Systems, Devices, and Processes

Abstract

A system for sequencing a nucleotide chain, the system comprising an apparatus having at least one electrode for measuring an overall electromagnetic signal from a portion of the nucleotide chain comprising a plurality of nucleotides, and a mechanism configured to move the nucleotide chain relative to the electrode to modulate the overall electromagnetic signal to identify electromagnetic signals associated with each individual nucleotide in the plurality of nucleotides.

Description

[Technical Field]

[0001] Related Applications This application is related to Australian Provisional Patent Application No. 2022902309 and Australian Complete Patent Application No. 2023202006, the specifications of which, as originally filed and any amended specifications, are incorporated herein by reference.

[0002] The present disclosure relates to systems, devices, and processes for sequencing nucleotide strands, including sequencing of molecules including nucleic acids, deoxyribonucleic acid (DNA), single-stranded (ssDNA), and / or ribonucleic acid (RNA). [Background technology]

[0003] Nucleic acid sequencing involves determining the order of nucleotides in a nucleotide chain, which is typically a nucleic acid molecule that may be DNA, ssDNA, RNA, and / or a protein / molecule that contains RNA, including determining the order of bases in the nucleotide chain, including adenine (A), guanine (G), cytosine (C), and thymine (T).

[0004] Rapid nucleotide sequencing has greatly accelerated biological and medical research and discovery.

[0005] However, existing systems and processes for nucleotide sequencing may not be fast / accurate / efficient enough for at least some applications.

[0006] It is desired to address or ameliorate one or more of the disadvantages or limitations associated with the prior art, or at least provide a useful alternative. Summary of the Invention

[0007] According to the present invention, there is provided a system for sequencing a nucleotide chain, comprising: at least one electrode for measuring an aggregate electromagnetic signal from a portion of the nucleotide chain comprising a plurality of nucleotides; a mechanism configured to move the nucleotide strand relative to the electrodes to modulate the aggregate electromagnetic signal to identify an electromagnetic signal associated with each individual nucleotide in the plurality of nucleotides; A system is provided, comprising an apparatus having:

[0008] According to the present invention, there is provided a process for sequencing a nucleotide chain, comprising: measuring, using at least one electrode, an overall electromagnetic signal from a portion of the nucleotide chain comprising a plurality of nucleotides; moving the nucleotide strand relative to at least one electrode to modulate the aggregate electromagnetic signal to identify electromagnetic signals associated with individual nucleotides in the plurality of nucleotides; Also provided is a process including: [Brief explanation of the drawings]

[0009] Some embodiments of the present invention will now be described with reference to the accompanying drawings.

[0010] [Figure 1] 1 is a right-side cross-sectional schematic diagram of an apparatus for sequencing nucleotide chains ("sequencer apparatus") according to a first embodiment of the present disclosure. [Figure 2] FIG. 2 is a schematic plan view of an anchor plate in the device according to the first embodiment of the present disclosure. [Figure 3] FIG. 2 is a schematic bottom view of a movable plate in the apparatus according to the first embodiment of the present disclosure. [Figure 4] 1 is a plan view schematic diagram of an actuation straightening device for straightening nucleotide strands (in solution) according to a first embodiment of the present disclosure; FIG. [Figure 5] FIG. 1 is a plan schematic view of a dual-electrode straightening device for straightening a nucleotide chain according to a first embodiment of the present disclosure. [Figure 6]1 is a right-side cross-sectional schematic diagram of an apparatus for sequencing nucleotide chains ("sequencer apparatus") according to a second embodiment of the present disclosure. [Figure 7] FIG. 10 is a schematic bottom view of a movable plate in an apparatus according to a second embodiment of the present disclosure. [Figure 8] FIG. 10 is a schematic diagram of an exemplary electronic system in an apparatus according to a second embodiment of the present disclosure. DETAILED DESCRIPTION OF THE INVENTION

[0011] I. Systems and Sequencing Devices Described herein is a system for sequencing nucleotide chains.

[0012] The system comprises an apparatus (100) for sequencing nucleotide chains ("sequencing apparatus", "sequencer", or "sequencer apparatus").

[0013] The device (100) has at least one electrode (the "sample electrode" or "probe") for measuring an electromagnetic signal (eg, charge, current, and / or voltage) from a portion of a nucleotide chain.

[0014] The device (100) further comprises a mechanism configured to move the nucleotide strand relative to the electrodes to modulate the electromagnetic signal.

[0015] II. First embodiment A first embodiment of a system comprising the device (100) is described below.

[0016] Basic structure of sequencing device As shown in FIG. 1, the sequencer device (100) according to this embodiment includes: a) at least one EM sample electrode (104A including 104Aa, 104Ab, ..., 104An) for measuring an electromagnetic (EM) signal ("sample EM signal") (e.g., charge, current, or voltage) from a portion (i.e., "sample portion") of the nucleotide chain (106A); b) a mechanism configured to move the nucleotide strand (106A) relative to the EM sample electrode (104A) to modulate the EM signal (i.e., a "modulation mechanism"); Equipped with.

[0017] At least one EM sample electrode (104A) may be a non-contact EM sensor, i.e., the sample electrode does not contact or touch the nucleotide strand (106A) during the measurement process.

[0018] The sequencer device (100) may include at least one EM reference electrode (102A, including 102Aa, 102Ab, ..., 102An) configured to generate or measure an EM signal ("reference EM signal") (e.g., charge and / or current) from a portion (i.e., "reference portion") of the nucleotide strand (106A) adjacent to each corresponding reference electrode (102A). The one or more reference electrodes (102A) may be controlled to substantially essentially clamp the nucleotide strand (106A) with nm precision to obtain a reliable electrical signal with an acceptable signal-to-noise ratio (SNR), as described below.

[0019] The EM signals measured may include one or more of the following: a) Fluctuations in the current (conductance) between the reference electrode (102X) and the sample electrode (104X) caused by the effect of different local charge densities for different nucleotides, which may generate dipole moments or modify the potential barrier to quantum tunneling of electrons. These fluctuations may, of course, be constant (DC) at determined frequencies (AC) due to specific molecular resonances, or may cover a broader portion of the frequency spectrum. b) Variation in the current conducted through a portion of the nucleotide chain (106A) from the anchor electrode (112X) to one of the sample electrodes (104Xx). c) A reference current conducted through a portion of the nucleotide chain (106A) from the anchor electrode (112X) to one of the reference electrodes (102Xx). This signal can be monitored to adjust / correct the signal from (b) for external noise sources, thermal drift, or Brownian motion. d) Variation of the current in (a) and (b) for a fixed frequency modulation applied to the anchor plate (200) via the plate actuator (124).

[0020] The modulation mechanism is a) a holder configured to hold / fix the nucleotide strand (106) so that the nucleotide strand (106) is stationary relative to an anchor portion of the sequencer device (100); b) at least one control actuator comprising a sensor actuator (108) and / or an anchor plate actuator (124), configured to move a sensor portion of the sequencer device (100) relative to an anchor portion, the sensor portion comprising at least one EM sample electrode (104A); Equipped with.

[0021] The anchor part is a) an anchor plate (200), i) an array of one or more reference electrodes (102Aa, 102Ab, ..., 102An); and ii) an anchor plate (200) having at least one anchor electrode (112A) configured to hold / immobilize the nucleotide strand (106A) at the anchor moiety (and conduct current along the nucleotide strand (106A) as described above); b) anchor plate actuator (124); c) an anchor support (114) that connects the anchor plate (200) to the base support structure (116), including via an anchor plate actuator (124), thereby supporting the anchor plate (200) during the measurement process of the sequencer device (100); Equipped with.

[0022] The holder is a) anchor electrode (112A), b) anchor molecules (118A) configured / selected to hold / immobilize the nucleotide strands (106A) to their corresponding anchor electrodes (112A), which may comprise self-assembled monolayers (SAMs) for DNA surface immobilization, as described in the article "A DNA Self-Assembled Monolayer for the Specific Attachment of Unmodified Double- or Single-Stranded DNA" by Cynthia Bamdad in the Biophysical Journal (V. 75, October 1998, pp. 1997-2003); and / or c) one or more reference electrodes (102Xx) that may be configured to attract / retain / fix the nucleotide strands (106X) to the anchor plate (200) to help hold the nucleotide strands (106X) stationary relative to the anchor plate (200) as the movable plate (300) moves relative to the anchor plate (200) and EM signals are generated and detected. Equipped with.

[0023] The sensor part is a) a movable plate (300) having one or more arrays of sample electrodes (104Aa, 104Ab, ..., 104An); b) a sensor-actuator (108) configured to movably hold / secure the movable plate (300) to the actuator support structure (122) and to secure the movable plate (300) in a controllably movable relationship relative to the base support structure (116); Equipped with.

[0024] 2, the anchor plate (200) comprises one or more parallel linear arrays of two or more reference electrodes (102Aa, 102Ab, ..., 102An), thus forming a two-dimensional (2D) array of reference electrodes (102Aa, 102Ab, ..., 102An, 102Ba, 102Bb, ..., 102Bn, 102Na, 102Nb, ..., 102Nn). The anchor plate (200) includes one or more of at least one anchor electrode (112A, 112B, ..., 112N), where each anchor electrode (112) holds one nucleotide strand (106) at a position on / along the corresponding linear array of reference electrodes (102a, 102b, ..., 102n), such that there is one anchor electrode (112) for each linear array of reference electrodes (102a, 102b, ..., 102n). Alternatively, the anchor plate (200) may include reference electrode plates corresponding to two or more of the sample electrodes (104Aa, 104Ab, ..., 104An), with widths and / or lengths corresponding to at least twice the widths and / or lengths of the corresponding sample electrodes (104Aa, 104Ab, ..., 104An). The reference electrode plates can accommodate nucleotide strands (106X) in use. The reference electrode tips may have a surface area at least 1%, 5%, or 10% larger than the surface area of ​​the corresponding sample electrode array for ease of handling and / or to accommodate larger amounts of DNA sample.

[0025] Optionally, the sensor portion of the sequencer device (100) may include an electrical insulator between each of the reference electrodes (102Xx) and the location where the nucleotide chain (106X) is held / fixed in place during the measurement process, for example, at least one insulator layer on the outer surface of each reference electrode (102Xx).

[0026] As shown in FIG. 3, the movable plate (300) comprises one or more parallel linear arrays of one or more sample electrodes (104Aa, 104Ab, ..., 104An), thus forming a 2D array of sample electrodes (104Aa, 104Ab, ..., 104An, 104Ba, 104Bb, ..., 104Bn, 104Na, 104Nb, ..., 104Nn).

[0027] The anchor portion of the sequencer device (100) may include an electrical insulator between each of the sample electrodes (104Xx) and the location where the nucleotide chain (106X) is held / fixed in place during the measurement process, for example, at least one insulator layer on the outer surface of each sample electrode (104Xx).

[0028] orthodontic device The system comprises at least one device ("correction device") for correcting the nucleotide chain (106A, 106B, ..., 106N) before insertion into the sequencer device (100). a) an operating nucleotide chain attractor for straightening nucleotide chains in solution, as shown in FIG. 4; and / or b) Dual-electrode nucleotide chain attractor for straightening nucleotide chains in solution, as shown in Figure 5. It may comprise:

[0029] During straightening, as shown in Figures 4 and 5, one or more nucleotide strands (106A, 106B, ..., 106N) are held / anchored by corresponding anchor molecules (118A, 118B, ..., 118N) and straightened / extended / stretched / unwound across / along a corresponding linear array of reference electrodes (102Xa, 102Xa, ..., 102Xn, where "X" is "A, B, ..., N") by attractor electrodes (404, 504) that apply attractive forces to the nucleotide strands (106A, 106B, ..., 106N) to straighten / extend / unwind the nucleotide strands (106X). The attractive forces are directed along each linear array of reference electrodes (102Xa, 102Xa, ..., 102Xn), away from the anchor molecules (118A, 118B, ..., 118N). The reference electrodes (102Xx) may be controlled to assist in straightening / stretching / extending / unwinding the nucleotide strands (106X) by applying a charge to the reference electrodes (102Xx) to attract the nucleotide strands (106X) across / along the linear array of reference electrodes (102Xx).

[0030] As shown in FIG. 4, one form of attractor electrode (404) is: a) an attractor electrode (404); b) an actuator ("attractor actuator") (402) mechanically coupled to the attractor electrode (404) and the additional support structure (406) such that the attractor electrode (404) is movable (by the attractor actuator (402) from a position close to the anchor molecule (118X) to a position farther from the anchor molecule (118X) after the nucleotide strand (106X) is held / fixed, for attracting / pulling and straightening / stretching / extending the nucleotide strand (106X) across / along a corresponding linear array of reference electrodes (102Xx, where "x" is "a, b, ..., n"); The actuation orthotic device (400) may include:

[0031] As shown in FIG. 5, one form of attractor electrode (504) is: a) an attractor electrode (504); b) a repulsor electrode (502) arranged to be biased by a voltage source (508) to form an electric field (506) surrounding the nucleotide strand (106X), the voltage source (508) being capable of pulsing / modulating the electric field (506) to gradually unwind the nucleotide strand (106X) during a preparation time prior to sensing an EM signal; The orthodontic device (500) may include a dual electrode.

[0032] As shown in Figures 4 and 5, the attractor electrodes (404, 504) apply a force (F) to the nucleotide strand (106X) along the linear array of reference electrodes (102Xx) in a direction away from the anchor molecule (118X). Because the linear array of reference electrodes (102Xx) is aligned and collinear with the corresponding linear array of sample electrodes (104Xx), the nucleotide strand (106X) is also corrected / stretched / elongated / positioned below / along the corresponding linear array of sample electrodes (104Xx) by the force (F) from the anchor molecule (118X) and the attractor electrodes (404, 504).

[0033] The dual electrode straightening device (500) and / or the actuation straightening device (400) may include an electrolyte solution in which the nucleotide strand (106X) is immersed, the electrolyte solution being selected to at least partially denature the nucleotide strand (106X), thereby allowing the force (F) to straighten / advance / extend the nucleotide strand.

[0034] The correction device (400, 500) may include a tray / vessel for holding an electrolyte solution around the nucleotide chain (106X).

[0035] The correction device (400, 500) may include a heater in and with the tray / vessel for heating the nucleotide strands (106X) so that they are denatured in a fluid and / or electrolyte solution, for example, to approximately 90°C.

[0036] Alternatively, the nucleotide chains (106X) may be fixed to the anchor plate (200) by other suitable methods / techniques capable of holding the nucleotide chains (106X) in place, such as using a surface treatment (e.g., hydrophobicity) of the anchor plate (200), assigning some of the reference electrodes (102Xx) as "nodes" to fix the nucleotide chains (106X) in place with electrostatic charges (so that these reference electrodes are not used for measurements), or by cryogenically cooling the nucleotide chains (106X) together with the anchor plate (200).

[0037] Actuators and Alignment The sensor actuator (108), anchor plate actuator (124), and attractor actuator (402) may include, for example, piezoelectric actuators manufactured by PI (Physik Instrumente), or piezoelectric ceramic actuators.

[0038] The anchor portion may comprise a stepper motor configured to move the anchor plate (200) into alignment with the movable plate (300) after the nucleotide strand (106) is fixed in place on / along the anchor plate (200), in three orthogonal directions X, Y, and Z, and additionally / optionally three rotational dimensions (e.g., Euler angles α, β, and γ). The sensor portion may comprise a stepper motor configured, together with the sensor actuator (108), to move the anchor plate (200) into alignment with the movable plate (300) after the nucleotide strand (106) is fixed in place on / along the anchor plate (200), in three orthogonal directions X, Y, and Z, and additionally / optionally three rotational dimensions (e.g., Euler angles α, β, and γ). Thus, the anchor plate (200) and the movable plate (300) are moved to an aligned state (or "in place" or "operated state") after the nucleotide strand (106X) is anchored and corrected in place, and in this aligned / operated state, there is a small gap between the plates (200, 300), which are approximately planar parallel, for example, this small gap can be less than 1 micrometer, less than 100 nanometers (nm), or less than 10 nm, for example, approximately the width of a DNA molecule (substantially 0.3 nm to 2.5 nm), but substantially larger than the width of a DNA molecule so that the movable plate (300) remains movable relative to the nucleotide strand (106X).

[0039] EM signal generation and detection When the sequencer device (100) is configured to measure current fluctuations due to the electrical resistance of the sample portion (due to quantum tunneling resistance) from the reference electrode (102A) or the sample electrode (104A) adjacent to the sample portion, the anchor plate (200) comprises at least one current source configured to supply current to the reference electrode (102A) and to the array of reference electrodes (102Xx) or the reference electrode plate, the current source including, for example, at least one wire / nanowire embedded in the substrate of the anchor plate (200), and the wire / nanowire is connected via an electrical circuit to a power source, for example a battery, external to the anchor plate (200).

[0040] Each sample electrode (104Xx) can be configured to measure a sample EM signal from a selected portion ("first portion") of the nucleotide strand (106X) directly or nearly directly adjacent to the sample electrode (104Xx). When the sample electrode (104Xx) is moved to modulate the sample EM signal, a different selected portion ("second portion") of the nucleotide strand (106X) becomes (nearly) directly adjacent to the sample electrode (104Xx). This different portion has a different charge and / or electromagnetic conductivity / absorption than the initial portion; thus, the sample EM signal is modulated by this difference. This difference depends on the difference in the nucleotide base / base pair moved between the sample electrodes (104Xx), and thus the detectable EM signal indicates the nucleotide sequence located between the first and second portions. By calculating / processing the sample EM signals from the multiple sample electrodes (104Xx), at least a portion of the nucleotide bases / base pairs can be deduced / detected from the nucleotide strand (106X).

[0041] Each reference electrode (102Xx) can be configured to measure a reference EM signal from a reference portion of the nucleotide strand (106X) that is directly or nearly directly adjacent to the reference electrode (102Xx). Because the reference electrode (102Xx) is not moved during the measurement process, the reference EM signal should not change substantially during movement of the sample electrode (104Xx), and therefore the reference EM signal can be substantially constant and provide a reference for use in the noise removal process. In the case of a reference plate electrode, the reference EM signal includes signals from multiple portions of the nucleotide strand (106X) that are directly or nearly directly adjacent to the reference electrode (102X).

[0042] The modulation mechanism may be configured to move each sample electrode (104Xx) along the linear array of reference electrodes (102X) a distance at least equal to the length (direction of movement) of each sample electrode (104Xx) plus the length of the spacing between the sample electrodes (104Xx), in other words, a distance at least equal to the period of the sample electrodes (104Xx), so that each portion of the nucleotide chain (106X) under the sample electrode (104Xx) is traversed along its length.

[0043] The length of the sample electrodes (104Xx) in the direction of movement can be substantially 2 to 50 nm. Each sample electrode (104Xx) can generate an EM signal from a selected portion containing substantially 1 to 150 base pairs when each nucleotide unit extends substantially 0.33 nm in the direction of movement along the nucleotide chain.

[0044] Preferably, and optionally, the sample electrodes (104Xx) may be arranged so that there is overlap in the detection range between different or adjacent sample electrodes (104Xx), in which case redundancy can be used to account for systematic variations. For example, differences in shape or local distribution of impurities may reduce the sensitivity of one sample electrode overall or only for specific base / charge densities. Overlap in the detection range between different or adjacent sample electrodes (104Xx) may reduce or eliminate one or more of these systematic variations, thereby improving the accuracy of nucleotide sequencing.

[0045] The modulation mechanism can move the sample electrodes (104Xx) at a selected frequency / periodicity so that the sample EM signal can be demodulated at a corresponding frequency to improve the signal-to-noise ratio (SNR) of the sample EM signal.

[0046] The modulation mechanism can move the sample electrodes (104Xx) based on signal feedback from the electrodes.

[0047] process In operation, the system: a) holding / fixing a nucleotide chain (106X) by an anchor molecule (118X); b) straightening / stretching / extending / unwinding the nucleotide chain (106X) by the action of the attractor electrodes (404, 504) using the actuation straightening device (400) and / or the dual electrode straightening device (500); c) receiving the nucleotide chain (106X) by a reference electrode (102Xx), which may include a reference electrode plate; d) straightening / stretching / extending / unwinding the nucleotide chain (106X) by actuation of the reference electrodes (102Xx) along each linear array; e) the reference electrodes (102Xx) are charged (e.g., by a current source or semiconductor doping) to attract the nucleotide strands (106X), thereby holding / fixing the nucleotide strands (106X) on / along the linear array of the reference electrodes (102Xx) by electrostatic attraction between the reference electrodes (102Xx) and the nucleotide strands (106X); f) draining the electrolyte; g) transferring the straightened / stretched / extended / rewound nucleotide chain (106X) from the actuation straightening device (400) or the dual electrode straightening device (500) to the sequencer device (100); h) moving the anchor plate (200) and the movable plate (300) into alignment, i.e., an operating state, by controlling the sensor actuator (108) and / or the anchor plate actuator (124); i) after discharging the electrolyte solution, sequencing the anchored and extended nucleotide chain (106X) by performing a measurement process in which an EM signal is detected; Execute a process that includes

[0048] The measurement process is a) measuring EM signals from a sample portion of a nucleotide chain (106X); b) moving the nucleotide chain (106X) to modulate the EM signal; Includes.

[0049] The measurement process is a) measuring the current variations (conductance) between a reference electrode (102X) and a corresponding sample electrode (104X) caused by the effect of different local charge densities on different nucleotides; b) measuring the variation in the current conducted through a portion of the nucleotide chain (106A) from the anchor electrode (112X) to one of the sample electrodes (104Xx); c) measuring a reference current conducted through a portion of the nucleotide chain (106A) from the anchor electrode (112X) to one of the reference electrodes (102Xx); and / or d) measuring the variation of the current in (a) and (b) in response to a fixed frequency modulation applied to the anchor plate (200) via the plate actuator (124); may include:

[0050] Implementation The movable plate (300) may be formed as a silicon chip / plate, and the sample electrodes (104Xx) may be formed using a photolithography process. The sample electrodes (104Xx) may be formed as electrodes in a square 2D grid located on the upper surface of the silicon chip / plate. The movable plate (300) may include a mixed-signal integrated circuit, for example integrated into the substrate below the silicon chip / plate, that amplifies, filters, and converts analog signals from the sample electrodes (104Xx) to amplified signals and, optionally, to digital signals for electronic demodulation / processing in an external computer.

[0051] The anchor plate (200) may be formed as a silicon chip / plate, and the reference electrodes (102Xx) may be formed using a photolithography process. The reference electrodes (102Xx) may be formed as electrodes in a square 2D grid located on the upper surface of the silicon chip / plate. The anchor plate (200) may include a mixed-signal integrated circuit, integrated into the substrate below the silicon chip / plate, for example, that amplifies, filters, and converts analog signals from the reference electrodes (102Xx) to amplified signals and, optionally, to digital signals for electronic demodulation / processing in an external computer.

[0052] Alternatively, other suitable fabrication methods / processes, such as guided growth, may be used to form the sample electrodes (104Xx) on the movable plate (300) and / or the reference electrodes (102Xx) on the anchor plate (200).

[0053] The sample electrodes (104Xx) and the reference electrodes (102Xx) may be field effect sensors. The sample electrodes (104Xx) and the reference electrodes (102Xx) are configured to measure EM signals, as further detailed hereinabove.

[0054] The electrodes of the sample electrodes (104Xx) and reference electrodes (102Xx) may be fabricated at the nanoscale on a silicon wafer using a photolithography process or any other suitable fabrication method / process, for example, guided growth.

[0055] Each sample electrode (104Xx) may be made of highly p-doped silicon and may include a 1-2 nm thin oxide layer, or a 1 nm sub-oxide layer, chemically grown by native oxidation on its surface to act as an electrical insulator between the sample electrode (104Xx) and the sample portion of the nucleotide chain (106). The sample electrodes (104Xx) may be rectangular prisms, each 1-50 nm long and 1-50 nm wide, or 2-50 nm long and 2-50 nm wide, with approximately equal lengths and widths (thus forming a square cross section). Each sample electrode (104Xx) may be vertically embedded in a silicon substrate to achieve a quasi-atomically flat surface across the entire chip, i.e., on the plane facing the nucleotide chain (106). The sample electrodes (104Xx) are spaced from each other by a selected distance on all sides to mitigate electrical interference and possible quantum tunneling effects. The selected distance (interval or separation distance) between adjacent sample electrodes (104Xx) may be approximately equal to the length or width of the sample electrodes (104Xx); for example, each sample electrode (104Xx) may have an approximately square area in a plane parallel to the 2D array, with the inter-electrode spacing approximately equal to the sides of each square in that plane. For example, if the electrodes are 1 × 1 nm, the inter-electrode distance or gap may be 1 nm, or if the electrodes are 2 × 2 nm, the inter-electrode distance or gap may be 2 nm. The inner side of each sample electrode (104Xx) facing away from the nucleotide chain (106) may be connected to the gate of a CMOS transistor that functions as a signal amplifier.

[0056] In a first embodiment, each reference electrode (102Xx) may be made of highly p-doped silicon and may include a 1-2 nm thin oxide layer, or a 1 nm sub-oxide layer, chemically grown on its surface by natural oxidation to act as an electrical insulator between the reference electrode (102Xx) and the sample portion of the nucleotide chain (106). The reference electrodes (102Xx) may be rectangular prisms. The reference electrodes (102Xx) may each be 1-50 nm long and 1-50 nm wide, or 2-50 nm long and 2-50 nm wide, and their lengths and widths may be approximately equal (thus forming a square cross section). The reference electrodes (102Xx) may also be referred to as "nanotips," "nanotip electrodes," or "probes" that include these lengths and widths. Each reference electrode (102Xx) may be vertically embedded in a silicon substrate to achieve a quasi-atomically flat surface across the entire tip, i.e., in the plane facing the nucleotide chain (106). The reference electrodes (102Xx) are spaced apart (i.e., spaced apart from one another) by a selected distance on all sides to reduce electrical interference and possible quantum tunneling effects between directly adjacent electrodes. The selected distance (interval, or mutual separation) may be equivalent to (i.e., approximately equal to) their length or width. For example, if the electrodes are 2 × 2 nm, the distance between the electrodes may be selected to be 2 nm. The inner side of each reference electrode (102Xx) facing the opposite side of the nucleotide chain (106) may be connected to the gate of a CMOS transistor that functions as a signal amplifier. Alternatively, the inner side of each reference electrode (102Xx) may be in electrical communication with an external computer via an electronic circuit including a mixed-signal integrated circuit.

[0057] The anchor electrode (112X) may be formed of n-doped silicon, such that the anchor molecule 118X is selected / configured to bond with the n-doped silicon. The anchor electrode (112X) may be connected / configured to conduct current through the anchor molecule (118X) and, therefore, the nucleotide chain (106A).

[0058] The base support structure 116 may comprise a material, such as a high-density block of granite, suspended on an air table to reduce external vibrations during operation of the sequencer device 100. The base support structure 116 may also comprise an EM insulating cage / shell (referred to as a "Faraday cage") that encloses the sample portion and anchor portion to reduce EM interference when the sequencer device 100 is operating, and the cage / shell may comprise a metal alloy of titanium carbonitride.

[0059] III. Second Embodiment A second embodiment of a system comprising the device (100) is described below.

[0060] Basic structure of sequencing device As shown in FIG. 6, the sequencer device (100) a) at least one EM sample electrode (104Aa, 104Ab, ..., 104An, "sample electrode" (104A)) for measuring an electromagnetic (EM) signal ("sample EM signal") (e.g., charge and / or current) from a portion (i.e., "sample portion") of the nucleotide chain (106A); b) a mechanism configured to move the nucleotide strand (106A) relative to the EM sample electrode (104A) to modulate the EM signal (i.e., a "modulation mechanism"); Equipped with.

[0061] At least one sample electrode (104A) may be a non-contact EM sensor, i.e., the sample electrode does not contact or touch the nucleotide strand (106A) during the measurement process. The sample electrode (104A) may, for example, take the form of a nanotip electrode, sometimes hereinafter referred to as a "nanotip" or "probe."

[0062] At least one sample electrode (104A) is provided in a sensor portion of the device (100). a) a sensor plate (700) having one or more arrays of sample electrodes (104Aa, 104Ab, ..., 104An); b) a sensor support structure (722) that secures the sensor plate (700) to the base support structure (116); Equipped with.

[0063] The mechanism comprises an actuator configured to move a common current collecting plate (600) on which the nucleotide strands (106A) are deposited.

[0064] More specifically, the device (100) comprises: a) a common current collecting plate (600) which is a conductive base plate for holding / supporting the nucleotide chains (106A); b) a current collector plate actuator (624) for moving the common current collector plate (600); c) a plate support structure (614) connecting the common current collecting plate (600) in a controllably movable relationship to the base support structure (116), including via a current collecting plate actuator (624), during the measurement process of the sequencer device (100); The current collecting plate portion includes:

[0065] 7, the sensor plate (700) may include one or more arrays of two or more sample electrodes (104An), thus forming a two-dimensional (2D) array of sample electrodes (104Aa, 104Ab, ..., 104An, 104Ba, 104Bb, ..., 104Bn, 104Na, 104Nb, ..., 104Nn), with the tip of each electrode facing a common current collecting plate (600). Each of the arrays in the 2D array may be generally linear in the sense of forming a line or chain or series of adjacent electrodes, with each electrode adjacent to the next electrode in the series. A 2D array may be formed from multiple of these linear arrays (which may be one-dimensional or may have some curvature so long as the electrodes form a substantial series), which may be parallel or substantially parallel (e.g., with a corresponding curvature) or adjacent or next to each other, including at least substantially adjacent to each other, thus forming the 2D array.

[0066] The nucleotide strands (106A), when deposited on the common current collecting plate (600), are positioned and fixed in place (e.g., by van der Waals forces) such that the nucleotide strands (106A) are linearly aligned beneath / along a corresponding linear array of sample electrodes (104a, 104a, ..., 104n), each of which addresses multiple nucleotides.

[0067] The EM signal measured by the sample electrode (104A) may include one or more of a charge, a current, and / or a voltage.

[0068] The measured EM signal exhibits electromagnetic fluctuations due to quantum tunneling, a quantum mechanical phenomenon in which, under certain conditions (e.g., distance, bias voltage), electrons have a finite probability of crossing a potential barrier determined by the physical gap between electrodes.

[0069] Due to quantum tunneling, DNA molecules perturb the condition when in proximity to the sample electrode (104A), thus generating a measurable electromagnetic signal resulting from the difference in electron flow. The disruption of the tunneling process has repeatable characteristics (i.e., "signatures") that can be used to distinguish between different types of nucleotides (e.g., by using an algorithm trained with the EM features or signatures for each type of nucleotide), thus enabling sequencing of the nucleic acid strand. For example, signal features / signatures can be extracted from the measured EM signal (e.g., in the form of current-voltage (IV) curves, current-time (It) traces, and / or other processed signals derived from these measurements).

[0070] In the embodiments described herein, relative movement between the nucleotide strands 106A and the EM sample electrode 104A is achieved by moving the nucleotide strands 106A, more specifically, by moving the common current collecting plate 600 on which the nucleotide strands 106A are deposited. Alternatively, this relative movement may be achieved by moving the sample electrode 104A instead of or in addition to the common current collecting plate 600. For example, the sensor actuator 118 described in the first embodiment may be provided to enable movement of the sensor plate 700 on which the sample electrode 104A is disposed. However, moving the common current collecting plate 600 rather than the sensor plate 700 reduces mechanical stress on the sensor plate 700 and avoids vibration of the sample electrode 104A that may result from movement, thereby achieving more accurate measurement and sequencing results.

[0071] orthodontic device The system may comprise at least one device ("straightening device") for straightening / stretching / extending / rewinding the nucleotide chain (106A, 106B, ..., 106N) before insertion into the sequencer device (100). This straightening device may have the same or similar structure as that described above in the first embodiment.

[0072] Alternatively, straightening of the nucleotide chain (106A, 106B, ..., 106N) may be performed using molecular threading techniques, for example, as described in "Molecular threading: mechanical extraction, stretching, and placement of DNA molecules from a liquid-air interface," Payne, Andrew C. et al., PloS one 8.7(2013):e69058.

[0073] Actuators and Alignment The current collecting plate actuator (624) may comprise, for example, a piezoelectric actuator manufactured by PI (Physik Instrumente) or a piezoelectric ceramic actuator.

[0074] The current collecting plate portion may comprise a stepper motor configured to move the common current collecting plate (600) into alignment with the sensor plate (700) after the nucleotide strands (106X) have been deposited in place on the common current collecting plate (600), including in all three orthogonal directions X, Y, and Z, and additionally / optionally in three rotational dimensions (e.g., Euler angles α, β, and γ). A small gap exists between the plates (600, 700), which are substantially planar-parallel; for example, this small gap may be less than 1 micrometer, less than 100 nanometers (nm), or less than 10 nm, e.g., about the width of a DNA molecule (substantially 0.3 nm to 2.5 nm), but substantially larger than the width of a DNA molecule, so that the nucleotide strands (106X) and the common current collecting plate (600) remain controllably movable relative to the sensor plate (700).

[0075] EM signal generation and detection When the sequencer device (100) is configured to measure a sample EM signal (e.g., charge, voltage, and / or current) from a sample electrode (104A) adjacent to the sample portion, the sensor plate (700) includes at least one current source configured to supply a respective voltage / current to the sample electrodes (104Xx), e.g., at least one wire / nanowire embedded in the substrate of the sensor plate (700) is connected via an electrical circuit to a power source, e.g., a battery, external to the sensor plate (700). On the other hand, the common current collecting plate (600), which is a conductive plate, is electrically connected to an electrode on the opposite side of the power source, either directly or via an electrical circuit.

[0076] Each sample electrode (104Xx) is configured to measure a sample EM signal from a selected portion (a "first portion") of the nucleotide strand (106X) that is immediately adjacent or nearly immediately adjacent to the sample electrode (104Xx).

[0077] The selected portion ("first portion") of the nucleotide chain (106X) may include multiple nucleotides. Thus, the measured sample EM signal is the aggregate value associated with all of these nucleotides in the sample portion. The sample EM signal is modulated to identify the EM signal associated with each individual nucleotide.

[0078] The sample EM signal may be modulated by moving (e.g., dithering) the common current collecting plate (600) so that the nucleotide strand (106X) is moved in the longitudinal direction of the nucleotide strand (106X), thereby causing relative movement between the nucleotide strand (106X) and the sample electrode (104Xx).

[0079] As a result of this movement, a different selected portion ("second portion") of the nucleotide strand (106X) becomes directly or substantially adjacent to the sample electrode (104Xx), this different portion having a different charge, current, and / or voltage than the initial portion (first portion), such that a change in the sample EM signal reflects this difference, which depends on the difference in the nucleotide base / base pair moved between the sample electrodes (104Xx), and thus the detectable EM signal is indicative of the nucleotide sequence located between the first and second portions. By calculating / processing the sample EM signals from the multiple sample electrodes (104Xx), at least a portion of the nucleotide bases / base pairs can be deduced / detected from the nucleotide strand (106X).

[0080] The modulation mechanism may be configured to move the common current collecting plate (600) in the longitudinal direction of the nucleotide chain (106X) by a distance at least equal to the length (direction of movement) of each sample electrode (104Xx) plus the length of the spacing between the sample electrodes (104Xx), in other words, by a distance at least equal to the period of the sample electrodes (104Xx), so that each portion of the nucleotide chain (106X) under the sample electrode (104Xx) is traversed along its length.

[0081] The length of the sample electrodes (104Xx) in the direction of movement can be substantially 2 to 50 nm. Each sample electrode (104Xx) can generate an EM signal from a selected portion containing substantially 1 to 150 base pairs when each nucleotide unit extends substantially 0.33 nm in the direction of movement along the nucleotide chain.

[0082] Preferably, and optionally, the sample electrodes (104Xx) may be arranged so that there is overlap in the detection range between different or adjacent sample electrodes (104Xx), in which case redundancy can be used to account for systematic variations. For example, differences in shape or local distribution of impurities may reduce the sensitivity of one sample electrode overall or only for specific base / charge densities. Overlap in the detection range between different or adjacent sample electrodes (104Xx) may reduce or eliminate one or more of these systematic variations, thereby improving the accuracy of nucleotide sequencing.

[0083] The modulation mechanism may move the common current collecting plate (600) at a selected frequency / periodicity so that the sampled EM signal can be demodulated at a corresponding frequency to improve the signal-to-noise ratio (SNR) of the sampled EM signal.

[0084] The movement of the common current collecting plate (600) may be induced, for example, by one or more standard off-the-shelf drivers configured to achieve a suitable modulation frequency. The modulation frequency may be selected based on the spectral composition of the noise. For example, it may be selected to encode the signal at a frequency with the lowest noise level. Most noise is expected to be in the lowest frequency range (up to tens of kHz), but this can be mitigated by a solid support substrate (e.g., a granite substrate) and a vibration isolation table. Preferably, the modulation mechanism allows for variation of the modulation frequency. This allows flexibility in selecting a modulation frequency that provides the best noise reduction effect and ensures robustness against possible noise sources. For example, the modulation frequency from the modulation mechanism may be in the range of 1 Hz to 100 Hz, or may be variable between two frequencies selected within the range, e.g., between 1 Hz and 100 Hz.

[0085] Optionally, in addition to the longitudinal modulation, the common current collecting plate (600) may be vertically dithered by vertical modulation. The vertical modulation may have a vertical modulation frequency in the range of 1 Hz to 100 Hz, and may optionally be variable between two frequencies selected within the range of 1 Hz to 100 Hz, for example, between 1 Hz and 100 Hz. The vertical modulation, if present, may be independent of the horizontal modulation mechanism.

[0086] Alternatively, or additionally, sample modulation may be performed by varying the input electromagnetic signal to the sample electrodes (104Xx), including electrically modulating the corresponding voltage / current supplied to the sample electrodes (104Xx), which may result in a change in the sample EM signal measured from each sample electrode (104Xx) and is referred to as "electrical modulation." Electrical modulation of the input voltage / current may be performed, for example, by standard off-the-shelf electronics / software (e.g., an off-the-shelf lock-in amplifier). The electrical modulation frequency may be selected to reduce or mitigate noise, including noise arising from in-circuit electronics (e.g., based on the spectral composition of the noise). For example, it may be selected to encode the signal at a frequency with the lowest noise level. The electrical modulation may include an electrical modulation frequency in the range of 1 Hz to 100 Hz and may optionally be variable between two frequencies selected within the range, e.g., between 1 Hz and 100 Hz. EM modulation, if present, may be independent of vertical and horizontal modulation.

[0087] By calculating / processing sample EM signals from a plurality of sample electrodes (104Xx) based on the input electromagnetic signals, at least a portion of the nucleotide bases / base pairs can be deduced / detected from the nucleotide chain (106X).

[0088] process In operation, the system according to this embodiment: a) depositing nucleotide chains (106X) on a common current collecting plate (600); b) moving the common current collecting plate (600) and the sensor plate (700) into alignment, i.e., an operative state, by controlling the current collecting plate actuator (624); c) sequencing the nucleotide strands (106X) on the common current collecting plate (600) by performing a measurement process in which EM signals are detected; Execute a process that includes

[0089] The measurement process is a) measuring an EM signal from a sample portion of a nucleotide chain (106X) using a sample electrode (104X); b) moving the nucleotide chain (106X) relative to the sample electrode (104X) to modulate the EM signal; Includes.

[0090] The measured EM signal includes one or more of charge, current, and voltage.

[0091] Preferably, the nucleotide chain (106X) is straightened / stretched / extended / unwound before being moved to the common current collecting plate (600). The straightening / stretching / extension / unwinding process may be performed by using a molecular threading technique. Alternatively, it may be performed by using a straightening device, such as the actuated straightening device (400) or the dual-electrode straightening device (500), as described above in the first embodiment. The straightening device may be integrated into the system of the present disclosure or may be separate from the system of the present disclosure. Alternatively, the straightening / stretching / extension / unwinding process may be performed by using any other nucleotide chain straightening technique / device applicable to the described embodiments.

[0092] The measured EM signal is a collective value associated with all of the nucleotides in the sample portion of the nucleotide strand (106X). To identify the EM signal associated with each individual nucleotide, the sample EM signal is demodulated based on a movement signal associated with the movement of the nucleotide strand (106X) relative to the sample electrode (104X). The movement signal may include a signal (which may also be referred to as a "dithering signal") indicating the distance and timing of the relative movement of the nucleotide strand (106X) relative to at least one sample electrode (104X) in the longitudinal direction of the nucleotide strand (106X). The modulation / demodulation process may also allow the sequencing information to be isolated from other forms of noise.

[0093] From the demodulated EM signals (e.g., in the form of current-voltage (IV) curves and / or current-time (It) traces), unique features or signatures associated with each known nucleotide type can be extracted. The extracted signal features or signatures are then used to resolve / identify the nucleotide types in a sample portion of the nucleotide chain (106X), which can be done using a database or an automatic classifier (e.g., a proprietary algorithm trained using the EM features / signatures for each nucleotide type).

[0094] Optionally, or alternatively, the sample EM signal may be modulated by controlling / varying the input EM signal to one or more of the sample electrodes (104X), for example, the current or voltage applied to one or more of the sample electrodes (104X). This may provide more extractable signal features or signatures for determining / identifying the type of each nucleotide in the sample portion of the nucleotide chain (106X), thus improving the accuracy and efficiency of sequencing.

[0095] Implementation The sensor plate (700) may be formed as a silicon chip / plate, and the sample electrodes (104Xx) may be formed using a photolithography process. The sample electrodes (104Xx) may be formed as electrodes in a square 2D grid located on the upper surface of the silicon chip / plate. Alternatively, other suitable fabrication methods / processes, such as guided growth, may be used to form the sample electrodes (104Xx) on the sensor plate (700). The sensor plate (700) may also include a mixed-signal integrated circuit integrated into the substrate, for example, in the silicon chip / plate, that amplifies, filters, and converts analog signals from the sample electrodes (104Xx) to amplified signals and, optionally, converts them to digital signals for electronic demodulation / processing in an external computer.

[0096] The common current collecting plate (600) may be formed as a conductive silicon chip / plate and may be connected to the same external computer via a current collecting plate actuator (624) for controlling / processing signals between the sensor plates (700).

[0097] The system comprises an electronic system (800). As shown in the schematic diagram of Figure 8, an external computer ("PC controller (802)") is electrically connected to a common current collecting plate (600) via a current collecting plate actuator in the form of a piezoelectric 3D stage (804). Sample electrodes (104Xx) in the form of a multi-electrode array plate (818) are controlled by the external computer (PC controller (802)) via multiple digital switches (816). EM signals collected by the multi-electrode array plate (818) are processed by a low-noise amplifier (814) and a lock-in current amplifier (812), synchronized to a moving signal ("dithering signal"), converted to digital data by an analog-to-digital converter ("ADC (810)"), and then collected by a data acquisition FPGA (808) and sent to the PC controller (802). The data acquisition FPGA (808) also collects data sent from the PC control unit (802) to the multi-digital switch (816) and controls the input signals to the multi-electrode array plate (818).

[0098] The sample electrodes (104Xx) are configured to emit / generate EM electric fields / charges or electrons (for quantum tunneling).

[0099] Using photolithography processes, sample electrodes (104Xx) may be fabricated on a silicon wafer at the nanoscale. Each sample electrode (104Xx) may be made of highly p-doped silicon and may include a 1-2 nm thin oxide layer, or a 1 nm sub-oxide layer, chemically grown on the surface by natural oxidation to act as an electrical insulator between the sample electrode (104Xx) and the sample portion of the nucleotide chain (106X). The sample electrodes (104Xx) may be rectangular prisms, each 1-50 nm long and 1-50 nm wide, or 2-50 nm long and 2-50 nm wide, with approximately equal lengths and widths (thus forming a square cross section). Sample electrodes (104Xx) with these lengths and widths may also be referred to as "nanotips," "nanotip electrodes," or "probes." Each sample electrode (104Xx) may be vertically embedded in a silicon substrate to achieve a quasi-atomically flat surface across the entire chip, i.e., in the plane facing the nucleotide chain (106). The sample electrodes (104Xx) are spaced apart from each other by a selected distance on all sides to reduce electrical interference and possible quantum tunneling effects between directly adjacent electrodes. The selected distance (interval, or mutual separation) may be equivalent to (i.e., approximately equal to) their length or width. For example, if the electrodes are 2 × 2 nm, the distance between the electrodes may be selected to be 2 nm. The inner side of each sample electrode (104Xx) facing the opposite side of the nucleotide chain (106X) may be connected to the gate of a CMOS transistor that functions as a signal amplifier. Alternatively, the inner side of each sample electrode (104Xx) may be in electrical communication with an external computer via an electronic circuit including a mixed-signal integrated circuit.

[0100] The base support structure 116 may comprise a material, such as a high-density block of granite, suspended on an air table to reduce external vibrations during operation of the sequencer device 100. The base support structure 116 may also comprise an EM insulating cage / shell (referred to as a "Faraday cage") that encloses the sample portion and the current collecting plate portion to reduce EM interference when the sequencer device 100 is operating, and the cage / shell may comprise a metal alloy of titanium carbonitride.

[0101] IV.Applications In the field of gene sequencing, there may be significant shortcomings in terms of length, time, accuracy, reliability, scalability, and price.In the field, there may be many devices with their own advantages and disadvantages, so that scientists are forced to use multiple devices / products and techniques to sequence samples, depending on their needs and what they can afford (time, accuracy, etc.).These problems prevent the field from fully realizing its potential, and therefore prevent humanity from benefiting from it.The system, device, and process described herein can address these problems.

[0102] Due to the highly parallel nature of the sequencer device (100) and the electromagnetic basis of the sample and reference electrodes (104, 102), large parallel "strands" / "strings" of DNA, ssDNA, or RNA can be analyzed with high uniformity in terms of length and time. Because genomes can be arranged in a 2D sheet within the sequencer device (100), electrode arrays (104X, 102X) can be formed using, for example, a grid of conductive electrodes, using 2D fabrication techniques and equipment from the semiconductor industry, for example. By creating vibrations with actuators (including the anchor plate actuators (124) and / or the sensor actuators (108), e.g., piezoelectric ceramic actuators), fluctuations in electromagnetic signals (e.g., charge, voltage, or current) become detectable via electronic circuitry. Because the entire genome can be sequenced simultaneously, this can reduce sequencing time from days to minutes. Due to the small vibrations in the relative motion of the sample electrode (104Xx) and the nucleotide chain (106X), as well as the small dimensions of the sample and reference electrodes (104, 102), single-base accuracy may be achievable. Because the sequencer device (100) does not need to be blocked during the sequencing process and can be easily cleaned and quickly reused, a single sequencer device (100) could potentially sequence multiple whole human genomes per day. Because polymerase chain reaction (PCR) amplification is not required, using this system may be substantially less expensive than conventional techniques.

[0103] V. Interpretation The drawings included herewith illustrate aspects of non-limiting, representative embodiments according to the present disclosure, although certain structural elements shown in the drawings may not be shown to scale or precisely to scale relative to each other. The depiction of a given element, or consideration or use of a particular element number in a particular drawing, or reference in corresponding descriptive material, can encompass identical, equivalent, similar, taxonomically similar, or like elements or element numbers identified in another drawing or its accompanying descriptive material. The presence of " / " in any drawing or text herein shall be understood to mean "and / or," i.e., "X / Y" shall mean "X" or "Y," or "both X and Y," unless otherwise indicated. The recitation of a particular numerical value or range of values ​​herein is understood to include or be the recitation of that approximate value or range of values ​​within, for example, + / -20%, + / -15%, + / -10%, + / -5%, + / -2.5%, + / -2%, + / -1%, + / -0.5%, or + / -0%. The terms "essentially all" or "substantially" can indicate a percentage of 50%, 60%, 70%, 80%, or 90% or more, e.g., 92.5%, 95%, 97.5%, 99%, or 100%.

[0104] Many modifications will be apparent to those skilled in the art without departing from the scope of the invention.

[0105] Throughout this specification and the claims that follow, unless the context requires otherwise, the word "comprise" and variations such as "comprises" and "comprising" will be understood to imply the inclusion of a stated integer or step or group of integers or steps, but not the exclusion of any other integer or step or group of integers or steps.

[0106] Reference in this specification to any prior publication (or information derived therefrom) or any known matter is not, and should not be construed as, an acknowledgement or admission, or any form of suggestion, that the prior publication (or information derived therefrom) or known matter forms part of the common general knowledge in the field of endeavor to which this specification pertains.

[0107] VI. Illustrative Embodiments Exemplary embodiments provide for direct sequencing of nucleotide chains (or "nucleic acid strands") by direct (i.e., "first-hand") real-time measurement of electronic configurations along the molecule using a 2D array of sample electrodes (also referred to as an "electrode matrix / grid"). In some exemplary embodiments, the sample electrodes have a generally square cross-section that tapers toward their free ends, forming a curved tip that may be easier to fabricate than, for example, a square-topped tip. The sample electrodes may be considered similar to scanning tunneling microscope (STM) tips, except that each sample electrode is not an atomic-scale nanotip (as in an STM), but rather a large nanotip that is easier to fabricate in an array format using standard commercially available photolithographic fabrication tools and processes. Each sample electrode can operate to provide electronic signatures of multiple nucleotides (or "assemblies") probed by the sample electrode by measuring current-voltage (IV) profiles in a manner similar to conventional STM-based methods for scanning DNA molecules with a single, ultra-sharp nanoscale conductive tip, as described, for example, in Shapir, E. (2007), "Electronic structure of single DNA molecules resolved by transverse scanning tunneling spectroscopy," Nature Materials, 7, 68.10.1038 / nmat2060. In some examples, the system can be configured to detect tunneling currents at each sample electrode from substantially 2 picoamperes (pA) to substantially 10 nanoamperes (nA), e.g., substantially 10-100 pA, at a fixed bias of less than 10 V. In some examples, the small gap between the plates is substantially the width of a DNA molecule (substantially 0.3 nm to 3 nm) or greater, although the gap from each sample to the nucleotide strand, in use, can be substantially equal to the width of a DNA molecule, e.g., substantially 1 nm or 3 nm.For example, the measured EM signals may include current over time (It) at a fixed bias voltage; current versus distance (Iz) at a fixed bias voltage and a sample electrode moved along a direction perpendicular to the substrate to repeatably approach the molecule and collect tunneling data (I) as a function of distance (z); current-voltage curves (IV) and derivatives thereof, where the tunneling current is recorded as a function of varying bias voltage using analysis techniques described, for example, in Tanaka, H. (2009), "Partial sequencing of a single DNA molecule with a scanning tunneling microscope," Nature Nanotechnology, 4, 518.10.1038 / NNANO.2009.155 and / or Tanaka, H. (2017), "Sequencing of adenine in DNA by scanning tunneling microscopy," Japanese Journal of Applied Physics, 56, 08LB02.10.7567 / JJAP.56.08LB02. The EM signals may be processed to identify nucleotides using deterministic signal processing analysis and software, as well as machine learning techniques. This process may include the use of density functional theory (DFT) to provide results as data points that aid in identification. This process may include transverse measurements, such as those described in Zwolak, M. (2005), "Electronic Signature of DNA Nucleotides via Transverse Transport," Nano Letters, 5, 421.10.1021 / nl048289w.Some embodiments may use machine learning methods to identify DNA molecules, as described, for example, in Albrecht, T. (2017), "Deep learning for single-molecule science," Nanotechnology, 28, 423001.10.1088 / 1361-6528 / aa8334, or Im, J. (2018), "Recognition Tunneling of Canonical and Modified RNA Nucleotides for Their Identification with the Aid of Machine Learning," ACS Nano, 12, 7067.10.1021 / acsnano.8b02819. An exemplary embodiment may have a 2D array formed by a 10 x 10 electrode array, each electrode printed with direct contacts to an external circuit, and connectors integrated with and covered by insulating material (e.g., SiO2). The movement mechanism may be provided by a commercially available 6-axis piezoelectric system. The moving mechanism also mechanically adjusts the common current collecting plate so that the DNA strands oscillate relative to the measurement electrodes. This modulation may be used to encode signal data (representing the molecules being measured) through a process of modulation encoding. This signal is then decoded using deterministic algorithms for signal processing, as well as machine learning techniques. The modulation frequency may be adjustable in the range of 1 Hz to 100 Hz to better distinguish the detected EM signal (e.g., current) from noise, and the detected EM signal can be filtered using a corresponding electronic filter or demodulator circuit corresponding to the electronic modulation (e.g., having a corresponding frequency or frequency pattern). In some embodiments, the EM signal collected by the multi-electrode array plate (818) may be connected to the low-noise amplifier (814) by an N-to-1 switch (e.g., including a multiplexer switch commercially available from Analog Devices) to reduce the consumption of the amplification module.This may reduce the sampling rate of the system, but costs may be reduced by multiplexing the amplifier module (e.g., by converting N×M parallel signals into N parallel signals with M continuous signals). In some embodiments, the low-noise amplifier (814) may comprise a picoamp input current quad operational amplifier commercially available from Analog Devices, the ADC (810) may comprise an ADC module commercially available from Texas Instruments, the data acquisition FPGA (808) may comprise an FPGA commercially available from Intel, and the lock-in current amplifier (812) may comprise a modulator / demodulator module commercially available from Texas Instruments.

Claims

1. 1. A system for sequencing a nucleotide chain, comprising: at least one electrode for measuring a total electromagnetic signal from a portion of the nucleotide strand comprising a plurality of nucleotides; a mechanism configured to move the nucleotide strand relative to the electrodes to modulate the aggregate electromagnetic signal to identify an electromagnetic signal associated with each individual nucleotide in the plurality of nucleotides; A system comprising an apparatus having:

2. the device comprises a conductive plate for holding the nucleotide chain; The system of claim 1 , wherein the mechanism is configured to move the conductive plate relative to the electrode.

3. The system of claim 1 or 2, wherein the at least one electrode comprises at least one nanotip electrode.

4. The system of any one of claims 1 to 3, wherein the at least one electrode comprises a two-dimensional array of electrodes.

5. the apparatus is configured to sequence a plurality of nucleotide strands; 5. The system of claim 4, wherein the two-dimensional array of electrodes comprises a corresponding plurality of lines of electrodes, each line of electrodes configured to measure a corresponding one of the plurality of nucleotide strands.

6. The system of any one of claims 1 to 5, wherein the aggregate electromagnetic signal comprises one or more of an electric charge, a current, and a voltage.

7. The system of any one of claims 1 to 6, wherein the total electromagnetic signal is further modulated by varying an input electromagnetic signal to the at least one electrode.

8. 8. The system of claim 1, wherein the mechanism is configured to move the nucleotide strand relative to the electrode in the longitudinal direction of the nucleotide strand to modulate the overall electromagnetic signal.

9. The at least one electrode comprises a plurality of electrodes, and the mechanism moves the nucleotide chain relative to the electrodes in a longitudinal direction of the nucleotide chain. (i) the length of the nucleotide chain of one of the electrodes in the longitudinal direction; and (ii) the distance between two adjacent electrodes in the longitudinal direction of the nucleotide chain 9. The system of claim 8, configured to move the object a distance equal to or greater than the sum of

10. The system of any one of claims 1 to 9, wherein the mechanism is configured to move the nucleotide strand at a selected frequency.

11. The system of claim 10 , wherein the frequency is variable by the mechanism.

12. 12. The system of claim 10 or 11, wherein the frequency is selected based on the spectral composition of the measured noise.

13. 13. The system of claim 1, wherein the at least one electrode comprises a plurality of electrodes, the electrodes being arranged such that there is an overlap in detection ranges between two adjacent ones of the electrodes.

14. measuring, with at least one electrode, an overall electromagnetic signal from a portion of the nucleotide chain comprising a plurality of nucleotides; moving the nucleotide strand relative to the at least one electrode to modulate the aggregate electromagnetic signal to identify electromagnetic signals associated with individual nucleotides in the plurality of nucleotides; A process for sequencing a chain of nucleotides, comprising:

15. 15. The process of claim 14, further comprising demodulating the measured total electromagnetic signal based on a movement signal associated with movement of the nucleotide strand relative to the at least one electrode.

16. 16. The process of claim 15, wherein the movement signal comprises a signal associated with movement of the nucleotide strand relative to the at least one electrode in the longitudinal direction of the nucleotide strand.

17. The nucleotide chain is aligned with the electrode in the longitudinal direction of the nucleotide chain. (i) the length of the nucleotide chain of one of the electrodes in the longitudinal direction; and (ii) the distance between two adjacent electrodes in the longitudinal direction of the nucleotide chain The process of any one of claims 14 to 16, wherein the object is moved a distance equal to or greater than the sum of

18. The process of any one of claims 14 to 17, wherein the nucleotide chain is moved at a selected frequency.

19. 20. The process of claim 18, wherein the frequency is selected based on the spectral composition of the measured noise.

20. The process of any one of claims 14 to 19, wherein the aggregate electromagnetic signal comprises one or more of an electric charge, a current, and a voltage.

21. 21. The process of any one of claims 14 to 20, further comprising varying an input electromagnetic signal to the at least one electrode to modulate the measured total electromagnetic signal.

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