WO2021197481A1

WO2021197481A1 - Gene sequencing array structure and gene sequencing apparatus

Info

Publication number: WO2021197481A1
Application number: PCT/CN2021/085322
Authority: WO
Inventors: 董晨洁; 蒋可; 苏云鹏; 邹耀中; 秦亚杰; 江逸舟
Original assignee: 成都今是科技有限公司; 复旦大学
Priority date: 2020-04-02
Filing date: 2021-04-02
Publication date: 2021-10-07
Also published as: CN113493735A; CN113493735B

Abstract

The present application provides a gene sequencing array structure and a gene sequencing apparatus. The gene sequencing array structure comprises at least one column unit. The column unit comprises: at least one detection unit, the detection unit comprising a test cavity and a first control unit connected to the test cavity; a redundant unit, the redundant unit comprising a redundant cavity and a second control unit connected to the redundant unit; and a readout circuit connected to the first control unit and the second control unit and configured to transfer and amplify accumulated charges of the test cavity by means of correlated double sampling of the accumulated charges of the test cavity and the redundant cavity. The gene sequencing apparatus can be used to determine the type of a nucleotide passing through a nanopore at a high speed, so as to achieve more accurate nucleic acid sequencing.

Description

Gene sequencing array structure and gene sequencing device

Technical field

The present disclosure belongs to the technical field of biological detection. Specifically, it relates to a gene sequencing array structure and a gene sequencing device, which can be used for high-speed detection of nucleotide base types to achieve sequencing, and is easy to expand on a large scale.

Background technique

The concept of nucleic acid sequencing based on nanopores was proposed in 1995. Researchers have discovered that certain transmembrane proteins, such as the bacterial toxin α-hemolysin, can form stable channels with a diameter of about 1-2 nanometers on the phospholipid membrane, called nanopores. Single-stranded DNA (or RNA) molecules will spontaneously pass through the nanopore in an electric field due to their own charged properties, and cause changes in the resistance of the nanopore during the passage, resulting in a so-called blocking current. The four different bases of DNA (RNA), A, T (U), C, and G, have recognizable differences in their blocking effects on current generation when they pass through the nanopore due to their own chemical structure differences, resulting in their own corresponding characteristics. Block the current. Accurate detection of the characteristic blocking current can determine the type of the corresponding base to determine the nucleic acid sequence.

There are two main methods for sequencing through nanopores. One method is represented by the Oxford Nanopore Technologies system, which directly allows single-stranded DNA molecules to pass through the nanopore and sequentially reads the characteristic blocking current corresponding to their bases. . However, due to the small difference in the characteristic currents given by different bases, multiple bases can stay in the nanopore at the same time, making the characterization of the blocking current very complicated, which puts forward extremely high requirements for the current data analysis in the later stage of sequencing. More importantly, this system has insurmountable difficulties for the determination of a continuous DNA sequence (homopolymer) of the same base. Another method is represented by the system adopted by Genia Technologies (currently belonging to Roche Sequencing Solutions), which uses modified nucleotide analogs to perform sequencing while nucleic acid synthesis. Although tagging the nucleotides used for replication can improve the recognition of the characteristic blocking current corresponding to different bases, the time interval at which a single nucleotide tag enters the nanopore also helps to determine the nucleic acid of the same consecutive base. Sequence (homopolymer), but this system is difficult to ensure that the tag of each nucleotide used for synthesis enters the nanopore to give a blocking current, which causes deletion errors in the sequencing process; it is also difficult to avoid nucleosides The acid tag blocks the current from being read, but the nucleotide itself does not really participate in the synthesis reaction, causing the signal to be read redundantly (insertion error). To solve this problem, in addition to detecting the nucleotide tag, it should be able to directly detect the nucleotide itself. Since nucleotides pass through the nanopore faster, a higher-speed detection circuit is needed to detect it.

Summary of the invention

The embodiments of the present disclosure provide a gene sequencing array structure and a gene sequencing device, which are used to achieve a higher speed sampling rate under the same noise level and more accurate nucleic acid sequencing.

In a first aspect, an embodiment of the present disclosure proposes a gene sequencing array structure, including: at least one column unit; the column unit includes:

At least one detection unit, the detection unit including a test cavity and a first control unit connected to the test cavity;

A redundant unit, the redundant unit including a redundant cavity and a second control unit connected to the redundant cavity;

The readout circuit is connected to the first control unit and the second control unit, and is used to transfer and amplify the accumulated charge of the test cavity through the correlated double sampling of the accumulated charge of the test cavity and the redundant cavity.

In an alternative embodiment, both the test chamber and the redundant chamber include a first compartment and a second compartment separated by a membrane, and a first electrode connected to the first compartment and a first electrode connected to the first compartment. The second electrode of the second compartment; the first electrode of the test cavity and the redundant cavity is connected to the common electrode terminal, the second electrode of the test cavity is connected to the first control unit, and the second electrode of the redundant cavity The electrode is connected to the second control unit; wherein the membrane of the test cavity has nanopores, and the membrane of the redundant cavity has no nanopores.

In an alternative embodiment, the first control unit and the second control unit both include a first reset switch and a readout switch; the first terminal of the first reset switch is connected to the column reference voltage, and the second terminal is connected to the The second electrode is used to reset the capacitor voltage of the film; the first end of the readout switch is connected to the second electrode, and the second end is connected to the readout circuit, which is used to reset the capacitor voltage of the film. The charge is guided to the readout circuit.

In an alternative embodiment, in the at least one column unit, the detection units or redundant units located in the same row are connected to a shared row reset signal, and the detection units and redundant units located in the same column are connected to the shared column reference voltage and Column output signal.

In an alternative embodiment, the column unit further includes a column reset prohibition switch, and the detection unit and the redundant unit are connected to a shared column reference voltage via the column reset prohibition switch.

In an alternative embodiment, the column unit further includes a column output prohibition switch, and the detection unit and the redundant unit are connected to the readout circuit via the column output prohibition switch.

In an alternative embodiment, the row reset signal is connected to the control terminal of the first reset switch of the detection unit or redundant unit in the same row, and is used to reset the detection unit or redundant unit in the same row. Capacitance voltage of the cell membrane.

In an alternative embodiment, the readout circuit includes:

The first amplifying circuit is used to transfer and amplify the membrane capacitance charges of the test cavity and the redundant cavity;

A correlated double sampling (CDS) circuit is used for correlated double sampling of the membrane capacitance charges of the test cavity and the redundant cavity to eliminate the offset voltage of the first amplifying circuit;

The second amplifying circuit is used to transfer and amplify the output of the CDS circuit.

In an alternative embodiment, the first amplifying circuit includes a first operational amplifier, a first feedback capacitor, and a second reset switch; wherein, the column reference voltage is input to the non-inverting input terminal of the first operational amplifier, The inverting input terminal is connected to the second terminal of the readout switch, the first feedback capacitor and the second reset switch are connected in parallel to the inverting input terminal and the output terminal of the first operational amplifier; The charge of the film capacitor is transferred and amplified under the action of the first feedback capacitor, and the second reset switch is used to reset the first feedback capacitor.

In an alternative embodiment, the CDS circuit includes a sampling capacitor, a sampling switch, a holding switch, a CDS capacitor, and a CDS sampling switch; wherein, the first end of the sampling switch is connected to the output of the first amplifying circuit, and The two ends are connected to the sampling capacitor and the first end of the CDS capacitor, and are used to guide the output voltage of the first amplifying circuit to the sampling capacitor or the CDS capacitor; the first end of the holding switch is connected to the The second end of the CDS capacitor is connected to the second amplifying circuit; the second end of the sampling capacitor is connected to the column reference voltage; the first end of the CDS sampling switch is connected to the second end of the CDS capacitor , The second terminal is connected to the column reference voltage for storing the output of the first amplifying circuit.

In an alternative embodiment, the second amplifying circuit includes a second operational amplifier, a second feedback capacitor, and a third reset switch; wherein the non-inverting input terminal of the second operational amplifier is connected to the column reference voltage , The second feedback capacitor and the third reset switch are connected in parallel with the inverting input terminal and the output terminal of the second operational amplifier; the second operational amplifier affects the CDS circuit under the action of the second feedback capacitor Charge transfer and amplification are performed on the output of, and the third reset switch is used to reset the second feedback capacitor.

In an alternative embodiment, the column unit further includes an analog-to-digital conversion circuit connected to the second amplifying circuit for converting the output of the second amplifying circuit into a digital signal and sampling.

In an alternative embodiment, the first control unit of the detection unit and the second control unit of the redundant unit located in the same column are sequentially connected to the readout circuit in a time division multiplexing manner.

In an alternative embodiment, in the at least one column unit, row reset signals located in different rows act sequentially in a time-division multiplexed manner to control the first reset switch to reset the detection units or redundant detection units in the same row. The membrane capacitor voltage of the remaining unit.

In the second aspect, an embodiment of the present disclosure proposes a gene sequencing device, including the gene sequencing array structure described in any of the foregoing embodiments.

The embodiment of the present disclosure adopts the method of charge transfer to directly read the accumulated charges of the film capacitance of the test cavity and the redundant cavity, and adopts the method of correlated double sampling to eliminate the offset voltage of the readout circuit amplification process, thereby reducing the sampling noise, so the same can be achieved Higher speed sampling rate under noise level, thereby improving the accuracy of nucleic acid sequencing.

Description of the drawings

In order to explain the embodiments of the present disclosure or the technical solutions in the prior art more clearly, the following will briefly introduce the drawings that need to be used in the description of the embodiments or the prior art one by one. Obviously, the drawings in the following description are For some of the embodiments of the present disclosure, for those of ordinary skill in the art, other drawings may be obtained based on these drawings without creative labor.

FIG. 1 is a schematic structural diagram of a single column unit in a gene sequencing array structure according to an embodiment of the present disclosure;

2 is a schematic diagram of an equivalent circuit model of a test cavity and a redundant cavity in a gene sequencing array structure according to an embodiment of the present disclosure;

3 is a schematic diagram of the circuit structure of a detection unit and a redundant unit in a gene sequencing array structure according to an embodiment of the present disclosure;

Figure 4 is a schematic diagram of a gene sequencing array structure according to an embodiment of the present disclosure;

5 is a schematic diagram of the circuit structure of a readout circuit according to an embodiment of the present disclosure;

FIG. 6 is a schematic diagram of signal waveforms according to the readout circuit shown in FIG. 5.

Detailed ways

In order to make the objectives, technical solutions, and advantages of the embodiments of the present disclosure clearer, the technical solutions in the embodiments of the present disclosure will be described clearly and completely in conjunction with the accompanying drawings in the embodiments of the present disclosure. Obviously, the described embodiments are part of the embodiments of the present disclosure, rather than all of the embodiments. Based on the embodiments in the present disclosure, all other embodiments obtained by those of ordinary skill in the art without creative work shall fall within the protection scope of the present disclosure.

In the present disclosure, it should be understood that terms such as "including" or "having" are intended to indicate the existence of the features, numbers, steps, actions, components, parts, or combinations thereof disclosed in this specification, and are not intended to exclude one The possibility of existence or addition of multiple other features, numbers, steps, behaviors, components, parts or combinations thereof.

The present disclosure proposes a gene sequencing array structure, which directly measures the accumulated charge stored in the detection unit and the redundant unit by means of charge transfer, and eliminates the offset voltage of the amplification process through correlated double sampling to reduce noise, so that it can be used for high-speed passing The nucleotide type of the nanopore is judged to achieve more accurate nucleic acid sequencing.

Fig. 1 is a schematic structural diagram of a single column unit in a gene sequencing array structure according to an embodiment of the present disclosure. As shown in FIG. 1, a single column unit in the gene sequencing array structure of the embodiment of the present disclosure includes:

At least one detection unit including a test cavity 102 and a control unit 104 connected to the test cavity 102;

A redundant unit, which includes a redundant cavity 108 and a control unit 104 connected to the redundant cavity;

The readout circuit 110, which connects the detection unit and the control unit 104 of the redundant unit, is used to accumulate the charge in the test cavity through correlated double sampling (CDS) of the accumulated charge of the test cavity 102 and the redundant cavity 108 Perform transfer and enlargement.

In the embodiment of the present disclosure, the readout circuit transfers and amplifies the charges accumulated in the test cavity 102 of the detection unit and the redundant cavity 108 of the redundant unit, and uses correlated double sampling to eliminate the offset voltage in the amplification process to reduce noise. When the column units constitute a gene sequencing array structure, the gene sequencing array structure can be used to judge the type of nucleotides passing through the nanopore at high speed, so as to achieve more accurate nucleic acid sequencing.

Wherein, the test chamber 102 includes a first compartment 106 and a second compartment 107 separated by a membrane 109, and a first electrode 101 (common electrode) connected to the first compartment 106 and a first electrode 101 (common electrode) connected to the second compartment 107 The second electrode 103 (working electrode), the first electrodes 101 of all test chambers are connected to the same command voltage VCMD, and the second electrode 103 is connected to the control unit 104. In one embodiment, the membrane 109 may be a phospholipid bimolecular membrane with nanopores 105 thereon.

Similarly, the redundant cavity 108 also includes a first compartment 106 and a second compartment 107 separated by a membrane 109, and a first electrode 101 (common electrode) connected to the first compartment 106 and connected to the second compartment The second electrode 103 (working electrode) of the chamber 107, the first electrode 101 is connected to the same command voltage VCMD, and the second electrode 103 is connected to the control unit 104. Unlike the test cavity 102, the membrane 109 in the redundant cavity 108 does not have nanopores 105.

The control unit 104 connected to the test cavity 102 is used to periodically reset the nanoporous membrane capacitance in the test cavity 102 to the column reference voltage, and form a positive or negative voltage difference with the first electrode 101 to promote nucleotide molecules The nanopore 105 on the membrane 109 moves bidirectionally between the first compartment 106 and the second compartment 107. As the nucleotide molecules pass through the nanopore 105, different nanopore resistances (for example, 250MΩ-20GΩ) will be formed, and the membrane capacitance will be charged or discharged, and thus different characteristic voltages will be formed in the sampling period (for example, 100μs).

Similarly, the control unit 104 connected to the redundant cavity 108 is used to periodically reset the film capacitance in the redundant cavity 108 to the column reference voltage.

In an alternative embodiment, as shown in FIG. 1, the control units 104 in different detection units and redundant units can also be connected to the readout circuit 110 sequentially in a time-division multiplexing manner, corresponding to the test cavity 102 and the redundant cavity. The charge accumulated on the film 109 in 108 is transferred to the readout circuit 110 for filtering and amplification, and finally output to the analog-to-digital converter 111 for sampling.

FIG. 2 is a schematic diagram of an equivalent circuit model of a test cavity and a redundant cavity in the array structure of gene sequencing according to an embodiment of the present disclosure. As shown in Figure 2, in terms of electrical characteristics, the test cavity 201 with nanopores can be equivalent to the nanopore equivalent model 217, where the common electrode 202 corresponds to the common electrode 207, the working electrode 203 corresponds to the working electrode 210, and the phospholipid double The molecular membrane 205 and the nanopore 204 are equivalent to the equivalent membrane capacitance 206 (for example, 1~2pF) and the parallel nanopore equivalent resistance 209 (for example, 250MΩ~20GΩ). The solution in the first compartment 205 is equivalent to the first The compartment equivalent resistance 208 (for example, 1KΩ-10KΩ), and the solution in the second compartment 206 is equivalent to the second compartment equivalent resistance 211 (for example, 1KΩ-10KΩ).

The redundant cavity without nanoholes (108 in FIG. 1), since there are no nanoholes, it is equivalent to the redundant unit equivalent model 218. The common electrode 202 corresponds to the common electrode 213, the working electrode 203 corresponds to the working electrode 215, the phospholipid bimolecular membrane 205 is equivalent to the equivalent membrane capacitance 212 (for example, 1 to 2 pF), and the solution in the first compartment 205 is equivalent to the first The equivalent resistance of one compartment 214 (for example, 1KΩ-10KΩ), and the solution in the second compartment 206 is equivalent to the equivalent resistance of the second compartment 216 (for example, 1KΩ-10KΩ).

In an alternative embodiment, as shown in FIG. 3, the detection unit 301 includes a reset switch 303, the first end of which is connected to the column reference voltage 302, and the second end is connected to the working electrode 210 of the test cavity equivalent model 217. It also includes a readout switch 304, the first end of which is connected to the working electrode 210 of the test cavity equivalent model 217, and the second end is connected to the readout circuit 110. The reset switch 303 is used to periodically reset the equivalent film capacitor 206, and the readout switch 304 is used to periodically transfer the charge accumulated in the equivalent film capacitor 206 to the readout circuit 110. In an alternative embodiment, the control terminal (not shown in FIG. 3) of the reset switch 303 may be connected to a reset signal, and the reset switch 303 periodically resets the equivalent film capacitance 206 under the action of the reset signal. The reset period of the reset switch 303 and the read period of the read switch 304 are determined by the system sampling rate and the acceptable system signal-to-noise ratio. In an alternative embodiment, the reset period and the readout period are at least 100 μs.

In an alternative embodiment, as shown in FIG. 3, the redundant unit 308 includes a reset switch 305, the first end of which is connected to the column reference voltage 306, and the second end is connected to the working electrode 215 of the redundant cavity equivalent model 218. It also includes a readout switch 307, the first end of which is connected to the working electrode 215 of the redundant cavity equivalent model 218, and the second end is connected to the readout circuit 110. The reset switch 305 is used to periodically reset the equivalent film capacitor 212, and the readout switch 307 is used to periodically transfer the charge accumulated in the equivalent capacitor 212 to the readout circuit 110. In an alternative embodiment, the control terminal (not shown in FIG. 3) of the reset switch 305 may be connected to a reset signal, and the reset switch 305 periodically resets the equivalent film capacitance 212 under the action of the reset signal. The reset period of the reset switch 305 and the read period of the read switch 307 are determined by the system sampling rate and the acceptable system signal-to-noise ratio. In an alternative embodiment, the reset period and the readout period are at least 100 μs.

Fig. 4 is a schematic diagram of a gene sequencing array structure according to an embodiment of the present disclosure. As shown in FIG. 4, in the gene sequencing array structure of this embodiment, the detection unit 406 (corresponding to the detection unit 301 in FIG. 3) and the redundant unit 407 (corresponding to the redundant unit 308 in FIG. 3) can pass through shared rows The reset signal 401, the shared column reference voltage signal 403, and the shared column output signal 404 constitute an array.

Wherein, the shared row reset signal 401 is connected to the control terminal of the first reset switch 303 of each detection unit 406 in the same line, and is used to control its opening and closing to achieve the purpose of resetting the film capacitance of the corresponding detection unit, or is connected to the same line The control terminal of the reset switch 305 of each redundant unit 407 is used to control its opening and closing so as to achieve the purpose of resetting the film capacitance of the corresponding redundant unit. The shared column reference voltage 403 is connected to the first end of the reset switch 303 of each detection unit 406 of each column and the first end of the reset switch 305 of the redundant unit 407, and is used to close the reset switch 303 or the reset switch 305. At this time, the column reference voltage is transferred to the working electrode of the corresponding detection unit or redundant unit. The row reset signals 401 of different rows act sequentially in a time-division multiplexed manner. The time width of time division multiplexing is determined by the number of detection units and redundant units contained in each column and the sampling period. The column reference voltage 403 shared by each column remains unchanged during the operation of the array. The column output signal 404 shared by each column is at the same time when the cells of each row are reset, according to the scanning direction of the pipeline array, the readout switch 304 of the detection unit of the adjacent row (the upper row or the next row) or the readout of the redundant cell is closed. The switch 307 outputs the accumulated charge of the membrane capacitor of the corresponding cell to the readout circuit 406, so as to ensure that the charge and discharge time of each cell from reset to charge output is maximized in a pipeline operation mode.

In an alternative embodiment, as shown in FIG. 4, since the working state of the corresponding unit may be abnormal, the unit needs to be prohibited during the working process. Therefore, a column reset prohibition switch 402 can be further added to the column reference voltage input terminal. The first end of the column reset prohibition switch 402 is connected to the column reference voltage 403, and the second end is connected to the first end of the reset switch 303 of the detection unit 406 in each column. The terminal and the first terminal of the reset switch 305 of the redundant unit 407 are used to disconnect the reference voltage from the outside of the array when resetting a certain row of cells. Correspondingly, a column output prohibition switch 405 can be added in the middle where the column output signal 404 is connected to the readout circuit 406. The first end of the column output prohibition switch 405 is connected to the shared column output signal 404, and the second end is connected to the readout circuit 406. It is used to disconnect the connection between the cell and the readout circuit when the cell output of a certain row is selected. The column reset prohibition switch 402 and the column output prohibition switch 405 can ensure that the working electrode 210 of the detection unit that needs to be prohibited is always in a floating state during reset, readout, and charging and discharging, thereby ensuring that the unit's nanopore is prohibited without current passing through.

In an alternative embodiment, as shown in FIG. 5, the readout circuit includes a first amplifying circuit, a correlated double sampling (CDS) circuit and a second amplifying circuit.

Correlated double sampling (CDS) is a noise suppression technology commonly used in CCD imaging. Because the output signal of each pixel of the CCD contains both a photosensitive signal and a reset pulse signal, if at the beginning of the integration of the photoelectric signal and At the end of integration, the output signal is sampled separately, and the noise voltages of the two samples are almost the same. Subtracting the two sampled values can basically eliminate the interference of reset noise and obtain the actual effective amplitude of the signal level. The embodiment of the present disclosure introduces the CDS circuit in the readout circuit based on the CDS sampling principle, which is used to transfer, amplify, store and subtract the film capacitance charges in the test cavity of the detection unit and the redundant cavity of the redundant unit, and eliminate the first An amplifying circuit generates offset voltage and low-frequency noise, thereby reducing the overall noise of the circuit.

The first amplifier circuit includes an operational amplifier 513, a feedback capacitor 512 (Cf1), and a reset switch 511 (RST1). The non-inverting input terminal of the operational amplifier 513 is connected to the column reference voltage VCM514, and the inverting input terminal is connected to the column output inhibit switch. The second end of 523, the first end of the column output prohibition switch 523 is connected to the second end of the readout switch 507 (corresponding to 304 in Figure 3) of the detection unit in the array and the readout switch 510 of the redundant unit (corresponding to Figure 3 307) the second terminal, the feedback capacitor 512 and the reset switch 511 are connected in parallel to the inverting input and output terminals of the operational amplifier 513; the operational amplifier 513 transfers the charge stored in the detection unit 406 in the array under the action of the feedback capacitor 512 The amplification and reset switch 511 is used to periodically reset the feedback capacitor 512. The typical value of the feedback capacitor 512 is 100μF. Considering the constant nanopore resistance, for a fixed nanopore electrode pressure difference, the charge and discharge amplitude of the nanopore is inversely proportional to the membrane capacitance, and the gain of the first amplifier circuit is inversely proportional to the membrane capacitance. The value is proportional, so the amplified signal finally obtained by the first amplifying circuit is only related to the nanopore resistance and has a one-to-one correspondence, and has nothing to do with the film capacitance value.

The CDS circuit includes a sampling capacitor 516, a sampling switch 515, a holding switch 519, a CDS capacitor 517, and a CDS sampling switch 518. Among them, the first end of the sampling switch 515 is connected to the output end of the operational amplifier 513, and the second end is connected to the first end of the sampling capacitor 516 and the CDS capacitor 517, which is used for sampling or introducing the voltage of the first amplifying circuit to the sampling capacitor during CDS sampling. 516 or CDS capacitor 517. The first end of the holding switch 519 is connected to the second end of the CDS capacitor 517 for transferring the charge stored in the sampling capacitor 516 and the CDS capacitor 517 after CDS sampling to the second amplifying circuit. The first end of the sampling capacitor 516 is connected to the second end of the sampling switch 515, and the second end is connected to the column reference voltage VCM514. The first end of the CDS capacitor 517 is connected to the second end of the sampling switch 515, and the second end is connected to the first end of the holding switch 519. The first end of the CDS sampling switch 518 is connected to the second end of the CDS capacitor 517, and the second end is connected to the column reference voltage VCM514 for storing the output of the first amplifying circuit during CDS sampling.

The second amplifying circuit includes an operational amplifier 522, a feedback capacitor 521 (Cf2), and a reset switch 520 (RST2). The non-inverting input terminal of the operational amplifier 522 is connected to the column reference voltage VCM514, and the inverting input terminal is connected to the second terminal of the holding switch 519. The feedback capacitor 521 and the reset switch 520 are connected in parallel to the inverting input terminal and the output terminal of the operational amplifier 522. The operational amplifier 522 performs charge transfer and further amplification of the output of the CDS circuit under the action of the feedback capacitor 521. The reset switch 520 is used to periodically reset the feedback capacitor 521. The gain of the second amplifying circuit is determined by the ratio of the capacitance value of the series capacitance of the CDS capacitor 517 and the sampling capacitor 516 to the capacitance value of the feedback capacitor 521. The capacitance of the feedback capacitor 521 is typically 100μF, and the gain of the second amplifying circuit is typically 2 ~3, so the typical value of the series capacitance of the CDS capacitor 517 and the sampling capacitor 516 is 200-300 μF.

FIG. 6 is a schematic diagram of signal waveforms according to the readout circuit shown in FIG. 5, showing the specific timing of the readout circuit shown in FIG. 5 during operation. As shown in Figure 6, for any detection unit in the array, its working sequence is divided into automatic zero 602, sampling 603, and holding 604. The total length unit sampling period T601 is the system sampling period/the number of column detection units. Unless otherwise specified, the column output prohibition switch 523 is always closed in the subsequent description. In the waveform diagram, RST is the control signal of the column reset switch 506 in FIG. 5, RST_DUMMY is the control signal of the redundant unit reset switch 508 in FIG. 5, SMP is the control signal of the column readout switch 507 in FIG. 5, and SMP_DUMMY is the control signal in FIG. The redundant unit reads the control signal of the switch 510, RST1 is the control signal of the reset switch 511 of the first amplifying circuit in FIG. 5, RST2 is the control signal of the reset switch 520 of the second amplifying circuit in FIG. 5, and CDS2 is the CDS sampling in FIG. For the control signal of the switch 518, the above switch control signal turns on the corresponding switch when it is at a high level, and turns off the corresponding switch when it is at a low level. CDS1 is the control signal of the sampling switch 515 and the holding switch 519 in FIG. 5, the holding switch 519 is closed when the high level, the sampling switch 515 is open, and the holding switch 519 is opened at the low level, and the sampling switch 515 is closed.

Among them, when the circuit is in the auto-zero 602 state, the sampling switch 515 and the CDS sampling switch 518 are always closed, the holding switch 519 is open, and the column readout switch 507 is open. The reset switch 511 first quickly resets the feedback capacitor 512 and then turns off, and then turns on the redundant unit readout switch 510. The stored voltage value of the membrane capacitor in the redundant cavity 505 and the possible offset voltage of the first amplifying circuit are superimposed and transferred to the CDS capacitor 517 and amplify, and then turn off the redundant cell readout switch 510 and the CDS sampling switch 518. Since the voltage stored before the redundant cavity sampling is the reference voltage VCM, the offset value of the amplified first amplifying circuit is stored in the CDS capacitor 517 at this time.

When the circuit is in the sampling state 603, the reset switch 511 first quickly resets the feedback capacitor 512 and then opens, and then turns on the detection unit column readout switch 507. Since the sampling switch 515 is closed, the membrane capacitor in the test cavity 502 is in the sampling period. The voltage accumulated in will be transferred to the sampling capacitor 516 and amplified. Before entering the holding state 604, the second amplifying circuit and the redundant unit circuit will be reset. In addition, when the charge of the detection unit is read, the detection unit in the previous row has been read out in the previous sampling period, and the current period will be restarted and the next round of charging and discharging will start. At the end of this state, what is stored in the sampling capacitor 516 is the amplified result of the accumulated voltage of the film capacitor of the test cavity superimposed on the offset voltage of the operational amplifier 513.

When the circuit is in the holding state 604, the detection unit column readout switch 507 is opened, the sampling switch 515 is opened, and the holding switch 519 is closed. At this time, the voltage stored in the sampling capacitor 516 will be reduced by the voltage stored in the CDS capacitor 517. It is transferred to the feedback capacitor 521 for further amplification, and the final voltage is formed and sent to the analog-to-digital conversion unit for sampling. Since the films of the redundant unit and the detection unit are generated under the same biochemical conditions, they can be expected to have similar capacitance values. Therefore, the gain multiples of the first amplifying circuit for the detection unit and the redundant unit should be approximately equal. Through the conversion of the CDS circuit, it can be considered that the offset voltage of the operational amplifier 513 will be basically eliminated in the output of the operational amplifier 522.

The present disclosure also provides a gene sequencing device, which includes the array structure as described in any of the foregoing embodiments.

The gene sequencing array structure and device of the embodiment of the present disclosure directly measure the accumulated charge of the test cavity and the redundant cavity by means of charge transfer, and adopt the method of correlated double sampling to eliminate the offset voltage during the amplification process of the readout circuit and reduce the detection noise , So that the type of nucleotides passing through the nanopore at high speed can be judged to achieve more accurate nucleic acid sequencing.

It should be noted that the above embodiments can be freely combined as required. The above are only the preferred embodiments of the present disclosure. It should be pointed out that for those of ordinary skill in the art, without departing from the principles of the present disclosure, several changes and improvements can be made, and these changes and improvements are also It should be regarded as falling into the protection scope of the present disclosure.

Claims

A gene sequencing array structure, characterized in that it comprises: at least one column unit; the column unit comprises:

At least one detection unit, the detection unit including a test cavity and a first control unit connected to the test cavity;

A redundant unit, the redundant unit including a redundant cavity and a second control unit connected to the redundant cavity;

The readout circuit is connected to the first control unit and the second control unit, and is used to transfer and amplify the accumulated charge of the test cavity through the correlated double sampling of the accumulated charge of the test cavity and the redundant cavity.
The gene sequencing array structure according to claim 1, wherein the test cavity and the redundant cavity each comprise a first compartment and a second compartment separated by a membrane, and a connection connected to the first compartment The first electrode and the second electrode connected to the second compartment; the first electrode of the test cavity and the redundant cavity are connected to the common electrode terminal, and the second electrode of the test cavity is connected to the first control unit, The second electrode of the redundant cavity is connected to the second control unit; wherein the membrane of the test cavity has nanopores, and the membrane of the redundant cavity has no nanopores.
The gene sequencing array structure according to claim 2, wherein the first control unit and the second control unit both comprise a first reset switch and a readout switch; the first end of the first reset switch is connected to the column Reference voltage, the second terminal is connected to the second electrode for resetting the capacitance voltage of the film; the first terminal of the readout switch is connected to the second electrode, and the second terminal is connected to the readout circuit, It is used to guide the capacitive charge of the film to the readout circuit.
The gene sequencing array structure according to claim 3, wherein in the at least one column unit, the detection units or redundant units located in the same row are connected to a shared row reset signal, and the detection units located in the same column and redundant units are connected to a shared row reset signal. The cells are connected to the shared column reference voltage and column output signal.
The gene sequencing array structure according to claim 4, wherein the column unit further comprises a column reset prohibition switch, and the detection unit and the redundant unit are connected to a shared column reference voltage via the column reset prohibition switch.
The gene sequencing array structure according to claim 5, wherein the column unit further comprises a column output prohibition switch, and the detection unit and the redundant unit are connected to the readout circuit via the column output prohibition switch.
The gene sequencing array structure according to claim 4, wherein the row reset signal is connected to the control terminal of the first reset switch of the detection unit or redundant unit of the same row, and is used to reset the The capacitance voltage of the membrane of the detection unit or redundant unit in the same row.
The gene sequencing array structure of claim 3, wherein the readout circuit comprises:

The first amplifying circuit is used to transfer and amplify the membrane capacitance charges of the test cavity and the redundant cavity;

A correlated double sampling (CDS) circuit is used for correlated double sampling of the membrane capacitance charges of the test cavity and the redundant cavity to eliminate the offset voltage of the first amplifying circuit;

The second amplifying circuit is used to transfer and amplify the output of the CDS circuit.
The gene sequencing array structure of claim 8, wherein the first amplifying circuit comprises a first operational amplifier, a first feedback capacitor, and a second reset switch; wherein, the non-inverting input of the first operational amplifier The column reference voltage is input to the terminal, the inverting input terminal is connected to the second terminal of the readout switch, the first feedback capacitor and the second reset switch are connected in parallel to the inverting input terminal and the output terminal of the first operational amplifier The first operational amplifier transfers and amplifies the charge of the film capacitor under the action of the first feedback capacitor, and the second reset switch is used to reset the first feedback capacitor.
The gene sequencing array structure according to claim 8, wherein the CDS circuit comprises a sampling capacitor, a sampling switch, a holding switch, a CDS capacitor, and a CDS sampling switch; wherein the first end of the sampling switch is connected to the The output of the first amplifying circuit, the second end is connected to the first end of the sampling capacitor and the CDS capacitor, and is used to guide the output voltage of the first amplifying circuit to the sampling capacitor or the CDS capacitor; The first end of the holding switch is connected to the second end of the CDS capacitor, and the second end is connected to the second amplifying circuit; the second end of the sampling capacitor is connected to the column reference voltage; the first end of the CDS sampling switch The terminal is connected to the second terminal of the CDS capacitor, and the second terminal is connected to the column reference voltage for storing the output of the first amplifying circuit.
The gene sequencing array structure of claim 8, wherein the second amplifying circuit comprises a second operational amplifier, a second feedback capacitor, and a third reset switch; wherein the non-inverting input of the second operational amplifier Terminal is connected to the column reference voltage, the second feedback capacitor and the third reset switch are connected in parallel with the inverting input terminal and the output terminal of the second operational amplifier; the second operational amplifier is connected to the second feedback capacitor The output of the CDS circuit is charged and amplified under the action of, and the third reset switch is used to reset the second feedback capacitor.
The gene sequencing array structure according to claim 8, wherein the column unit further comprises an analog-to-digital conversion circuit, and the analog-to-digital conversion circuit is connected to the second amplifying circuit for amplifying the second amplifying circuit. The output of the circuit is converted into a digital signal and sampled.
The gene sequencing array structure according to claim 7, wherein the first control unit of the detection unit and the second control unit of the redundant unit located in the same column are sequentially connected to the readout unit in a time division multiplexing manner. Circuit.
The gene sequencing array structure according to claim 13, wherein in the at least one column unit, row reset signals located in different rows act sequentially in a time-division multiplexed manner to control the reset switch of the first reset switch. Describe the film capacitor voltage of the detection unit or redundant unit in the same row.
A gene sequencing device, characterized by comprising the gene sequencing array structure according to any one of claims 1-14.