Ag, Pd, Rh, Ru, or alloys thereof.
In various embodiments, the nanogap is from about 1 nm to about 50 nm.
In various embodiments, the single-molecule biosensor further comprises a dielectric, ceramic or polymer coating layer disposed on the Si-material layer on a side opposite the first and second electrodes, wherein the dielectric, ceramic or polymer coating layer includes an opening on the region of the Si-material layer suspended over the nanogap, the opening leaving an exposed portion of the Si-material layer therein, the opening dimensioned to accommodate only one enzyme molecule.
In various embodiments, the dielectric, ceramic or polymer layer comprises PMMA or SiO2.
In various embodiments, the enzyme molecule comprises a DNA polymerase or an RNA polymerase.
In various embodiments, the enzyme molecule is attached to the Si-material layer via a biotin-streptavidin linkage.
In various embodiments, the Si-material layer was obtained from a silicon-on-insulator (SOI) wafer.
In various embodiments, the substrate and Si-Material layer comprise a silicon-on-insulator (SOI) wafer.
In various embodiments of the present disclosure, a DNA or RNA sequencing device comprises: an electrode array of conducting electrode pairs disposed on a substrate, each pair of electrodes comprising a source electrode and a drain electrode spaced-apart from the source electrode by a nanogap; a contiguous Si-material layer disposed on the first electrode and on the second electrode, the Si-material layer configured as a bridge suspended over the nanogap; a polymerase enzyme molecule attached to a region of the Si-material layer over the nanogap; and a microfluidic system encasing the conducting electrode pair, the contiguous Si-material layer and the enzyme molecule attached thereto.
In the detailed description, references to “various embodiments”, “one embodiment”, “an embodiment”, “an example embodiment”, etc., indicate that the embodiment described may include a particular feature, structure, or characteristic, but every embodiment may not necessarily include the particular feature, structure, or characteristic. Moreover, such phrases are not necessarily referring to the same embodiment. Further, when a particular feature, structure, or characteristic is described in connection with an embodiment, it is submitted that it is within the knowledge of one skilled in the art to affect such feature, structure, or characteristic in connection with other embodiments whether or not explicitly described. After reading the description, it will be apparent to one skilled in the relevant art(s) how to implement the disclosure in alternative embodiments.
Steps recited in any of the method or process descriptions may be executed in any order and are not necessarily limited to the order presented. Furthermore, any reference to singular includes plural embodiments, and any reference to more than one component or step may include a singular embodiment or step. Also, any reference to attached, fixed, connected, coupled or the like may include permanent (e.g., integral), removable, temporary, partial, full, and/or any other possible attachment option. Any of the components may be coupled to each other via friction, snap, sleeves, brackets, clips or other means now known in the art or hereinafter developed. Additionally, any reference to without contact (or similar phrases) may also include reduced contact or minimal contact.
Benefits, other advantages, and solutions to problems have been described herein with regard to specific embodiments. However, the benefits, advantages, solutions to problems, and any elements that may cause any benefit, advantage, or solution to occur or become more pronounced are not to be construed as critical, required, or essential features or elements of the disclosure. The scope of the disclosure is accordingly to be limited by nothing other than the appended claims, in which reference to an element in the singular is not intended to mean “one and only one” unless explicitly so stated, but rather “one or more.” Moreover, where a phrase similar to ‘at least one of A, B, and C’ or ‘at least one of A, B, or C’ is used in the claims or specification, it is intended that the phrase be interpreted to mean that A alone may be present in an embodiment, B alone may be present in an embodiment, C alone may be present in an embodiment, or that any combination of the elements A, B and C may be present in a single embodiment; for example, A and B, A and C, B and C, or A and B and C.
All structural, chemical, and functional equivalents to the elements of the above-described various embodiments that are known to those of ordinary skill in the art are expressly incorporated herein by reference and are intended to be encompassed by the present claims. Moreover, it is not necessary for an apparatus or component of an apparatus, or method in using an apparatus to address each and every problem sought to be solved by the present disclosure, for it to be encompassed by the present claims. Furthermore, no element, component, or method step in the present disclosure is intended to be dedicated to the public regardless of whether the element, component, or method step is explicitly recited in the claims. No claim element is intended to invoke 35 U.S.C. 112(f) unless the element is expressly recited using the phrase “means for.” As used herein, the terms “comprises”, “comprising”, or any other variation thereof, are intended to cover a non-exclusive inclusion, such that a chemical, chemical composition, process, method, article, or apparatus that comprises a list of elements does not include only those elements but may include other elements not expressly listed or inherent to such chemical, chemical composition, process, method, article, or apparatus.
This application is a continuation-in-part of U.S. patent application Ser. No. 16/840,755 filed Apr. 6, 2020, entitled “Method, Apparatus and System for Single-Molecule Polymerase Biosensor with Transition Metal Nanobridge.” This application also claims priority to, and the benefit of, U.S. Provisional Patent Application Ser. No. 62/954,292 filed Dec. 27, 2019, entitled “Pre-Aligned and Transferred, SOI Based Nano-Bridge Biosensor Array and Memory, Comprising Single Biomolecule Sensor, and Method and Uses Thereof.” The '755 Application claims priority to, and the benefit of, U.S. Provisional Patent Application Ser. No. 62/830,231 filed Apr. 5, 2019, entitled “Single-Molecule Polymerase Biosensor Comprising Transition Metal Dichalcogenide Nanobridge for Sequencing, Information Storage and Reading.” These disclosures are incorporated herein by reference in their entireties for all purposes.
| Number | Date | Country | |
|---|---|---|---|
| 62954292 | Dec 2019 | US | |
| 62830231 | Apr 2019 | US |
| Number | Date | Country | |
|---|---|---|---|
| Parent | 16840755 | Apr 2020 | US |
| Child | 17135375 | US |
