Molecular detection unit and chip

Abstract

The utility model discloses a molecule detection unit and a chip. The molecule detection unit comprises a substrate, a first structural layer, a second structural layer, a sensing electrode, a third structural layer, a first buffer solution flow channel, a sample flow channel, a single-hole liquid storage cavity, a liquid resistance flow channel and a second buffer solution flow channel, the liquid resistance flow channel and the single-hole liquid storage cavity are arranged in the first structure layer, and the bottom of the single-hole liquid storage cavity is communicated with one end of the liquid resistance flow channel; the sample runner is arranged in the second structural layer and is communicated with the single-hole liquid storage cavity; the first buffer solution runner is arranged in the third structural layer; the bottom of the second buffer solution runner is communicated with the other end of the liquid resistance runner, and the top of the second buffer solution runner is communicated with the first buffer solution runner; the sensing electrode is arranged at the communication position of the single-hole liquid storage cavity and the liquid resistance flow channel. According to the invention, the arrangement density of the nanopore device in unit area can be improved, the chip manufacturing efficiency is improved, and the flux of the nanopore sequencing device is improved.

Description

TECHNICAL FIELD

[0001] The present application relates to the technical field of biological detection, and in particular to a molecular detection unit and a chip. BACKGROUND

[0002] A nanopore sequencing device is a device that determines the base pair sequence of a DNA molecule by detecting the change in electrical signal generated when the DNA molecule passes through a nanopore. This type of device generally includes a thin film that can isolate two reservoirs and a nanopore inlaid in the thin film. The nanopore can use a protein molecule with a pore structure that exists in nature or is artificially compiled (for example, a nanopore protein), or can be physically machined by an ion beam or the like to form a hole in the thin film. When a nanopore sequencing device is used to sequence a biological molecule (for example, a DNA molecule), the reservoirs on both sides of the thin film are at different potentials to drive the DNA molecule to pass through the nanopore. When the target DNA for sequencing passes through the nanopore, the electrical signal between the two reservoirs is recorded at the same time. By analyzing these signals, the sequencing of the DNA structure can be completed.

[0003] In the prior art, the nanopore device mainly uses the method of monitoring the change in ionic current through the nanopore protein between the two reservoirs to calculate the base pair sequence of the target DNA for sequencing. The length of the DNA molecule in the stretched state is relatively long, and it takes a certain amount of time for the DNA molecule to pass through the nanopore. Within a certain period of time, only a small part of the base sequence of the DNA molecule is in the nanopore. Since each base has a different effect on the change in ionic current, the size of the ionic current will change accordingly due to the change in the small base sequence in the nanopore. Therefore, by analyzing the change in ionic current during the entire process of the DNA passing through the nanopore, the base sequence of the target DNA for sequencing can be calculated using a corresponding calculation method.

[0004] When using a sequencing method based on ionic current, the nanopore device of this type has a relatively high requirement for the accuracy of the detection of ionic current. The nanopore protein used in the biological nanopore sequencing device in the prior art has a resistance of about 1 gigohm in the sequencing environment, which makes the size of the ionic current during sequencing about 100 picosiemens. Therefore, changes in the device structure and the test environment can significantly affect the accuracy of the measurement of ionic current, thereby placing higher requirements on the amplification performance and stability of the device circuit. At the same time, the measurement of ionic current is affected by the size of the electrode area, thereby limiting the total number of nanopore devices per unit area, and thereby limiting the size of the throughput of commercial nanopore devices.

[0005] In addition, in the voltage sequencing method using the principle of a voltage divider in the prior art, a buffer flow channel connected by a liquid resistance flow channel is added to a liquid storage cavity on one side of a film embedded with a nanopore to form a voltage divider, so that the device can obtain the resistance change in the nanopore by detecting the potential change of the liquid storage cavity on one side of the film, and then calculate the base sequence of the sequencing target DNA through a corresponding method.

[0006] In the prior art, the liquid resistance flow channel and the buffer flow channel are usually realized by forming a through hole in a substrate. However, in the field of micro-nano processing technology, the commonly used through hole forming method has certain requirements for the thickness of the material and the geometry of the through hole. Most of the nanopore sequencing devices in the prior art are based on silicon wafers, and the thickness is usually several hundred microns, and the required hole diameter in the structure is usually in the range of several microns. In the existing mature high aspect ratio micro-nano processing technology, due to factors such as load effect and mask layer area, only deep hole etching with a maximum aspect ratio of 15:1 can be effectively completed in actual production. Therefore, how to effectively manufacture such a through hole structure in the substrate has become an important technical problem for the current realization of the chip manufacturing and mass production of the voltage sequencing principle. Practical new type

[0007] Embodiments of the present disclosure provide a molecular detection unit and a chip.

[0008] In a first aspect, embodiments of the present disclosure provide a molecular detection unit, comprising: a substrate, a first structure layer, a second structure layer, a sensing electrode, a third structure layer, a first buffer flow channel, a sample flow channel, a single-hole liquid storage cavity, a liquid resistance flow channel, and a second buffer flow channel.

[0009] The first structure layer is arranged on the top of the substrate;

[0010] The second structure layer is arranged above the first structure layer and the insulating layer;

[0011] The third structure layer is arranged above the second structure layer;

[0012] The liquid resistance flow channel and the single-hole liquid storage cavity are arranged in the first structure layer;

[0013] The bottom of the single-hole liquid storage cavity is in communication with one end of the liquid resistance flow channel;

[0014] The sample flow channel is arranged in the second structure layer, above the single-hole liquid storage cavity and in communication with the single-hole liquid storage cavity;

[0015] The first buffer flow channel is arranged in the third structure layer;

[0016] The bottom of the second buffer liquid flow channel is communicated with the other end of the liquid resistance flow channel; and the top of the second buffer liquid flow channel is communicated with the first buffer liquid flow channel.

[0017] The sensing electrode is arranged at the communication position of the single-hole liquid storage cavity and the liquid resistance flow channel.

[0018] In a second aspect, embodiments of the present disclosure provide a molecular detection chip, comprising: a molecular detection array;

[0019] The molecular detection array comprises at least one molecular detection unit group; each molecular detection unit group comprises a plurality of molecular detection units according to any one of the preceding embodiments;

[0020] The sample flow channels of the molecular detection units are communicated with each other to form a common sample flow channel; and / or,

[0021] The first buffer liquid flow channels of the molecular detection units are communicated with each other to form a first common buffer liquid flow channel; and / or,

[0022] The second buffer liquid flow channels of the molecular detection units are communicated with each other to form a second common buffer liquid flow channel.

[0023] It should be understood that the content described in this part is not intended to identify key or important features of the embodiments of the present disclosure, nor to limit the scope of the present disclosure. Other features of the present disclosure will become apparent from the following description. BRIEF DESCRIPTION OF DRAWINGS

[0024] The accompanying drawings are used to better understand the present scheme, and do not constitute a limitation on the present disclosure. Among them:

[0025] Figure 1 It is a structure schematic diagram of the molecular detection unit in an embodiment of the present disclosure.

[0026] Figure 2 It is a structure schematic diagram of the molecular detection unit in an embodiment of the present disclosure. Figure 1

[0027] Figure 3 It is a structure schematic diagram of the molecular detection unit in another embodiment of the present disclosure.

[0028] Figure 4 It is a structure schematic diagram of the molecular detection unit in another embodiment of the present disclosure.

[0029] Figure 5 It is a structure schematic diagram of the molecular detection unit provided with a film layer in an embodiment of the present disclosure.

[0030] Figure 6 It is a working principle schematic diagram of the molecular detection unit in an embodiment of the present disclosure.​

[0031] Figure 7 This is a schematic diagram of the molecular detection unit in another specific embodiment of the present disclosure.

[0032] Figure 8 for Figure 7 A schematic diagram of the BB' cross-section.

[0033] Figure 9 This is a schematic diagram of the structure of a molecular detection unit with a membrane layer in another specific embodiment of this disclosure.

[0034] Figure 10 This is a schematic diagram of the molecular detection array in a specific embodiment of the present disclosure.

[0035] Figure 11 for Figure 10 A schematic diagram of the CC' section.

[0036] Figure 12 This is a schematic diagram of the molecular detection array in another specific embodiment of the present disclosure.

[0037] Figure 13 for Figure 12 A schematic diagram of the DD' cross-section.

[0038] Figure 14 This is a schematic diagram of the structure of a molecular detection array with a membrane layer in another specific embodiment of this disclosure.

[0039] Figure 15 This is a schematic diagram of the molecular detection array in another specific embodiment of the present disclosure.

[0040] Figure 16 for Figure 15 A schematic diagram of the EE' cross-section.

[0041] Figure 17 This is a schematic diagram of the structure of a molecular detection array with a membrane layer in another specific embodiment of this disclosure.

[0042] Figure 18 This is a schematic diagram of a molecular detection array in another specific embodiment of this disclosure.

[0043] Figure 19 This is a schematic diagram of the fabrication process of the molecular detection chip in a specific embodiment of this disclosure. Figure 1 .

[0044] Figure 20 This is a schematic diagram of the fabrication process of the molecular detection chip in a specific embodiment of this disclosure. Figure 2 .

[0045] Figure 21Schematic diagram of a preparation process of a molecular detection chip in one embodiment of the present disclosure Figure 3 .

[0046] Figure 22 Schematic diagram of a preparation process of a molecular detection chip in one embodiment of the present disclosure Figure 4 .

[0047] Figure 23 Schematic diagram of a preparation process of a molecular detection chip in one embodiment of the present disclosure Figure 5 .

[0048] Figure 24 Schematic diagram of a preparation process of a molecular detection chip in one embodiment of the present disclosure Figure 6 .

[0049] Figure 25 Schematic diagram of a preparation process of a molecular detection chip in one embodiment of the present disclosure Figure 7 .

[0050] Figure 26 Schematic diagram of a preparation process of a molecular detection chip in one embodiment of the present disclosure Figure 8 .

[0051] Figure 27 Schematic diagram of a preparation process of a molecular detection chip in one embodiment of the present disclosure Figure 9 .

[0052] Figure 28 Schematic diagram of a preparation process of a molecular detection chip in one embodiment of the present disclosure Figure 10 .

[0053] Figure 29 Schematic diagram of a preparation process of a molecular detection chip in one embodiment of the present disclosure Figure 1 . DETAILED DESCRIPTION

[0054] It should be noted that the following detailed description is merely exemplary and is intended to provide further description of the present disclosure. Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this present disclosure belongs.

[0055] It is to be understood that the singular forms “a,” “an,” and “the” include plural referents unless the context clearly dictates otherwise. It should be noted that the terms “comprises,” “comprising,” “includes,” “including,” and the like, when used in this specification, specify the presence of stated features, steps, operations, devices, components, and / or combinations thereof, but do not preclude the presence or addition of one or more other features, steps, operations, devices, components, and / or combinations thereof.

[0056] The embodiments in the present disclosure and the features in the embodiments can be combined with each other in the case of no conflict.

[0057] In order to make the technical solutions and advantages of the present disclosure clearer, the present disclosure will be further described in detail below with reference to the drawings and specific embodiments.

[0058] A molecular detection unit is provided in the present disclosure.

[0059] As shown in Figure 2 and Figure 5 In one specific embodiment in the present disclosure, the molecular detection unit can include a substrate 1, a first structure layer 2, a second structure layer 3, a sensing electrode 4, a third structure layer 5, a first buffer flow channel 101, a sample flow channel 102, a single-hole liquid storage cavity 103, a liquid resistance flow channel 104, and a second buffer flow channel 105.

[0060] The first structure layer 2 is arranged on top of the substrate 1.

[0061] The second structure layer 3 is arranged above the first structure layer 2.

[0062] The third structure layer 5 is arranged above the second structure layer 3.

[0063] The liquid resistance flow channel 104 and the single-hole liquid storage cavity 103 are arranged in the first structure layer 2.

[0064] The bottom of the single-hole liquid storage cavity 103 is in communication with one end of the liquid resistance flow channel 104.

[0065] The sample flow channel 102 is arranged in the second structure layer 3, above the single-hole liquid storage cavity 103 and in communication with the single-hole liquid storage cavity 103.

[0066] The first buffer flow channel 101 is arranged in the third structure layer 5.

[0067] The bottom of the second buffer flow channel 105 is in communication with the other end of the liquid resistance flow channel 104, and the top of the second buffer flow channel 105 is in communication with the first buffer flow channel 101.

[0068] The sensing electrode 4 is arranged at the communication between the single-hole liquid storage cavity 103 and the liquid resistance flow channel 104.

[0069] In addition, in the technical solutions of the present disclosure, a corresponding membrane layer and nanopore can be further arranged in the above-mentioned molecular detection unit according to the needs of actual application scenarios.

[0070] The film layer can separate the single-hole liquid storage cavity from the sample channel, and the nanopore can liquidly connect the sample flow channel and the single-hole liquid storage cavity.

[0071] For example, as an example, as shown in Figure 3 In one specific embodiment of the present disclosure, the above-mentioned molecular detection unit can further include a film layer 301.

[0072] The film layer 301 is arranged at the connection between the sample flow channel 102 and the single-hole liquid storage cavity 103, and the film layer 301 is provided with a nanopore 302.

[0073] In addition, in the technical solution of the present disclosure, different film layers can be formed in the above-mentioned molecular detection unit according to the needs of actual application scenarios.

[0074] For example, as an example, in one specific embodiment of the present disclosure, the film layer 301 can be a bilayer or a monolayer.

[0075] For example, when the material of the film layer is phospholipid, the film layer 301 is a bilayer; and when the material of the film layer is other polymer material, the film layer 301 is a monolayer.

[0076] In addition, as an example, as shown in Figure 4 and Figure 3 In one specific embodiment of the present disclosure, the first structure layer 2 can include a first insulating layer 21 and a second insulating layer 22.

[0077] The first insulating layer 21 is arranged on the top of the substrate 1.

[0078] The second insulating layer 22 is arranged on the top of the first insulating layer 21.

[0079] The liquid resistance flow channel 104 is arranged in the first insulating layer 21.

[0080] The single-hole liquid storage cavity 103 is arranged in the second insulating layer 22.

[0081] In the technical solution of the present disclosure, the first insulating layer 21 and the second insulating layer 22 can be further arranged in the above-mentioned first structure layer 2, and then the liquid resistance flow channel 104 and the single-hole liquid storage cavity 103 are arranged in the first insulating layer 21 and the second insulating layer 22, respectively.

[0082] In addition, in the technical solution of the present disclosure, the specific positional relationship between the second structure layer 3 and the second buffer flow channel 105 can also be arranged according to the needs of actual application scenarios.

[0083] For example, as an example, as shown in Figure 4As shown, in one specific embodiment of the present disclosure, the second structural layer 3 can be disposed on top of the second insulating layer 22.

[0084] The second buffer flow channel 105 penetrates through the second structural layer 3, the second insulating layer 22 and the first insulating layer 21.

[0085] In this embodiment, the second structural layer 3 is disposed on top of the first structural layer, and the bottom of the second structural layer 3 abuts against the top of the second insulating layer 22 of the first structural layer.

[0086] For another example, as an example, as shown in FIG. 1, the second structural layer 3 can be disposed on top of the first insulating layer 21 and the second insulating layer 22, and the bottom of the second structural layer 3 abuts against the top of the first insulating layer 21 through the second insulating layer 22. Figure 5 As shown, in one specific embodiment of the present disclosure, the second structural layer 3 can be disposed on top of the second insulating layer 22.

[0087] The second buffer flow channel 105 penetrates through the second structural layer 3 and the first insulating layer 21.

[0088] In this embodiment, the bottom of the second structural layer 3 is embedded into the first structural layer, and abuts against the top of the first insulating layer 21 through the second insulating layer 22.

[0089] For another example, as an example, as shown in FIG. 1, the second structural layer 3 can be disposed on top of the first insulating layer 21 and the second insulating layer 22, and the bottom of the second structural layer 3 abuts against the top of the first insulating layer 21 through the second insulating layer 22. Figure 5 As shown, in one specific embodiment of the present disclosure, the above-mentioned molecular detection unit can further comprise a driving electrode 6.

[0090] The driving electrode 6 is disposed in the third structural layer 5 and connected with the first buffer flow channel 101.

[0091] Before using the above-mentioned molecular detection unit, considering the requirements of storage, transportation and the like, buffer solution (a polar solution containing required electrolyte) can be injected and filled into the sample flow channel 102, the first buffer flow channel 101 and the second buffer flow channel 105, respectively. Since the single-hole liquid storage cavity 103 is in communication with the second buffer flow channel 105 through the liquid resistance flow channel 104, the buffer solution will also be filled into the single-hole liquid storage cavity 103 and the liquid resistance flow channel 104.

[0092] When the above-mentioned molecular detection unit needs to be used, sample solution can be injected into the sample flow channel 102 to replace the original buffer solution in the sample flow channel 102.

[0093] In this process, the liquid flow can be driven by positive or negative pressure provided by an external pump. For example, when the sample solution is driven into the sample flow channel 102 by positive pressure, the sample solution in the sample solution storage device (e.g., a sample solution bottle, etc.) will flow into the sample flow channel 102 from one end of the sample flow channel 102, flow through the sample flow channel 102, and exit from the other end of the sample flow channel 102 under the pressure provided by the external pump, and be collected by the waste liquid collection device (e.g., a waste liquid bottle, etc.) to prevent contamination.

[0094] Under pressure driving, the Reynolds number of the sample solution when moving in the sample flow channel 102 is low, showing a laminar flow state, and the sample solution will not pass through the membrane layer 301 into the single-hole solution storage cavity 103. When there is no pressure driving, the membrane layer 301 as a thin film between the single-hole solution storage cavity 103 and the sample flow channel 102 can also prevent mutual diffusion between the sample solution in the sample flow channel 102 and the buffer solution in the single-hole solution storage cavity 103.

[0095] When the above-mentioned molecular detection unit is in a working state, a voltage can be applied to one side of the sample flow channel 102 (e.g., a voltage is applied to the sample solution in the sample flow channel 102, which can be denoted as V3, as shown in FIG. 6A), and a voltage is applied to one side of the first buffer solution flow channel 101 (e.g., a voltage can be applied to the buffer solution in the first buffer solution flow channel 101 by the driving electrode 6, which can be denoted as V2, as shown in FIG. 6A). Figure 5 Figure 6 Since the first buffer solution flow channel 101 is in communication with the single-hole solution storage cavity 103 through the second buffer solution flow channel 105 and the liquid resistance flow channel 104, the voltage V1 applied to one side of the single-hole solution storage cavity 103 will eventually have a certain difference from the voltage V2. Since different voltages can be applied in different liquids, the implementation of the voltage sequencing principle can be ensured. Therefore, only when the preset voltages V2 and V3 are applied to the buffer solution and the sample solution, the electrolyte and the target sample molecule will pass through the nanopore 302 and the liquid resistance flow channel 104 under the action of the electric field force.

[0096] When the DNA molecule 303 in the sample solution passes through the nanopore (e.g., nanopore protein) 302 embedded in the membrane layer 301, the resistance R1 of the nanopore 302 can be regarded as a variable resistance when the DNA molecule 303 passes through, and the resistance of the liquid resistance flow channel 104 can be regarded as a fixed resistance. As the base sequence of the DNA molecule 303 passes through the nanopore 302 in turn, the resistance R1 will change correspondingly, thereby causing the voltage V1 between the two resistances to change. Therefore, according to the change rule of the voltage V1 detected by the sensing electrode 4, the base sequence of the DNA molecule 303 can be calculated, and the sequencing of the DNA molecule 303 can be realized.

[0097] ​For example, as an example, in another embodiment of the present disclosure, the resistance between the above-mentioned voltages V2 and V3 is mainly composed of the resistance R1 of the nano-pore-embedded membrane layer 301 and the resistance R2 of the liquid resistance flow channel 2, and the voltage VI will be collected and amplified by an amplification circuit 43, and transmitted to the electrical signal database of the sequencing system 44 (for example, a computer or the like), as shown in Figure 7 During the sequencing process, the DNA molecule passes through the nano-pore, and the combination of DNA bases passing through the nano-pore at different times is different, resulting in that the overall resistance R1 of the nano-pore-embedded membrane layer 301 changes over time, that is, the resistance R1 can be regarded as a variable resistance. Therefore, by the voltage VI at a given time, the total resistance of the nano-pore-embedded membrane layer 301 and the current base combination passing through the nano-pore can be determined, and by using a deep learning algorithm, the base sequence of the DNA molecule passing through the pore protein can be calculated.

[0098] In the above-mentioned molecular detection unit of the present disclosure, the first buffer flow channel 101 is arranged in the third structure layer 5, the sample flow channel 102 is arranged in the second structure layer 3, and the single-hole liquid storage cavity 103 is arranged in the first structure layer 2, forming a stacked multi-layer structure. Since the first buffer flow channel 101, the sample flow channel 102 and the single-hole liquid storage cavity 103 are arranged in different layers, and the first buffer flow channel 101 is located in the third structure layer 5 above the sample flow channel 102 and the single-hole liquid storage cavity 103, the arrangement density of the nano-pore device per unit area can be effectively improved (for example, the channel density of the nano-pore chip can be improved to the level of one hundred thousand devices per square centimeter), and the nano-pore sequencing device flux size is greatly improved, so that ultra-high flux nano-pore-based gene sequencing can be realized. In addition, since the above-mentioned molecular detection unit is a stacked multi-layer structure, each layer structure can be separately manufactured by using mature semiconductor processing technology, and then the multi-layer structure is formed by bonding, so that the production and manufacturing efficiency can be effectively improved.

[0099] In addition, the single-hole liquid storage cavity 103, the liquid resistance flow channel 104, the first buffer flow channel 101, the second buffer flow channel 105 and the sample flow channel 102 in the above-mentioned molecular detection unit are all arranged on the same side of the substrate 1, so that the liquid resistance flow channel and the buffer flow channel can be constructed on the same surface of the substrate, thereby further effectively improving the chip manufacturing efficiency based on the voltage sequencing method, and further improving the flux of the nano-pore sequencing device.

[0100] In addition, in the above-mentioned molecular detection unit of the present disclosure, the sample flow channel 102 and the first buffer flow channel 101 are isolated from each other, and the sample solution can flow in from one end of the sample flow channel 102 and flow out from the other end of the sample flow channel 102, without entering the first buffer flow channel 101; similarly, the buffer can flow in from one end of the first buffer flow channel 101 and flow out from the other end of the first buffer flow channel 101, and the buffers in the first buffer flow channel 101 and the second buffer flow channel 105 will not enter the sample flow channel 102.

[0101] Since the above-mentioned two flow channels are separated, different voltages can be applied to different liquids in the two flow channels, thereby ensuring the implementation of the basic principle of voltage sequencing. In addition, the above-mentioned two separated flow channels can greatly reduce cross contamination and electrical leakage between samples, and improve the signal-to-noise ratio of the signal.

[0102] In addition, as an example, as shown in Figure 8 and Figure 7 in one specific embodiment of the present disclosure, the above-mentioned molecular detection unit can further include two sample solution outlets 1021 and two buffer outlets 1011;

[0103] The two sample solution outlets 1021 are respectively in communication with the sample flow channel 102;

[0104] The two buffer outlets 1011 are respectively in communication with the first buffer flow channel 101.

[0105] For example, as shown in Figure 8 and Figure 9 the above-mentioned sample solution outlet 1021 can pass through the third structural layer 5 and be in communication with the sample flow channel 102 in the second structural layer 3; and the buffer outlet 1011 can be arranged in the third structural layer 5 and be in communication with the first buffer flow channel 101 in the third structural layer 5.

[0106] In the above-mentioned molecular detection unit, the sample flow channel 102 and the two sample solution outlets 1021 can constitute a micro-flow channel system of the sample, while the first buffer flow channel 101, the second buffer flow channel 105 and the two buffer outlets 1011 can constitute a micro-flow channel system of the buffer. The liquids of the two micro-flow channel systems enter from one outlet of the system and flow out from the other outlet, and are isolated from each other. For example, the sample solution only enters the sample flow channel 102 from one sample solution outlet 1021 and then flows out of the sample flow channel 102 from the other sample solution outlet 1021; in this process, the sample solution does not enter the micro-flow channel system of the buffer. Similarly, the buffer can enter the first buffer flow channel 101 and the second buffer flow channel 105 from one buffer outlet 1011 and then flow out of the first buffer flow channel 101 from the other buffer outlet 1011; in this process, the buffer in the micro-flow channel system of the buffer does not enter the micro-flow channel system of the sample.

[0107] Since the above-mentioned two micro-flow channel systems are separated, different voltages can be applied to different liquids in the two micro-flow channel systems, thereby ensuring the implementation of the basic principle of voltage sequencing. In addition, the above-mentioned two separated micro-flow channel systems can greatly reduce cross contamination and electrical leakage between samples and improve the signal-to-noise ratio of the signal.

[0108] In addition, as an example, as shown in Figure 1 In one specific embodiment of the present disclosure, the above-mentioned molecular detection unit can further include a sample flow channel interface 12 and a buffer flow channel interface 11.

[0109] The sample flow channel interface 12 is provided in the sample solution outlet 1021, and the top of the sample flow channel interface 12 protrudes from the top of the sample solution outlet 1021; the bottom of the sample flow channel interface 12 communicates with the sample flow channel 102.

[0110] The buffer flow channel interface 11 is provided in the buffer outlet 1011, and the top of the buffer flow channel interface 11 protrudes from the top of the buffer outlet 1011; the bottom of the buffer flow channel interface 11 communicates with the first buffer flow channel 101.

[0111] In the technical solution of the present disclosure, a sample flow channel interface 12 can be provided in each sample solution outlet 1021, and a buffer flow channel interface 11 can be provided in each buffer outlet 1011, so that the sample flow channel interface 12 communicates with the sample solution outlet 1021, and the buffer flow channel interface 11 communicates with the buffer outlet 1011.

[0112] In addition, as an example, in one specific embodiment of the present disclosure, a first driving electrode can also be arranged in the sample flow channel interface, and a second driving electrode can also be arranged in the buffer flow channel interface.

[0113] At this time, the sample flow channel interface and the buffer flow channel interface described above can be a structure allowing liquid to enter and exit and a driving electrode to access, so that the corresponding liquid can be injected into the sample solution outlet 1021 or caused to flow out of the sample solution outlet 1021 through the sample flow channel interface, and the corresponding voltage V3 can be applied to the liquid in the sample flow channel through the first driving electrode in the sample flow channel interface. Similarly, the corresponding liquid can be injected into the buffer outlet 1011 or caused to flow out of the buffer outlet 1011 through the buffer flow channel interface, and the corresponding voltage V2 can be applied to the liquid in the first buffer flow channel through the second driving electrode in the buffer flow channel interface.

[0114] In addition, in the technical solution of the present disclosure, the shape of the liquid resistance flow channel 104 can be set according to the needs of the actual application scene.

[0115] For example, in one specific embodiment of the present disclosure, the shape of the liquid resistance flow channel 104 can be a straight line type (as shown in Figure 10 , a curved type, an arc type, or a surrounding type.

[0116] For another example, in one specific embodiment of the present disclosure, the liquid resistance flow channel 104 can be a shape in which a part is in a curved type or an arc type, and another part is in a straight line type.

[0117] In addition, in the technical solution of the present disclosure, a molecular detection chip is also proposed.

[0118] For example, as an example, in another specific embodiment of the present disclosure, the molecular detection chip comprises a molecular detection array;

[0119] The molecular detection array comprises at least one molecular detection unit group; each molecular detection unit group comprises a plurality of molecular detection units; the molecular detection unit can be the molecular detection unit in any one of the above-mentioned embodiments;

[0120] Among them, the sample flow channels of the respective molecular detection units are connected to each other to form a common sample flow channel; and / or,

[0121] The first buffer flow channels of the respective molecular detection units are connected to each other to form a first common buffer flow channel and / or,

[0122] The second buffer flow channels of the respective molecular detection units are connected to each other to form a second common buffer flow channel.

[0123] In the molecular detection chip, a molecular detection array can be included, and one or more groups of molecular detection units can be arranged in the molecular detection array, and each group of molecular detection units can include a plurality of molecular detection units.

[0124] Since the sample flow channels in each molecular detection unit are arranged in the same structural layer (i.e., the second structural layer), in each group of molecular detection units, the sample flow channels of the molecular detection units can be connected to each other according to their respective arrangement orders to form a common sample flow channel in the second structural layer; or the sample flow channels of all the molecular detection units in the molecular detection array can be connected to each other to form a common sample flow channel in the second structural layer. Similarly, since the first buffer flow channels in each molecular detection unit are arranged in the same structural layer (i.e., the third structural layer), the first buffer flow channels of the molecular detection units in each group of molecular detection units can be connected to each other according to their respective arrangement orders to form a first common buffer flow channel in the third structural layer; or the first buffer flow channels of all the molecular detection units in the molecular detection array can be connected to each other to form a first common buffer flow channel in the third structural layer; similarly, the second buffer flow channels of the molecular detection units in the molecular detection array can be connected to each other according to their respective arrangement orders to form a second common buffer flow channel in the second structural layer; or the second buffer flow channels of all the molecular detection units in the molecular detection array can be connected to each other to form a second common buffer flow channel in the second structural layer. Therefore, each molecular detection unit in each group of molecular detection units or the entire molecular detection array can share the common sample flow channel, and / or share the first common buffer flow channel, and / or share the second common buffer flow channel. Therefore, the required liquid (e.g., buffer and / or sample solution) can be injected into each molecular detection unit through the common sample flow channel, and the required liquid (e.g., buffer) can be injected into each molecular detection unit through the first common buffer flow channel.

[0125] In addition, in the technical solution of the present disclosure, the number of molecular detection units in a group of molecular detection units can be set according to the needs of actual application scenarios.

[0126] For example, as an example, in another specific embodiment of the present disclosure, a group of molecular detection units can include 2, 4, 6, 8, or more molecular detection units. In the technical solution of the present disclosure, the number of molecular detection units in a group of molecular detection units can be set according to the length of the second buffer flow channel and / or the second common buffer flow channel and the size of the group of molecular detection units, and therefore will not be listed one by one here.

[0127] For example, as shown in FIG. 1, the molecular detection array 100 includes a group of molecular detection units 110, and the group of molecular detection units 110 includes a plurality of molecular detection units 111, 112, 113, 114, 115, and 116. The molecular detection array 100 further includes a common sample flow channel 120 and a first common buffer flow channel 130. Figure 12As shown, in another specific embodiment of the present disclosure, one group of molecular detection units can include 4 molecular detection units.

[0128] Of course, the number of molecular detection units in one group of molecular detection units can also be other suitable values, which are not listed one by one here.

[0129] In addition, as an example, in one specific embodiment of the present disclosure, the molecular detection chip can further include: two common sample solution outlets 51 and / or two common buffer outlets 52; the two common sample solution outlets 51 are respectively in communication with the common sample flow channels; the two common buffer outlets 52 are respectively in communication with the first common buffer flow channels.

[0130] That is, when each molecular detection unit in the molecular detection chip shares the common sample flow channel and shares the first common buffer flow channel, only two common sample solution outlets and / or two common buffer outlets can be provided in the entire molecular detection chip, the two common sample solution outlets are respectively in communication with the common sample flow channels shared, and the two common buffer outlets are respectively in communication with the first common buffer flow channels shared, as shown in Figure 13 and Figure 14 Therefore, all molecular detection units in the molecular detection array share the two common sample solution outlets and / or the two common buffer outlets. When the above-mentioned molecular detection array needs to be used, the buffer or sample solution can be injected into the sample flow channel of each molecular detection unit from one common sample solution outlet and flows out of the sample flow channel from the other common sample solution outlet; the buffer can be injected into the first buffer flow channel and the second buffer flow channel of each molecular detection unit from one common buffer outlet and flows out of the first buffer flow channel from the other common buffer outlet.

[0131] In addition, as an example, as shown in Figure 15 in one specific embodiment of the present disclosure, the molecular detection chip can further include: a common sample flow channel interface 501 and / or a common buffer flow channel interface 502;

[0132] The common sample flow channel interface 501 is provided in the common sample solution outlet 51, the top of the common sample flow channel interface 501 protrudes from the top of the common sample solution outlet 51; the bottom of the common sample flow channel interface 501 is in communication with the common sample flow channel;

[0133] The common sample flow channel interface 501 is arranged in the common sample solution outlet 51, and the top of the common sample flow channel interface 501 protrudes from the top of the common sample solution outlet 51; the bottom of the common sample flow channel interface 501 is in communication with the first sample flow channel.

[0134] In the technical solution of the present disclosure, a common sample flow channel interface 501 can be arranged in each common sample solution outlet 51, and a common buffer flow channel interface 502 can be arranged in each common buffer outlet 52, so that the common sample flow channel interface 501 is in communication with the common sample solution outlet 51, and the common buffer flow channel interface 502 is in communication with the common buffer outlet 52.

[0135] In addition, as an example, in one specific embodiment of the present disclosure, a first common driving electrode can also be arranged in the common sample flow channel interface 501, and a second common driving electrode can also be arranged in the common buffer flow channel interface 502.

[0136] At this time, the common sample flow channel interface 501 and the common buffer flow channel interface 502 described above can be a structure that allows liquid to enter and exit and a driving electrode to access, so that the corresponding liquid can be injected into the common sample solution outlet 51 or caused to flow out of the common sample solution outlet 51 through the common sample flow channel interface 501, and the first common driving electrode in the common sample flow channel interface 501 can apply a corresponding voltage V3 to the liquid in the common sample flow channel and the sample flow channel. Similarly, the corresponding liquid can be injected into the common buffer outlet 52 or caused to flow out of the common buffer outlet 52 through the common buffer flow channel interface 502 described above, and the second common driving electrode in the common buffer flow channel interface 502 can apply a corresponding voltage V2 to the liquid in the first common buffer flow channel and the first buffer flow channel.

[0137] In addition, in the technical solution of the present disclosure, the second buffer flow channels in each molecular detection unit in the molecular detection array can be independent of each other or directly in communication with each other.

[0138] For example, as an example, in another specific embodiment of the present disclosure, the first buffer flow channels in each molecular detection unit in the molecular detection array are in communication with each other to form a first common buffer flow channel, and the second buffer flow channels in each molecular detection unit can be in communication with the first common buffer flow channel, respectively.

[0139] For example, in another embodiment of the present disclosure, the first buffer flow channels in each of the molecular detection units in the molecular detection array can be connected to each other in the third structure layer to form a first common buffer flow channel, and the second buffer flow channels in each of the molecular detection units can be independently connected to the first common buffer flow channel. At this time, the second buffer flow channel in each of the molecular detection units is equivalent to a hollow needle tube or pipeline extending downward from the first common buffer flow channel, penetrating through the second structure layer and inserted into the liquid resistance flow channel of each of the molecular detection units, so as to connect the liquid resistance flow channel of each of the molecular detection units to the first common buffer flow channel.

[0140] In addition, in the technical solution of the present disclosure, the plurality of molecular detection units in a molecular detection unit group can have a plurality of different arrangement modes.

[0141] For example, as an example, in another embodiment of the present disclosure, the plurality of molecular detection units in the same molecular detection unit group can be uniformly arranged around a common center.

[0142] For example, as an example, in another embodiment of the present disclosure, the plurality of molecular detection units in the same molecular detection unit group can be uniformly arranged around a common center.

[0143] In addition, as an example, in another embodiment of the present disclosure, when the plurality of molecular detection units in the same molecular detection unit group are uniformly arranged around a common center (for example, 6 molecular detection units are uniformly arranged in a hexagon, as shown in Figure 18 and Figure 15 The plurality of molecular detection units in the same molecular detection unit group can share a first buffer flow channel and share a second buffer flow channel.

[0144] The arrangement mode can also be regarded as: the first buffer flow channels of each of the molecular detection units in the same molecular detection unit group are connected to each other to form a first common buffer flow channel, and the second buffer flow channels of each of the molecular detection units in the same molecular detection unit group are connected to each other to form a second common buffer flow channel.

[0145] Figure 18 and Figure 15 The arrangement mode shown in the above can form a molecular detection unit group by a hexagonal close-packed mode, and a vertical second buffer flow channel or a second common buffer flow channel can be arranged at the center position of the molecular detection unit group, so as to save the total volume occupied by the second buffer flow channel or the second common buffer flow channel, and further improve the arrangement density of the nanopore device per unit area.

[0146] In addition, in the technical solution of the present disclosure, since the liquid resistance flow channel 104 is arranged in each molecular detection unit, in the array design of the molecular detection chip, dummy holes 61 can be introduced to realize the consistency of the array. Of course, the dummy holes can also not be introduced.

[0147] In addition, in the technical solution of the present disclosure, the sample flow channels in each molecular detection unit in the molecular detection array can be independent of each other and connected to a common sample flow channel through corresponding channels; or the sample flow channels in each molecular detection unit can be directly connected to each other and directly serve as part of the common sample flow channel.

[0148] For example, as shown in FIGS. 1 and 2, in one embodiment of the present disclosure, the shape of the sample flow channel in each molecular detection unit can be a rectangular shape, and the sample flow channels in each molecular detection unit are independent of each other and connected to a common sample flow channel through corresponding channels arranged in the second structure layer. Figure 18 Figure 15 As shown in FIGS. 3 and 4, in another embodiment of the present disclosure, the shape of the sample flow channel in each molecular detection unit can be a cylindrical shape, and the sample flow channels in each molecular detection unit are independent of each other and connected to a common sample flow channel through corresponding channels arranged in the second structure layer.

[0149] In addition, in the technical solution of the present disclosure, the first buffer flow channels in each molecular detection unit in the molecular detection array can be independent of each other and connected to a first common buffer flow channel through corresponding channels; or the first buffer flow channels in each molecular detection unit can be directly connected to each other and directly serve as part of the first common buffer flow channel.

[0150] For example, as shown in FIGS. 5 and 6, in one embodiment of the present disclosure, the shape of the first buffer flow channel in each molecular detection unit can be a rectangular shape, and the first buffer flow channels in each molecular detection unit are independent of each other and connected to a first common buffer flow channel through corresponding channels arranged in the second structure layer. Figure 16 Figure 19 As shown in FIGS. 7 and 8, in another embodiment of the present disclosure, the shape of the first buffer flow channel and the second buffer flow channel in each molecular detection unit can be a cylindrical shape, and the first buffer flow channels in each molecular detection unit in the same group of molecular detection units are connected to a first common buffer flow channel, and the second buffer flow channels in each molecular detection unit in the same group of molecular detection units are connected to a second common buffer flow channel.

[0151] ​​For another example, in another embodiment of the present disclosure, the shape of the first buffer solution flow channel in each molecular detection unit can also be cylindrical, the first buffer solution flow channels in each molecular detection unit are independent of each other, and the first buffer solution flow channels are connected to form a first common buffer solution flow channel through corresponding channels (not shown in the figure) provided in the third structure layer. The shape of the first buffer solution flow channel is also a non-parallel flow channel design instead of a series of parallel narrow flow channels, which can simplify the introduction of the buffer solution solution of the first buffer solution flow channel.

[0152] In addition, for example, in another embodiment of the present disclosure, the plurality of molecular detection units in the same group of molecular detection units can be sequentially formed in a linear array structure.

[0153] In the molecular detection chip, a molecular detection array can be provided, and one or more groups of molecular detection units can be provided in the molecular detection array to form one or more groups of molecular detection units.

[0154] The sample flow channels of each molecular detection unit in each row of molecular detection units (i.e., one group of molecular detection units) can be connected to each other according to the respective arrangement order to form a common sample flow channel in the second structure layer; and the first buffer solution flow channels of each molecular detection unit can also be connected to each other according to the respective arrangement order to form a first common buffer solution flow channel in the third structure layer. Therefore, each molecular detection unit in each row of molecular detection units can share the common sample flow channel and the first common buffer solution flow channel.

[0155] In addition, in the technical solution of the present disclosure, one row of molecular detection units (i.e., one group of molecular detection units) can be provided in the molecular detection array according to the needs of the actual application scene, or multiple rows of molecular detection units (i.e., multiple groups of molecular detection units) can be provided in the molecular detection array. In addition, the number of common sample flow channels and / or first common buffer solution flow channels in the molecular detection array can also be pre-set according to the needs of the actual application scene.

[0156] For example, for example, in one embodiment of the present disclosure, when the molecular detection array includes one or more groups of molecular detection units, the number of common sample flow channels and / or first common buffer solution flow channels in the molecular detection array can be equal to the number of groups of molecular detection units in the molecular detection array.

[0157] For another example, for example, in another embodiment of the present disclosure, when the molecular detection array includes multiple groups of molecular detection units, the number of common sample flow channels and / or first common buffer solution flow channels in the molecular detection array can be less than the number of groups of molecular detection units in the molecular detection array.

[0158] For example, as an example, in another specific embodiment of this disclosure, when the molecular detection array includes one or more groups of molecular detection units, the number of first common buffer channels in the molecular detection array may be equal to the number of groups of molecular detection units in the molecular detection array.

[0159] For example, as an example, in another specific embodiment of this disclosure, when the molecular detection array includes multiple molecular detection unit groups, the number of first common buffer channels in the molecular detection array can be less than the number of molecular detection unit groups in the molecular detection array.

[0160] In the technical solutions disclosed herein, the above specific embodiments can be combined in accordance with the needs of actual application scenarios to obtain various different specific implementation methods, which will not be listed one by one here.

[0161] In addition, in the technical solution disclosed herein, the number of molecular detection units in each molecular detection unit group in the molecular detection array can be flexibly set according to the needs of the actual application scenario.

[0162] For example, as an example, in a specific embodiment of this disclosure, when the molecular detection array includes multiple molecular detection unit groups, the number of molecular detection units in each molecular detection unit group may be equal or unequal.

[0163] Furthermore, in the technical solution disclosed herein, the aforementioned molecular detection chip can be prepared through the following steps:

[0164] Step A1: Prepare a substrate and form sensing electrodes for each molecular detection unit in the molecular detection chip on the top of the substrate.

[0165] For example, as an example, such as Figure 20 As shown, in a specific embodiment of this disclosure, a substrate layer (i.e., substrate 1) can be fabricated first using semiconductor processes. This substrate layer can be used to contain the circuit structure required by the device. Furthermore, during the fabrication of substrate 1, sensing electrodes 4 for each molecular detection unit in the molecular detection chip can be formed on the top of substrate 1 using semiconductor processes. These sensing electrodes 4 can be connected to different types of circuit boards or circuit chips through different semiconductor packaging methods (e.g., they can be connected to different types of circuit boards or circuit chips through metal leads disposed in substrate 1), which will not be elaborated further here.

[0166] Step A2: A first structural layer is formed on the substrate, and a corresponding single-well liquid reservoir and a bonding hole for the second buffer flow channel are formed in the first structural layer for each molecular detection unit in the molecular detection chip.

[0167] In this step, first, a first structure layer 2 is formed on the substrate 1, and then a corresponding single-hole liquid storage cavity 103 and a bonding hole for the second buffer solution flow channel 105 are formed in the first structure layer 2 for each molecular detection unit in the molecular detection chip.

[0168] In addition, in the technical solution of the present disclosure, the above-mentioned step A2 can be implemented by using various implementation manners. In the following, several specific implementation manners will be taken as examples to introduce the technical solution of the present disclosure.

[0169] For example, as an example, in one specific embodiment of the present disclosure, when the first structure layer includes a first insulating layer and a second insulating layer, the above-mentioned step A2 can include the following steps:

[0170] Step A201, for each molecular detection unit in the molecular detection chip, a liquid resistance flow channel sacrificial structure is formed at a preset position on the substrate and the sensing electrode.

[0171] For example, as an example, as shown in Figure 21 in one specific embodiment of the present disclosure, for each molecular detection unit in the molecular detection chip, a liquid resistance flow channel sacrificial structure 41 can be formed at a preset position on the substrate 1 and the sensing electrode 4 (i.e. the position where the liquid resistance flow channel 104 of each molecular detection unit needs to be formed) by a certain pattern transfer method (such as electron beam exposure or laser direct writing, etc.) and / or a material deposition process (such as physical vapor deposition or chemical vapor deposition, etc.).

[0172] For example, as an example, in one specific embodiment of the present disclosure, the liquid resistance flow channel sacrificial structure 41 can be a sacrificial material such as amorphous silicon.

[0173] Step A202, forming a first insulating layer 21 on the substrate, the sensing electrode and the liquid resistance flow channel sacrificial structure.

[0174] For example, as an example, as shown in Figure 22 in one specific embodiment of the present disclosure, when the first structure layer 2 includes a first insulating layer 21 and a second insulating layer 22, a first insulating layer 21 covering the substrate 1, the sensing electrode 4 and the liquid resistance flow channel sacrificial structure 41 can be formed by a certain material deposition process (such as physical vapor deposition or chemical vapor deposition, etc.).

[0175] Step A203, processing the first insulating layer to expose part of the sensing electrode and the liquid resistance flow channel sacrificial structure at the preset position.

[0176] For example, as an example, as shown in Figure 23As shown, in one specific embodiment of the present disclosure, the first insulating layer can be processed by a certain pattern transfer method (e.g., photolithography, etc.) and / or a certain etching process (e.g., reactive ion etching, etc.) to expose part of the sensing electrode 4 and the liquid resistance flow channel sacrificial structure 41 at the preset position.

[0177] Step A204, forming a second insulating layer on the first insulating layer, and forming a corresponding single-hole liquid storage cavity and a bonding hole for the second buffer flow channel in the second insulating layer for each molecular detection unit in the molecular detection chip.

[0178] For example, as an example, as Figure 24 As shown, in one specific embodiment of the present disclosure, the second insulating layer 22 can be formed on the first insulating layer 21 by a certain material deposition process (e.g., physical vapor deposition or chemical vapor deposition, etc.); then, a certain pattern transfer method (e.g., photolithography, etc.) and / or a certain etching process (e.g., reactive ion etching, etc.) are used to form a corresponding single-hole liquid storage cavity 103 and a bonding hole for the second buffer flow channel 105 in the second insulating layer 22 for each molecular detection unit in the molecular detection chip. Wherein, the single-hole liquid storage cavity 103 and the bonding hole for the second buffer flow channel 105 are isolated from each other and do not directly communicate with each other. For example, the single-hole liquid storage cavity 103 corresponds to the position of the sensing electrode 4, and the bonding hole for the second buffer flow channel 105 corresponds to the middle position of the liquid resistance flow channel sacrificial structure 41.

[0179] Therefore, by the above steps A201-A204, the above-mentioned insulating layer 2 and the first structure layer 2 can be formed, and the single-hole liquid storage cavity 103 and the bonding hole for the second buffer flow channel 105 of each molecular detection unit in the molecular detection chip can be formed in the first structure layer 2.

[0180] Step A3, forming a corresponding cavity of the sample flow channel and the second buffer flow channel for each molecular detection unit in the molecular detection chip on the base material for the second structure layer, forming the second structure layer.

[0181] For example, as an example, as Figure 25 and Figure 26As shown, in one specific embodiment of the present disclosure, the cavity of the sample flow channel 102 and the corresponding through hole as the second buffer flow channel 105 for each molecular detection unit in the molecular detection chip can be formed on the base material 30 (e.g., glass, silicon dioxide, quartz, or other materials) for manufacturing the second structure layer by some pattern transfer method (e.g., photolithography, etc.) and / or some material etching process (e.g., reactive ion etching, etc.), thereby forming the second structure layer 3. Wherein, the sample flow channel 102 and the second buffer flow channel 105 are isolated from each other and not directly connected to each other.

[0182] Step A4, bonding the second structure layer and the first structure layer to form a bonded structure.

[0183] For example, as an example, as Figure 27 As shown, in one specific embodiment of the present disclosure, when the first structure layer 2 includes the first insulating layer 21 and the second insulating layer 22, and the bottom of the second structure layer 3 is embedded into the second insulating layer 22, the second structure layer 3, the first insulating layer 21 and the second insulating layer 22 can be bonded, the second buffer flow channel of the second structure layer is inserted into the bonding hole in the first structure layer, to form a bonded structure, and the construction of the sample flow channel 102, the single-hole liquid storage cavity 103 and the second buffer flow channel 105 is realized. Wherein, the bottom of the second structure layer 3 abuts against the top of the first insulating layer 21 through the second insulating layer 22, the bottom of the second buffer flow channel 105 can be in communication with the top of the liquid resistance flow channel sacrificial structure 41, and the sample flow channel 102 is located above the single-hole liquid storage cavity 103 and in communication with the single-hole liquid storage cavity 103.

[0184] For another example, as an example, in another specific embodiment of the present disclosure, when the bottom of the second structure layer abuts against the top of the first structure layer (or the second insulating layer) and is not embedded into the first structure layer (or the second insulating layer), the second structure layer and the first structure layer can also be bonded, the second structure layer is directly bonded on the top of the first structure layer (or the second insulating layer) to form a bonded structure, and the construction of the sample flow channel 102, the single-hole liquid storage cavity 103 and the second buffer flow channel 105 can also be realized, so that the bottom of the second buffer flow channel 105 can be in communication with the top of the liquid resistance flow channel sacrificial structure 41, and the sample flow channel 102 is located above the single-hole liquid storage cavity 103 and in communication with the single-hole liquid storage cavity 103.

[0185] Step A5, forming a corresponding liquid resistance flow channel for each molecular detection unit in the molecular detection chip in the first structure layer.

[0186] In this step, a corresponding liquid resistance flow channel 104 for each molecular detection unit in the molecular detection chip can be formed in the first structure layer 2.

[0187] For example, as shown in FIG. 1A, in one embodiment of the present disclosure, the liquid resistance flow channel 104 connecting the single-hole liquid storage cavity 103 and the second buffer solution flow channel 105 can be formed by removing the liquid resistance flow channel sacrificial structure 41 in each molecular detection unit using a certain etching process (e.g., a xenon fluoride silicon etching process, etc.). Figure 28

[0188] Step A6, forming a corresponding first buffer solution flow channel for each molecular detection unit in the molecular detection chip on the base material for the third structure layer, and forming the third structure layer.

[0189] For example, as shown in FIG. 1A, in one embodiment of the present disclosure, the liquid resistance flow channel 104 connecting the single-hole liquid storage cavity 103 and the second buffer solution flow channel 105 can be formed by removing the liquid resistance flow channel sacrificial structure 41 in each molecular detection unit using a certain etching process (e.g., a xenon fluoride silicon etching process, etc.). Figure 29

[0190] In addition, in one embodiment of the present disclosure, the forming the third structure layer can further include: forming a corresponding driving electrode 6 for each molecular detection unit in the molecular detection chip on the base material for the third structure layer (i.e., in the third structure layer 5).

[0191] Step A7, bonding the third structure layer and the bonding structure.

[0192] For example, as shown in FIG. 1A, in one embodiment of the present disclosure, the third structure layer 5 and the bonding structure including the second structure layer 3 and the first structure layer 2 can be bonded so that the second buffer solution flow channel 105 communicates with the first buffer solution flow channel 101, thereby forming the molecular detection chip. ​ Therefore, by the above steps A1-A7, the required molecular detection chip can be prepared.

[0193] In addition, in the technical solution of the present disclosure, a film layer 301 can be further formed at the connection between the single-hole liquid storage cavity 103 and the sample flow channel 102 of each molecular detection unit, and a nanopore 302 can be further formed on the film layer 301.

[0194] For example, as shown in FIG. 1A, in one embodiment of the present disclosure, the liquid resistance flow channel 104 connecting the single-hole liquid storage cavity 103 and the second buffer solution flow channel 105 can be formed by removing the liquid resistance flow channel sacrificial structure 41 in each molecular detection unit using a certain etching process (e.g., a xenon fluoride silicon etching process, etc.).

[0195]

[0196] Step A8, forming a film layer at the connection between the single-hole liquid storage cavity and the sample flow channel of each molecular detection unit, and forming a nanopore on the film layer.​​​

[0197] In the technical solution of the present disclosure, the membrane layer 301 can be formed at the connection between the single-well liquid storage cavity 103 and the sample flow channel 102 of each molecular detection unit according to their positions.

[0198] In addition, the molecular detection unit and the molecular detection chip described above can be widely applied to the preparation of a nanopore sensor or used in nanopore characterization of analytes, and have great application potential in the field of biological detection technology.

[0199] In summary, in the technical solution of the present disclosure, the first buffer flow channel, the sample flow channel and the single-well liquid storage cavity are arranged in different layers, and the first buffer flow channel is located in the third structure layer above the sample flow channel and the single-well liquid storage cavity. Therefore, the arrangement density of the nanopore device per unit area can be effectively improved, and the nanopore sequencing device throughput can be greatly improved, so that ultrahigh-throughput nanopore-based gene sequencing can be realized. In addition, the molecular detection unit described above is a stacked multilayer structure, so each layer structure can be separately manufactured using mature semiconductor processing technology, and then the multilayer structure can be formed by bonding, thereby effectively improving the production efficiency

[0200] In addition, the two sets of independent microchannel systems are constructed by using multilayer flow channels, and interconnection can be achieved by removing the liquid resistance flow channel sacrificial structure between the two sets of systems before the final application, so that the liquid resistance flow channel and the buffer flow channel can be manufactured on the same side of the substrate. The single-well liquid storage cavity, the liquid resistance flow channel, the buffer flow channel and the sample flow channel are located on the same side of the substrate, so that the liquid resistance flow channel and the buffer flow channel can be constructed on the same surface of the substrate, thereby effectively improving the chip manufacturing efficiency based on the voltage sequencing method, and further improving the throughput of the nanopore sequencing device.

[0201] In addition, in the technical solution of the present disclosure, the sample flow channel and the buffer flow channel are isolated from each other, so that different voltages can be applied in different liquids in the two flow channels, thereby ensuring the realization of the basic principle of voltage sequencing. In addition, the two separated flow channels can greatly reduce cross contamination and electrical leakage between samples, and improve the signal-to-noise ratio.

[0202] The above only describes the preferred embodiments of the present disclosure and is not intended to limit the present disclosure. Any modification, equivalent replacement, improvement, etc. made within the spirit and principle of the present disclosure shall be included in the protection scope of the present disclosure.

Claims

1. A molecular detection unit, characterized by, The molecular detection unit comprises a substrate, a first structure layer, a second structure layer, a sensing electrode, a third structure layer, a first buffer solution flow channel, a sample flow channel, a single-hole liquid storage cavity, a liquid resistance flow channel and a second buffer solution flow channel; The first structure layer is arranged on the top of the substrate; The second structure layer is arranged above the first structure layer; The third structure layer is arranged above the second structure layer; The liquid resistance flow channel and the single-hole liquid storage cavity are arranged in the first structure layer; the liquid resistance flow channel is in a linear or curved shape; The bottom of the single-hole liquid storage cavity is in communication with one end of the liquid resistance flow channel; The sample flow channel is arranged in the second structure layer, above the single-hole liquid storage cavity and in communication with the single-hole liquid storage cavity; The first buffer solution flow channel is arranged in the third structure layer; The bottom of the second buffer solution flow channel is in communication with the other end of the liquid resistance flow channel; the top of the second buffer solution flow channel is in communication with the first buffer solution flow channel; The sensing electrode is arranged at the communication position of the single-hole liquid storage cavity and the liquid resistance flow channel.

2. The molecular detection unit of claim 1, wherein, The molecular detection unit further comprises a membrane layer; The membrane layer is arranged at the connection position of the sample flow channel and the single-hole liquid storage cavity; Nanopores are arranged on the membrane layer.

3. The molecular detection unit of claim 1, wherein, The molecular detection unit further comprises a driving electrode; The driving electrode is arranged in the third structure layer and connected with the first buffer solution flow channel.

4. The molecular detection unit of claim 1, wherein, The molecular detection unit further comprises two sample solution outlets and two buffer solution outlets; The two sample solution outlets are respectively in communication with the sample flow channel; The two buffer solution outlets are respectively in communication with the first buffer solution flow channel.

5. The molecular detection unit of claim 4, wherein, The molecular detection unit further comprises a sample flow channel interface and a buffer solution flow channel interface; The sample flow channel interface is arranged in the sample solution outlet, and the top of the sample flow channel interface extends from the top of the sample solution outlet; the bottom of the sample flow channel interface is in communication with the sample flow channel; The buffer solution flow channel interface is arranged in the buffer solution outlet, and the top of the buffer solution flow channel interface extends from the top of the buffer solution outlet; the bottom of the buffer solution flow channel interface is in communication with the first buffer solution flow channel.

6. The molecular detection unit according to claim 5, wherein: The first driving electrode is arranged in the sample flow channel interface, and the second driving electrode is arranged in the buffer solution flow channel interface.

7. A molecular detection chip, characterized by, The molecular detection chip comprises a molecular detection array; The molecular detection array comprises at least one molecular detection unit group; each molecular detection unit group comprises a plurality of molecular detection units according to any one of claims 1-6; wherein the sample flow channels of the molecular detection units are in communication with each other to form a common sample flow channel; and / or the first buffer solution flow channels of the molecular detection units are in communication with each other to form a first common buffer solution flow channel; and / or the second buffer solution flow channels of the molecular detection units are in communication with each other to form a second common buffer solution flow channel.

8. The molecular detection chip of claim 7, wherein, The molecular detection chip further comprises two common sample solution outlets and / or two common buffer solution outlets; The two common sample solution outlets are respectively in communication with the common sample flow channel; The two common buffer solution outlets are respectively in communication with the common buffer solution flow channel. The two common buffer inlets are respectively communicated with the first common buffer flow channel.

9. The molecular detection chip of claim 8, wherein, The molecular detection chip further comprises a common sample flow channel interface and / or a common buffer flow channel interface. The common sample flow channel interface is arranged in the common sample solution inlet, and the top of the common sample flow channel interface extends from the top of the common sample solution inlet; the bottom of the common sample flow channel interface is communicated with the common sample flow channel. The common buffer flow channel interface is arranged in the common buffer inlet, and the top of the common buffer flow channel interface extends from the top of the common buffer inlet; the bottom of the common buffer flow channel interface is communicated with the first common buffer flow channel.

10. The molecular detection chip according to claim 9, wherein: The first common driving electrode is arranged in the common sample flow channel interface. The second common driving electrode is arranged in the common buffer flow channel interface.

11. The molecular detection chip according to claim 7, wherein: The first buffer flow channels in each of the molecular detection units in the molecular detection array are communicated with each other to form a first common buffer flow channel. The second buffer flow channels in each of the molecular detection units in the molecular detection array are respectively communicated with the first common buffer flow channel.

12. The molecular detection chip according to claim 7, wherein: The plurality of molecular detection units in the same molecular detection unit group are uniformly arranged around a common center.

13. The molecular detection chip according to claim 12, wherein: The sample flow channel, the first buffer flow channel and / or the second buffer flow channel in each of the molecular detection units are in a cylindrical shape.

14. The molecular detection chip according to claim 7, wherein: The plurality of molecular detection units in the same molecular detection unit group are sequentially arranged in a straight line.

15. The molecular detection chip according to claim 14, wherein: When the molecular detection array comprises one or more molecular detection unit groups, the number of common sample flow channels and / or first common buffer flow channels in the molecular detection array is equal to the number of molecular detection unit groups in the molecular detection array.

16. The molecular detection chip according to claim 14, wherein: When the molecular detection array comprises a plurality of molecular detection unit groups, the number of common sample flow channels and / or first common buffer flow channels in the molecular detection array is less than the number of molecular detection unit groups in the molecular detection array.

Images