Spatial omics detection tool, preparation method thereof and spatial omics detection method

Abstract

The invention provides a space omics detection tool, a preparation method thereof and a space omics detection method. In the detection tool, detection site electrodes arranged in an array are communicated with a liquid changing micro-channel; the control system controls the power-on state of the detection site electrode by using the site power-on structure; the micro-fluidic liquid changing system, the microelectrode array and the site electrifying structure are MEMS (Micro Electro Mechanical System) structures. A space omics detection tool is obtained by adopting an MEMS structure, the space resolution precision of space omics detection is improved to a subsingle cell level, and the space omics detection cost and detection complexity are reduced; meanwhile, the microelectrode array and a site power-on structure MEMS are integrated through a field effect transistor and a capacitor, the number of bonding pads is reduced, the packaging difficulty is lowered, and the spatial resolution precision of spatial omics detection is further improved; in addition, a detection site electrode is electrically connected with an external bonding pad from the bottom, so that the spatial resolution precision of spatial omics detection is further improved; and finally, a detachable probe card is arranged as a packaging structure, so that the replacement efficiency of the space omics detection tool is improved.

Description

Technical Field

[0001] The present invention belongs to the fields of biotechnology and semiconductor integrated circuit manufacturing technology, and in particular relates to a spatial omics detection tool and a preparation method thereof and a spatial omics detection method. Background Art

[0002] Spatial omics technologies such as spatial transcriptomics and genomics can implement non-targeted detection of genes (mutation information) and their expression characteristics at single-cell or even subcellular resolution, and accurately obtain the spatial distribution characteristics of gene expression at the molecular level, which can accurately describe the heterogeneity of cell composition and spatial cell interactions, molecular interactions, etc., so as to analyze the differences in individual drug sensitivity and side effects with higher precision and more data dimensions. In recent years, the application of spatial omics technologies in basic life science research and major disease research in immunity, development, neurology, and brain science has achieved many heavyweight results, reflecting the huge development potential of spatial omics technologies such as spatial transcriptomics and genomics.

[0003] The acquisition of spatial position information is the core of spatial omics technology. In the prior art, the spatial information acquisition methods of spatial transcriptomics and genomics technology can be divided into microdissection, imaging and spatial barcodes. Among them, the microdissection method combined with NGS (Next generation sequencing) can perform high-resolution and in-depth analysis of the whole transcriptome, but the equipment used for microdissection is very expensive, and its sequencing throughput is still low due to factors such as the operating equipment of microdissection; the method of obtaining spatial information through imaging includes in situ hybridization (such as seqFISH, merFISH, etc.) and in situ sequencing (STARMap, Barseq, etc.), both of which use fluorescent probes to hybridize with genes or transcripts in situ and collect images to obtain sequence and position information, but due to spectral bandwidth limitations, multiple rounds of hybridization operations are required, which makes the operation relatively complex and the image data volume is huge, inefficient, and requires expensive high-resolution microscopes and molecular probes; in contrast, spatial barcode technology no longer relies on imaging, but captures genes / transcripts through nucleotide coding arrays, and then combines NGS sequencing technology to obtain sequence and position information, which can reduce equipment costs and attract widespread attention in the industry.

[0004] Spatial barcode technology through inkjet printing Methods such as microbead arrays (HDST, Slide-seq) and microfluidics-assisted (DBiT-seq, Decoder-seq) form spatial barcode arrays, and the spatial resolution accuracy is also reduced from 55μm to close to the single-cell level. However, due to the structural density limitations of inkjet, microbeads and other structures themselves, these methods are also limited in improving the spatial resolution accuracy of spatial omics analysis. In 2022, the Stereo-seq reported by the BGI research team in my country pushed the resolution of spatial omics analysis to the submicron level, but its equipment cost and operation complexity have increased.

[0005] Therefore, there is an urgent need for a structure or method that can improve the spatial resolution accuracy of spatial omics analysis while reducing equipment costs and improving operational efficiency.

[0006] It should be noted that the above introduction to the technical background is only for the convenience of providing a clear and complete description of the technical solutions of the present application and for the convenience of understanding by those skilled in the art. It cannot be considered that the above technical solutions are well known to those skilled in the art simply because these solutions are described in the background technology part of the present application. Summary of the invention

[0007] In view of the above shortcomings of the prior art, an object of the present invention is to provide a spatial omics detection tool and a preparation method thereof and a spatial omics detection method, so as to solve the problems in the prior art.

[0008] To achieve the above-mentioned object, the present invention provides a spatial omics detection tool, which comprises: a microfluidic fluid exchange system, a microelectrode array, an electrical structure on a site, and a control system;

[0009] The microfluidic fluid exchange system includes a fluid exchange microchannel with a controllable on-off state, and the microelectrode array includes n detection site electrodes arranged in an array, where n is an integer greater than or equal to 1; the fluid exchange microchannel is connected to each of the detection site electrodes to pass label raw materials from the fluid exchange microchannel to the detection site electrodes, and the detection site electrodes are used to electrochemically synthesize label nucleic acid sequences from the label raw materials passed into the detection site electrodes after power is turned on, and the label nucleic acid sequences are used for capturing DNA and preparing libraries in spatial omics technology;

[0010] The electrical structure on the site is electrically connected to each of the detection site electrodes, the control system is electrically connected to the electrical structure on the site, and the control system controls the power-on state of each of the detection site electrodes through the electrical structure on the site;

[0011] The microfluidic fluid exchange system, the microelectrode array and the electrical structure on the site are MEMS structures.

[0012] Optionally, the spatial omics detection tool includes n electrical structures on the sites, each of the electrical structures on the sites includes electrically connected connecting wires and external solder pads, and the external solder pads are electrically connected to the control system; each of the detection site electrodes is electrically connected to the connecting wires of one of the electrical structures on the sites, and the control system controls the power-on state of each of the detection site electrodes through the connecting wires and the external solder pads.

[0013] Optionally, the microelectrode array and the electrical structure at the site are MEMS integrated circuits; the electrical structure at the site includes n capacitors, n identical field effect transistors arranged in an array of a rows and b columns, a row control lines, b column control lines and n external pads, where a and b are both integers greater than or equal to 1;

[0014] Each gate electrode of the field effect transistors in each row is connected to a row control line, each source electrode of the field effect transistors in each column is connected to a column control line, and one end of each row control line and column control line not connected to the field effect transistor is electrically connected to one of the external pads;

[0015] Each of the capacitors is connected to a corresponding field effect transistor and a detection site electrode; each of the capacitors includes a first electrode, a second electrode and an insulating medium, the first electrode is electrically connected to the corresponding detection site electrode, the second electrode is electrically connected to the drain of the corresponding field effect transistor, and the insulating medium is located between the first electrode and the second electrode; the control system controls the power-on state of the corresponding detection site electrode by controlling the input electrical signals of the row control line and the column control line of the field effect transistor.

[0016] Optionally, the first electrode is a polysilicon region at a preset position around the drain connected to the capacitor, the second electrode is the drain connected to the capacitor, and the insulating medium is a silicon dioxide layer located between the first electrode and the second electrode.

[0017] Optionally, the field effect transistor is an NMOS transistor.

[0018] Optionally, the spatial omics detection tool also includes a probe card, which is used to encapsulate the microelectrode array and the electrical structure at the site; the probe card includes n probes and a PCB circuit; one end of each of the probes is electrically connected to the external pad corresponding to one of the detection site electrodes, and the other end of each of the probes leads the electrical connection of the external pad to the packaging pins on the outer surface through the PCB circuit, and the control system is electrically connected to the packaging pins to control the power-on state of the detection site electrodes.

[0019] Optionally, n is greater than or equal to 5000.

[0020] Optionally, the spacing between two adjacent detection site electrodes is less than or equal to 10 microns, or the spacing between two adjacent detection site electrodes is less than or equal to 5 microns.

[0021] The present invention also provides a spatial omics detection method, which is performed using any one of the above-mentioned spatial omics detection tools, and the detection method comprises:

[0022] Controlling the liquid exchange microchannel in the microfluidic liquid exchange system to pass the label raw material to the detection site electrode in the microelectrode array; controlling the detection site electrode through which the label raw material is passed to be energized by the control system, so as to electrochemically synthesize the label nucleic acid sequence from the label raw material at the detection site electrode;

[0023] The control system controls the detection site electrode to be powered off; after perforating the cell membrane of the tissue slice to be detected, the tissue slice to be detected is transferred to the detection site electrode;

[0024] Performing DNA unbinding and DNA cutting on the tissue slice to be detected transferred to the detection site electrode to obtain multiple DNA fragments, so that each DNA fragment obtained after cutting is captured by the label nucleic acid sequence corresponding to the detection site electrode;

[0025] The tag nucleic acid sequence of the DNA fragment captured by the detection site electrode is subjected to library preparation, so that the DNA fragment captured by the tag nucleic acid sequence replicates the tag structure of the corresponding tag nucleic acid sequence, and the DNA fragment with the tag structure is collected and processed into a structure that can be sequenced to obtain a transcript;

[0026] The DNA fragments with the tag structure in the transcripts obtained by library preparation and collection are subjected to high-throughput sequencing to obtain the spatial position information of the DNA fragments in the tissue section to be detected.

[0027] The present invention also provides a method for preparing a spatial omics detection tool, the method for preparing the aforementioned spatial omics detection tool, the method comprising:

[0028] Providing a substrate having a first oxide layer disposed on its upper surface;

[0029] Disposing a first metal layer on the first oxide layer;

[0030] Patterning the first metal layer;

[0031] Covering the gaps between and the upper surface of the patterned first metal layers with a second oxide layer;

[0032] Patterning the second oxide layer above the patterned first metal layer to expose a portion of the first metal layer below; providing a second metal layer in the gap between the patterned second oxide layers, wherein the second metal layer forms a corresponding electrical connection with the first metal layer below;

[0033] Disposing a third metal layer on the surface of the second metal layer, wherein the third metal layer covers the second metal layer and the second oxide layer exposed on the surface;

[0034] The third metal layer is patterned to obtain a microelectrode array and an external pad electrically connected to the second metal layer, wherein the microelectrode array includes n detection site electrodes arranged in an array, and each detection site electrode is electrically connected to the first metal layer and a corresponding external pad via the second metal layer below; the detection site electrode is used to electrochemically synthesize a label nucleic acid sequence for a label raw material passed into the detection site electrode after power is supplied through the external pad, and the label nucleic acid sequence is used for capturing DNA and preparing a library in spatial omics technology.

[0035] As described above, the spatial omics detection tool and preparation method thereof and the spatial omics detection method of the present invention have the following beneficial effects:

[0036] The present invention uses MEMS devices to obtain detection site electrodes that can electrochemically synthesize label nucleic acid sequences for DNA capture, thereby greatly improving the density of detection site electrodes, thereby reducing the spatial resolution accuracy of spatial omics detection to a sub-single-cell level, which can be adapted to the spatial omics analysis of larger area tissue sections, and greatly reduces the equipment cost for spatial omics detection, which is conducive to improving the efficiency of spatial omics detection;

[0037] The present invention integrates the microelectrode array and the electrical structure at the site through the structure of field effect tube and capacitor through MEMS integrated circuit, reduces the number of conductive pads required, reduces the packaging difficulty, and is conducive to further improving the electrode density at the detection site;

[0038] The present invention further increases the density of the detection site electrodes by electrically connecting the detection site electrodes to the external pads from the bottom metal, thereby improving the resolution of spatial omics detection;

[0039] The present invention improves the replacement efficiency of spatial omics detection tools by providing a detachable probe card as a packaging structure. BRIEF DESCRIPTION OF THE DRAWINGS

[0040] Figure 1 Shown is a schematic diagram of a partial explosion diagram of the three-dimensional structure of the spatial omics detection tool in Example 1 of the present invention.

[0041] Figure 2 Shown is a schematic diagram of a partial explosion diagram of the three-dimensional structure of the spatial omics detection tool in Example 2 of the present invention.

[0042] Figure 3 Shown is a schematic diagram of the electrical structure at the site of the spatial omics detection tool in Example 2 of the present invention.

[0043] Figure 4 Shown is a schematic diagram of the electrical connection structure of a single field effect transistor of the spatial omics detection tool in Example 2 of the present invention.

[0044] Figure 5 A schematic side cross-sectional view of a semiconductor structure showing the electrical structure at a site of a spatial omics detection tool in Example 2 of the present invention.

[0045] Figure 6 It shows a schematic diagram of the structure presented by the synthetic label nucleic acid sequence in step A1 of the spatial omics detection method in Example 3 of the present invention.

[0046] Figure 7 It shows a schematic diagram of the structure presented by providing a substrate in step B1 of the method for preparing a spatial omics detection tool in Example 4 of the present invention.

[0047] Figure 8 It is a schematic diagram of the structure presented by setting the first metal layer in step B2 of the preparation method of the spatial omics detection tool in Example 4 of the present invention.

[0048] Fig. 9 It is a schematic diagram showing the structure of the patterned first metal layer in step B3 of the method for preparing a spatial omics detection tool in Example 4 of the present invention.

[0049] Fig.10 It is a schematic diagram of the structure presented by covering the second oxide layer in step B4 of the preparation method of the spatial omics detection tool in Example 4 of the present invention.

[0050] Fig.11 It is a schematic diagram of the structure presented by setting the second metal layer in step B5 of the preparation method of the spatial omics detection tool in Example 4 of the present invention.

[0051] Fig.12 It is a schematic diagram of the structure presented by setting the third metal layer in step B6 of the preparation method of the spatial omics detection tool in Example 4 of the present invention.

[0052] Fig.13 It is a schematic diagram of the structure presented by the patterned third metal layer in step B7 of the preparation method of the spatial omics detection tool in Example 4 of the present invention.

[0053] Component number description

[0054] 110, microfluidic fluid exchange system; 111, fluid exchange microchannel; 112, label nucleic acid sequence; 113, label structure; 120, microelectrode array; 121, detection site electrode; 131, connecting wire; 132, external pad; 140, control system; 210, capacitor; 211, first electrode; 212, second electrode; 213, insulating medium; 214, silicon dioxide layer; 215, polysilicon region; 220, field effect tube; 221, drain; 222, gate; 223, source; 224, channel region; 231, row control line; 232, column control line; 233, ground line; 234, row addressing integrated circuit; 235, column addressing integrated circuit; 310, substrate; 311, first oxide layer; 312, second oxide layer; 321, first metal layer; 322, second metal layer; 323, third metal layer; 324, titanium layer; 325, platinum layer. DETAILED DESCRIPTION

[0055] The following describes the embodiments of the present invention through specific examples, and those skilled in the art can easily understand other advantages and effects of the present invention from the contents disclosed in this specification. The present invention can also be implemented or applied through other different specific embodiments, and the details in this specification can also be modified or changed in various ways based on different viewpoints and applications without departing from the spirit of the present invention.

[0056] For example, when describing the embodiments of the present invention in detail, for the sake of convenience, the schematic diagrams showing the device structure will not be partially enlarged according to the general scale, and the schematic diagrams are only examples, which should not limit the scope of protection of the present invention. In addition, in actual production, the three-dimensional dimensions of length, width and depth should be included.

[0057] For convenience of description, spatially relative terms such as "under", "below", "below", "below", "above", "on", etc. may be used herein to describe the relationship of one element or feature shown in the drawings to other elements or features. It will be understood that these spatially relative terms are intended to encompass other orientations of the device in use or operation in addition to the orientation depicted in the drawings.

[0058] In the context of the present application, a structure in which a first feature is described as being "above" a second feature may include embodiments in which the first and second features are in direct contact, and may also include embodiments in which additional features are formed between the first and second features, such that the first and second features may not be in direct contact.

[0059] It should be noted that the illustrations provided in this embodiment are only used to illustrate the basic concept of the present invention in a schematic manner, and therefore the illustrations only show components related to the present invention rather than being drawn according to the number, shape and size of components in actual implementation. In actual implementation, the type, quantity and proportion of each component may be changed arbitrarily, and the component layout may also be more complicated.

[0060] Embodiment 1:

[0061] This embodiment provides a spatial omics detection tool, such as Figure 1 As shown, the spatial omics detection tool includes: a microfluidic fluid exchange system 110, a microelectrode array 120, and an electrical structure and control system 140 on the site;

[0062] The microfluidic fluid exchange system 110 includes a fluid exchange microchannel 111 whose on / off state can be controlled. The microelectrode array 120 includes n detection site electrodes 121 arranged in an array, where n is an integer greater than or equal to 1. The fluid exchange microchannel 111 is connected to each of the detection site electrodes 121 to allow label raw materials to be passed from the fluid exchange microchannel 111 to the detection site electrodes 121. The detection site electrodes 121 are used to electrochemically synthesize label nucleic acid sequences 112 ( Figure 6 ), the tag nucleic acid sequence 112 is used for capturing DNA and preparing libraries in spatial omics technology;

[0063] The electrical structure on the site is electrically connected to each of the detection site electrodes 121, and the control system 140 is electrically connected to the electrical structure on the site. The control system 140 controls the power-on state of each of the detection site electrodes 121 through the electrical structure on the site;

[0064] The microfluidic fluid exchange system 110 , the microelectrode array 120 and the electrical structure on the site are MEMS structures.

[0065] In the prior art, the spatial information acquisition methods of spatial transcriptome and genomics technology can be divided into microdissection, imaging and spatial barcoding. Among them, the microdissection method combined with NGS (Next generationsequencing) can perform high-resolution and in-depth analysis of the whole transcriptome, but the equipment used for microdissection is very expensive, and its sequencing throughput is still low due to factors such as the operating equipment of microdissection; the methods of obtaining spatial information through imaging include in situ hybridization (such as seqFISH, merFISH, etc.) and in situ sequencing (STARMap, Barseq, etc.), both of which use fluorescent probes to hybridize in situ with genes or transcripts and collect images to obtain sequence and position information, but due to spectral bandwidth limitations, multiple rounds of hybridization operations are required, which makes the operation relatively complex and the image data volume is huge, inefficient, and requires expensive high-resolution microscopes and molecular probes; in contrast, spatial barcoding technology no longer relies on imaging, but captures genes / transcripts through nucleotide coding arrays, and then combines NGS sequencing technology to obtain sequence and position information, which can reduce equipment costs. The current spatial barcoding technology uses inkjet printing to obtain sequence and position information. Methods such as microbead arrays (HDST, Slide-seq), microfluidics-assisted (DBiT-seq, Decoder-seq), etc. form spatial barcode arrays, and the spatial resolution accuracy is also reduced from 55μm to near the single-cell level. However, due to the structural density limitations of structures such as inkjet and microbeads themselves, these methods are also limited in improving the spatial resolution accuracy of spatial omics analysis. In 2022, the Stereo-seq reported by the BGI research team in my country pushed the resolution of spatial omics analysis to the submicron level, but its equipment cost and operation complexity have increased. At the same time, since the field of spatial omics analysis in the prior art generally uses traditional tools such as microscopes and probes for detection, MEMS structures and processes are not used for spatial omics analysis and detection.

[0066] The present invention adopts a cross-domain technical solution, breaking the technical prejudice and blind spots of detection tools in traditional spatial omics analysis. By adopting MEMS devices as spatial omics detection tools, a detection site electrode 121 that can electrochemically synthesize a label nucleic acid sequence 112 for DNA capture is obtained. The high integration that can be achieved using the mature MEMS process is far superior to the site density of spatial barcodes formed by inkjet, microbeads, etc. in the prior art, and the density of the detection site electrode 121 is greatly improved, thereby reducing the spatial resolution accuracy of spatial omics detection to the sub-single-cell level, which can be adapted to the spatial omics analysis of larger area tissue slices, and achieves a breakthrough in the field of spatial omics technology; at the same time, the batch preparation cost of MEMS devices is greatly reduced compared with the microscope equipment used in the prior art, thereby greatly reducing the equipment cost for spatial omics detection, and at the same time, the sequencing throughput that can be achieved by a single spatial omics detection tool is also improved; and because the MEMS device can be used as a spatial omics detection tool to directly process and label DNA by powering on and introducing a biochemical reaction solvent, the operation is simple and fast, which is conducive to improving the efficiency of spatial omics detection.

[0067] Specifically, the spatial resolution accuracy of spatial omics detection also refers to the spatial resolution, that is, the minimum size that can be resolved in spatial omics detection.

[0068] Specifically, each detection site electrode 121 includes a circular wall and a solid circular protrusion, wherein the solid circular protrusion is located within the inner diameter of the circular wall, and there is a preset distance between the inner diameter of the circular wall and the circumference of the solid circle, and the circular groove formed by the preset distance is used for electrochemical synthesis of the label nucleic acid sequence 112; when the label nucleic acid sequence 112 is electrochemically synthesized, the solid circular protrusion is used as the positive electrode and the circular wall is used as the negative electrode, and hydrogen ions are generated near the solid circular protrusion after power is turned on, which are used to synthesize the label nucleic acid sequence 112, and the generated hydrogen ions are absorbed near the circular wall to prevent the hydrogen ions generated in one detection site electrode 121 from moving to the adjacent detection site electrode 121, thereby affecting the synthesis of the label nucleic acid sequence 112 in other detection site electrodes 121.

[0069] In one embodiment, the distance between the inner diameter and the outer diameter of the annular wall is 500 nanometers, and the outer diameter of the solid circular protrusion is 1 micrometer.

[0070] In this embodiment, the spatial omics detection tool includes n electrical structures on the sites, each of the electrical structures on the sites includes an electrically connected connecting wire 131 and an external solder pad 132, and the external solder pad 132 is electrically connected to the control system 140; each of the detection site electrodes 121 is electrically connected to the connecting wire 131 of one of the electrical structures on the sites, and the control system 140 controls the power-on state of each of the detection site electrodes 121 through the connecting wire 131 and the external solder pad 132.

[0071] In the present invention, the power-on state of the detection site electrode 121 is controlled directly by connecting the wire 131 and the external pad 132, which can further reduce the integrated circuit tape-out cost and reduce the spatial omics detection cost.

[0072] In one embodiment, the spatial omics detection tool also includes a probe card, which is used to encapsulate the microelectrode array 120 and the electrical structure at the site; the probe card includes n probes and a PCB circuit; one end of each of the probes is electrically connected to the external pad 132 corresponding to one of the detection site electrodes 121, and the other end of each of the probes leads the electrical connection of the external pad 132 to the packaging pins on the outer surface through the PCB circuit, and the control system 140 is electrically connected to the packaging pins to control the power-on state of the detection site electrode 121.

[0073] The present invention omits the need for wire bonding of integrated circuits by using a detachable probe card as the packaging structure of the spatial omics detection tool, thereby improving packaging efficiency. At the same time, since the probe card can be directly removed to package the new microelectrode array 120 and the electrical structure on the site, it is convenient to replace the integrated circuit inside the spatial omics detection tool, further reducing the maintenance and replacement costs and improving maintenance efficiency.

[0074] In one embodiment, the control system 140 is a single chip microcomputer or a FPGA (Field Programmable Gate Array), and the package pin header is electrically connected to the control system 140 via a DuPont line.

[0075] In one embodiment, the control system 140 may be a single power supply including only a controllable switch electrically connected to all the detection site electrodes 121. This approach has a simple structure, but cannot perform individual targeted control of the detection site electrodes 121. It can be selected based on application requirements.

[0076] In one embodiment, n is greater than or equal to 5,000.

[0077] The present invention realizes a spatial omics detection tool by using a MEMS structure, and can realize the setting of more than 5,000 detection site electrodes 121 on the same spatial omics detection tool, which greatly improves the spatial resolution accuracy of spatial omics detection and can adapt to the spatial omics analysis needs of larger area tissue slices.

[0078] In one embodiment, the distance between two adjacent detection site electrodes 121 is less than or equal to 10 micrometers, or the distance between two adjacent detection site electrodes 121 is less than or equal to 5 micrometers.

[0079] The present invention obtains a spatial omics detection tool by using a MEMS structure and a MEMS process, which can achieve an integration density of detection site electrodes 121 with a spacing of less than or equal to 10 microns, or even less than or equal to 5 microns, thereby greatly improving the spatial resolution accuracy of spatial omics detection.

[0080] Embodiment 2:

[0081] This embodiment provides a spatial omics detection tool, which is substantially the same as that in Embodiment 1 in other features, except that:

[0082] In this embodiment, Figure 2-Figure 5 As shown, Figure 2 is a schematic diagram of the three-dimensional structure of the spatial omics detection tool, Figure 3 is a schematic diagram of the electrical structure at the site, Figure 4 for Figure 3 The schematic diagram of the electrical connection structure of a single field effect tube 220 in the middle M part, Figure 5 A side cross-sectional view of a semiconductor structure of an electrical structure at each site; the microelectrode array 120 and the electrical structure at the site are MEMS integrated circuits; the electrical structure at the site includes n capacitors 210, n identical field effect transistors 220 arranged in an array of a rows and b columns, a row control lines 231, b column control lines 232 and n external pads 132, where a and b are both integers greater than or equal to 1;

[0083] Each gate electrode 222 of the field effect transistors 220 in each row is connected to a row control line 231, each source electrode 223 of the field effect transistors 220 in each column is connected to a column control line 232, and one end of each row control line 231 and column control line 232 that is not connected to the field effect transistor 220 is electrically connected to one of the external pads 132;

[0084] Each of the capacitors 210 is connected to a corresponding field effect transistor 220 and a detection site electrode 121; each of the capacitors 210 includes a first electrode 211, a second electrode 212 and an insulating medium 213, the first electrode 211 is electrically connected to the corresponding detection site electrode 121, the second electrode 212 is electrically connected to the drain 221 of the corresponding field effect transistor 220, and the insulating medium 213 is located between the first electrode 211 and the second electrode 212; the control system 140 controls the power-on state of the corresponding detection site electrode 121 by controlling the input electrical signals of the row control line 231 and the column control line 232 of the field effect transistor 220.

[0085] The present invention integrates the microelectrode array 120 and the electrical structure on the site through the structure of the field effect transistor 220 and the capacitor 210 through a MEMS integrated circuit, which greatly reduces the number of external pads 132 required, reduces the difficulty of packaging, and is conducive to further improving the density of the detection site electrodes 121, thereby further improving the spatial resolution accuracy of spatial omics detection; but at the same time, the method of integrating the microelectrode array 120 and the electrical structure on the site using a MEMS integrated circuit is more expensive to prepare than the method of directly connecting the microelectrode array 120 and the external pad 132 through the connecting wire 131 in Example 1. Those skilled in the art can choose between the two schemes based on the requirements of cost and spatial resolution accuracy, but overall, the cost of the scheme in this embodiment is also much lower than the cost required to achieve the same spatial resolution accuracy in the prior art.

[0086] Specifically, Figure 5 As shown, the channel region 224 of the field effect transistor 220 is located between the source 223 and the drain 221 .

[0087] In one embodiment, Figure 2-Figure 3 As shown, the row control line 231 is connected to the row addressing integrated circuit 234, and the column control line 232 is connected to the column addressing integrated circuit 235. The row addressing integrated circuit 234 and the column addressing integrated circuit 235 can be connected to the external pad 132 through the row control line 231 and the column control line 232, and then controlled by the control system 140 through the external pad 132; the row addressing integrated circuit 234 and the column addressing integrated circuit 235 can also be connected to the row control line 231 and the column control line 232 by connecting to the external pad 132. The row addressing integrated circuit 234 and the column addressing integrated circuit 235 can be placed in the control system 140 to further reduce the line density required to be set in the MEMS integrated circuit composed of the microelectrode array 120 and the electrical structure on the site. When problems occur later, only the MEMS integrated circuit can be replaced without replacing the control system 140, so as to reduce the maintenance cost.

[0088] Specifically, when the row control line 231 and the column control line 232 connected to a detection site electrode 121 are both selected, the detection site electrode 121 is powered on, otherwise it is in a power-off state.

[0089] In one embodiment, Figure 5 As shown, the first electrode 211 is a polysilicon region 215 at a preset position around the drain 221 corresponding to the capacitor 210, the second electrode 212 is the drain 221 corresponding to the capacitor 210, and the insulating medium 213 is a silicon dioxide layer 214 located between the first electrode 211 and the second electrode 212.

[0090] The present invention uses the drain 221 of the field effect transistor 220 itself as the second electrode 212 of the capacitor 210, and realizes the structure of the capacitor 210 through the polysilicon in the conventional semiconductor structure and the silicon dioxide layer 214 outside the polysilicon, so that it can be directly prepared using the mature processes and equipment in the existing MEMS integrated circuit process without the need for special customized materials or tools, thereby further improving the possibility of popularizing the production of spatial omics detection tools and reducing the preparation cost.

[0091] In one embodiment, Figure 4 As shown, the field effect transistor 220 is an NMOS transistor.

[0092] Specifically, the field effect transistor 220 can also be a PMOS tube or other types of field effect transistors 220, and its specific connection structure can be adjusted according to needs, but the NMOS tube is used as the field effect transistor 220 to realize the power-on state control of the detection site electrode 121. It can achieve the characteristics of low power consumption and high response speed of spatial omics detection tools through smaller on-resistance and faster switching speed, thereby improving the utilization efficiency of spatial omics detection tools.

[0093] Specifically, the field effect transistor 220 may also be replaced by other suitable semiconductor switch structures to realize power-on control of the detection site electrode 121, all of which are within the protection scope of the present invention.

[0094] Preferably, when the field effect transistor 220 is an NMOS transistor, the source 223 and the drain 221 are heavily N-type doped, the substrate 310 is a P-type silicon substrate, and the polysilicon region 215 is electrically connected to the ground wire 233 to achieve the same electric potential of the drain 221 of all field effect transistors 220, thereby ensuring the reliability of controlling the power-on state of the detection site electrode 121.

[0095] Embodiment 3:

[0096] This embodiment provides a spatial omics detection method, which is performed using any one of the spatial omics detection tools described in Embodiments 1-2.

[0097] The spatial omics detection method of the present invention will be described in detail below with reference to the accompanying drawings. It should be noted that the above sequence does not strictly represent the sequence of the spatial omics detection method protected by the present invention, and those skilled in the art may change it according to the actual preparation steps.

[0098] First, step A1 is performed to control the liquid exchange microchannel 111 in the microfluidic liquid exchange system 110 to pass the label raw material to the detection site electrode 121 in the microelectrode array 120; the control system 140 controls the detection site electrode 121 into which the label raw material is passed to be energized, so that the label raw material of the detection site electrode 121 is electrochemically synthesized as follows: Figure 6 The tag nucleic acid sequence 112 is shown.

[0099] Specifically, Figure 6 As shown, each tag nucleic acid sequence 112 includes multiple different tag structures 113 . The figure only shows a structure in which one tag nucleic acid sequence 112 includes multiple tag structures 113 . In fact, each tag nucleic acid sequence 112 includes multiple different tag structures 113 .

[0100] Then, step A2 is performed, wherein the control system 140 controls the detection site electrode 121 to be powered off; and after perforating the cell membrane of the tissue slice to be detected, the tissue slice to be detected is transferred to the detection site electrode 121 .

[0101] Specifically, by perforating the cell membrane of the tissue slice to be detected, the DNA inside the tissue slice to be detected can be more smoothly discharged for processing after being transferred to the detection site electrode 121, thereby improving the detection efficiency and ensuring the reliability of the detection result.

[0102] Next, step A3 is performed to perform DNA unbinding and DNA cutting on the tissue slice to be detected transferred to the detection site electrode 121 to obtain multiple DNA fragments, so that each DNA fragment obtained after cutting is captured by the label nucleic acid sequence 112 in the corresponding detection site electrode 121.

[0103] Specifically, the nucleic acid of the tissue section to be detected transferred to the detection site electrode 121 will be captured by the label nucleic acid sequence 112 on the detection site electrode 121 at the corresponding position in a hybridization manner.

[0104] In one embodiment, the DNA is dissolved and untied by chromosome histones. Specifically, other suitable methods can also be used to untie the DNA.

[0105] In one embodiment, the DNA is cut by Tn5 transposase to cut the DNA into multiple DNA fragments. Specifically, other suitable methods can also be used to cut the DNA.

[0106] Then, step A4 is performed to prepare a library for the label nucleic acid sequence 112 of the DNA fragment captured by the detection site electrode 121, so that the DNA fragment captured by the label nucleic acid sequence 112 replicates the corresponding label structure 113 of the label nucleic acid sequence 112, and the DNA fragments with the label structure 113 are collected and processed into a structure that can be sequenced to obtain transcripts.

[0107] Specifically, the number of label nucleic acid sequences 112 included in each detection site electrode 121 is set to be greater than the number of DNA fragments after unbinding and cleavage, so as to ensure that there are enough label nucleic acid sequences 112 in each detection site electrode 121 to capture the DNA fragments obtained after unbinding and cleavage one-to-one.

[0108] In one embodiment, each of the detection site electrodes 121 includes 10 7 -10 11 A tag nucleic acid sequence 112.

[0109] Finally, step A5 is performed to perform high-throughput sequencing on the DNA fragment with the tag structure 113 in the transcript obtained by library preparation and collection, so as to obtain the spatial position information of the DNA fragment in the tissue section to be detected.

[0110] In one embodiment, the high-throughput sequencing is a second-generation sequencing technology or a third-generation sequencing technology. Specifically, the scheme of the present invention can also be applied to the first-generation sequencing technology, but since the effect of improving the spatial resolution accuracy of the present invention cannot be fully reflected in the first-generation sequencing technology, the first-generation sequencing technology is not described.

[0111] The present invention uses MEMS structures and processes to obtain spatial omics detection tools that are applied across fields in spatial omics technology, so that the spatial barcode technology in spatial omics analysis no longer relies on imaging, but instead uses a nucleotide coding array (a label nucleic acid sequence 112 on a microelectrode array 120) to capture genes (DNA on the tissue section to be detected) to obtain transcripts, and then combines NGS sequencing technology to obtain the sequence and position information of the DNA on the tissue section to be detected. Compared with the prior art methods of using microdissection combined with NGS sequencing and in situ hybridization / in situ sequencing combined with high-resolution microscopy acquisition, the present invention has higher advantages in cost and resolution; at the same time, the present invention achieves a sub-single-cell resolution level that is difficult to achieve with other technical platforms based on spatial barcodes.

[0112] Embodiment 4:

[0113] This embodiment provides a method for preparing a spatial omics detection tool, which is used to prepare the spatial omics detection tool in Example 1, and the preparation method comprises:

[0114] Step B1: providing a substrate 310 having a first oxide layer 311 disposed on its upper surface;

[0115] Step B2: Disposing a first metal layer 321 on the first oxide layer 311;

[0116] Step B3: patterning the first metal layer 321;

[0117] Step B4: Covering the gaps between and the upper surface of the patterned first metal layer 321 with a second oxide layer 312 ;

[0118] Step B5: patterning the second oxide layer 312 above the patterned first metal layer 321 to expose a portion of the first metal layer 321 below; providing a second metal layer 322 in the gap between the patterned second oxide layers 312, and the second metal layer 322 forms a corresponding electrical connection with the first metal layer 321 below;

[0119] Step B6: disposing a third metal layer 323 on the surface of the second metal layer 322 , wherein the third metal layer 323 covers the exposed second metal layer 322 and the second oxide layer 312 ;

[0120] Step B7: The third metal layer 323 is patterned to obtain a microelectrode array 120 and an external pad 132 electrically connected to the second metal layer 322, wherein the microelectrode array 120 includes n detection site electrodes 121 arranged in an array, and each detection site electrode 121 is electrically connected to the first metal layer 321 and a corresponding external pad 132 through the second metal layer 322 below; the detection site electrode 121 is used to electrochemically synthesize a label nucleic acid sequence 112 for the label raw material passed into the detection site electrode 121 after power is supplied through the external pad 132, and the label nucleic acid sequence 112 is used for capturing DNA and preparing libraries in spatial omics technology.

[0121] The preparation method of the spatial omics detection tool of the present invention will be described in detail below with reference to the accompanying drawings. It should be noted that the above sequence does not strictly represent the sequence of the preparation method of the spatial omics detection tool protected by the present invention, and those skilled in the art may change it according to the actual preparation steps.

[0122] First, if Figure 7As shown, step B1 is performed to provide a substrate 310 having a first oxide layer 311 disposed on its upper surface.

[0123] In one embodiment, the substrate 310 is a silicon wafer.

[0124] In one embodiment, the first oxide layer 311 is a thermal oxide layer.

[0125] In one embodiment, the substrate 310 with the first oxide layer 311 is a 6-inch silicon oxide wafer, wherein the first oxide layer 311 is silicon dioxide, and the thickness of the silicon dioxide is greater than or equal to 1 micron.

[0126] Before performing step B2, the substrate 310 provided with the first oxide layer 311 is subjected to organic ultrasonic cleaning and thermal baking at 120° C. for 5 minutes to pre-treat the first oxide layer 311 and the substrate 310 to make the surface clean and free of pollution.

[0127] Then, if Figure 8 As shown, step B2 is performed to dispose a first metal layer 321 on the first oxide layer 311 .

[0128] In one embodiment, the material of the first metal layer 321 can be gold, platinum, aluminum or other suitable materials.

[0129] In one embodiment, the first metal layer 321 is a titanium / gold composite layer, wherein the thickness of the titanium layer is 20 nanometers, and the thickness of the gold layer is 200 nanometers.

[0130] In one embodiment, the first metal layer 321 is formed by magnetron sputtering.

[0131] Then, if Fig. 9 As shown, step B3 is performed to pattern the first metal layer 321 .

[0132] In one embodiment, a photoresist is disposed on the first metal layer 321; the photoresist is developed and exposed on the first metal layer 321 by a 6-inch stepper photoresist and the photoresist to expose a portion of the first metal layer 321 under the photoresist; IBE (ion beam etching) is performed on the exposed first metal layer 321 to achieve patterning of the first metal layer 321, and the line width and line spacing of the photoresist are both 500 nanometers; after the first metal layer 321 is patterned, NMP (N-methylpyrrolidone) is used to remove the remaining photoresist.

[0133] Then, if Fig.10 As shown, step B4 is performed to cover the gaps between and the upper surface of the patterned first metal layer 321 with a second oxide layer 312 .

[0134] In one embodiment, the first oxide layer 311 and the second oxide layer 312 are made of the same material to ensure adhesion between the two oxide layers and to ensure reliability of the semiconductor structure.

[0135] In one embodiment, the second oxide layer 312 is formed by PECVD (Plasma-Enhanced Chemical Vapour Deposition).

[0136] In one embodiment, the thickness of the second oxide layer 312 is 500 nanometers.

[0137] Then, if Fig.11 As shown, step B5 is performed to pattern the second oxide layer 312 above the patterned first metal layer 321 to expose a portion of the first metal layer 321 below; a second metal layer 322 is set in the gap between the patterned second oxide layers 312, and the second metal layer 322 forms a corresponding electrical connection with the first metal layer 321 below.

[0138] In one embodiment, a photoresist is disposed on the second oxide layer 312; the photoresist on the second oxide layer 312 is exposed and developed through a 6-inch stepper photoresist to expose a portion of the second oxide layer 312 under the photoresist; RIE (Reactive Ion Etching) is performed on the exposed second oxide layer 312 to achieve patterning of the second oxide layer 312, the line width and line spacing of the photoresist are both 500 nanometers, and the alignment error of the photoresist is less than 150 nanometers; and NMP (N-methylpyrrolidone) is used to remove the remaining photoresist.

[0139] In one embodiment, after the second oxide layer 312 is patterned, a second metal layer 322 is disposed on the second oxide layer 312, and the second metal layer 322 covers the second oxide layer 312 and the exposed surface of the first metal layer 321; a photoresist is disposed on the second metal layer 322, and the photoresist on the second metal layer 322 is exposed and developed using a photomask to expose a portion of the second metal layer 322 under the photoresist; IBE (Ion beametching) is performed on the exposed second metal layer 322 to obtain the second metal layer 322 between the gaps in the second oxide layer 312; and NMP (N-methylpyrrolidone) is used to remove the remaining photoresist.

[0140] Specifically, the same photomask may be used when patterning the second oxide layer 312 and the second metal layer 322 .

[0141] In one embodiment, the second metal layer 322 may be made of gold, platinum, aluminum or other suitable materials.

[0142] In one embodiment, the second metal layer 322 further includes a titanium layer, and the titanium layer contacts the first metal layer 321 to achieve a better adhesion effect and improve the reliability of the structure.

[0143] In one embodiment, the thickness of the titanium layer is 20 nanometers, and the thickness of the metal layer on the titanium layer is 200 nanometers.

[0144] Then, if Fig.12 As shown, step B6 is performed to dispose a third metal layer 323 on the surface of the second metal layer 322 , wherein the third metal layer 323 covers the exposed second metal layer 322 and the second oxide layer 312 .

[0145] In one embodiment, the material of the third metal layer 323 is platinum, and the platinum is located on the upper surface of the third metal layer 323 to ensure that when the detection site electrode 121 performs electrochemical synthesis of the label nucleic acid sequence 112, the detection site electrode 121 itself will not be corroded by the synthesis solution, so as to ensure the reliability and service life of the spatial omics detection tool and the reliability of the detection results.

[0146] In one embodiment, Fig.12 As shown, the material of the third metal layer 323 is a composite layer composed of a titanium layer 324 and a platinum layer 325, wherein the titanium layer 324 is in contact with the second metal layer 322 below to achieve a better adhesion effect and improve the reliability of the structure.

[0147] In one embodiment, the thickness of the titanium layer 324 in the third metal layer 323 is 35 nanometers, and the thickness of the platinum layer 325 is 200 nanometers.

[0148] Finally, if Fig.13 As shown, step B7 is performed to graph the third metal layer 323 to obtain a microelectrode array 120 and an external pad 132 electrically connected to the second metal layer 322, wherein the microelectrode array 120 includes n detection site electrodes 121 arranged in an array, and each of the detection site electrodes 121 is electrically connected to the first metal layer 321 and a corresponding external pad 132 through the second metal layer 322 below; the detection site electrode 121 is used to electrochemically synthesize a label nucleic acid sequence 112 for the label raw material passed into the detection site electrode 121 after power is supplied through the external pad 132, and the label nucleic acid sequence 112 is used for capturing DNA and preparing libraries in spatial omics technology.

[0149] The present invention uses the first metal layer 321 and the second metal layer 322 under the detection site electrode 121 and the external pad 132 for electrical connection to avoid excessive area occupied by too many single metal layers, thereby further improving the density of the detection site electrodes 121 that can be achieved by a single spatial omics detection tool, thereby further improving the achievable spatial resolution accuracy.

[0150] In one embodiment, before setting the third metal layer 323, a photoresist is first set on the second metal layer 322; the photoresist on the second metal layer 322 is exposed and developed through a 6-inch stepper photoresist to reveal a portion of the second metal layer 322 under the photoresist, and the alignment deviation of the photoresist is less than 150 nanometers; metal sputtering is performed on the patterned photoresist to obtain the third metal layer 323, and the third metal layer 323 fills the gaps in the photoresist and covers the surface of the exposed second metal layer 322; when removing the remaining photoresist, the third metal layer 323 covering the photoresist is simultaneously metal-lifted (Lift-Off) to leave the patterned third metal layer 323.

[0151] In one embodiment, after performing step B7, the microfluidic fluid exchange system 110 is set on the microelectrode array 120; the microfluidic fluid exchange system 110 includes a fluid exchange microchannel 111 with a controllable on-off state, and the fluid exchange microchannel 111 is connected to each of the detection site electrodes 121 in the microelectrode array 120 to pass the label raw material from the fluid exchange microchannel 111 to the detection site electrodes 121. Specifically, the microfluidic fluid exchange system 110 can also be set between other suitable steps of the preparation, all within the protection scope of the present invention.

[0152] In summary, the spatial omics detection tool and its preparation method and spatial omics detection method of the present invention can obtain detection site electrodes that can electrochemically synthesize label nucleic acid sequences for DNA capture by using MEMS devices, thereby greatly improving the density of detection site electrodes, thereby reducing the spatial resolution accuracy of spatial omics detection to the sub-single cell level, and can adapt to the spatial omics analysis of larger area tissue sections, and greatly reduce the equipment cost for spatial omics detection, which is beneficial to improving the efficiency of spatial omics detection; at the same time, the microelectrode array and the electrical structure on the site are integrated and prepared through the structure of field effect transistors and capacitors through MEMS integrated circuits, reducing the number of required conductive pads, reducing the difficulty of packaging, and facilitating further improvement of the density of detection site electrodes; in addition, by electrically connecting the detection site electrodes to the external pads from the bottom metal, the density of the detection site electrodes is further improved, thereby improving the resolution of spatial omics detection; finally, by setting a detachable probe card as a packaging structure, the replacement efficiency of the spatial omics detection tool is improved.

[0153] Therefore, the present invention effectively overcomes various shortcomings in the prior art and has high industrial utilization value.

[0154] The above embodiments are merely illustrative of the principles and effects of the present invention, and are not intended to limit the present invention. Anyone familiar with the art may modify or alter the above embodiments without departing from the spirit and scope of the present invention. Therefore, all equivalent modifications or alterations made by a person of ordinary skill in the art without departing from the spirit and technical concept disclosed by the present invention shall still be covered by the claims of the present invention.

Claims

1. A spatial omics detection tool, characterized in that: The spatial omics detection tool includes: a microfluidic fluid exchange system, a microelectrode array, a site electrical structure and a control system; The microfluidic fluid exchange system includes a fluid exchange microchannel with a controllable on-off state, and the microelectrode array includes n detection site electrodes arranged in an array, where n is an integer greater than or equal to 1; the fluid exchange microchannel is connected to each of the detection site electrodes to pass label raw materials from the fluid exchange microchannel to the detection site electrodes, and the detection site electrodes are used to electrochemically synthesize label nucleic acid sequences from the label raw materials passed into the detection site electrodes after power is turned on, and the label nucleic acid sequences are used for capturing DNA and preparing libraries in spatial omics technology; The electrical structure on the site is electrically connected to each of the detection site electrodes, the control system is electrically connected to the electrical structure on the site, and the control system controls the power-on state of each of the detection site electrodes through the electrical structure on the site; The microfluidic fluid exchange system, the microelectrode array and the electrical structure on the site are MEMS structures.

2. The spatial omics detection tool according to claim 1, characterized in that: The spatial omics detection tool includes n electrical structures on the sites, each of which includes electrically connected connecting wires and external pads, and the external pads are electrically connected to the control system; each detection site electrode is electrically connected to the connecting wire of one of the electrical structures on the sites, and the control system controls the power-on state of each detection site electrode through the connecting wires and the external pads.

3. The spatial omics detection tool according to claim 1, characterized in that: The microelectrode array and the electrical structure on the site are MEMS integrated circuits; the electrical structure on the site includes n capacitors, n identical field effect transistors arranged in an array of a rows and b columns, a row control lines, b column control lines and n external pads, where a and b are both integers greater than or equal to 1; Each gate electrode of the field effect transistors in each row is connected to a row control line, each source electrode of the field effect transistors in each column is connected to a column control line, and one end of each row control line and column control line not connected to the field effect transistor is electrically connected to one of the external pads; Each of the capacitors is connected to a corresponding field effect transistor and a detection site electrode; each of the capacitors includes a first electrode, a second electrode and an insulating medium, the first electrode is electrically connected to the corresponding detection site electrode, the second electrode is electrically connected to the drain of the corresponding field effect transistor, and the insulating medium is located between the first electrode and the second electrode; the control system controls the power-on state of the corresponding detection site electrode by controlling the input electrical signals of the row control line and the column control line of the field effect transistor.

4. The spatial omics detection tool according to claim 3, characterized in that: The first electrode is a polysilicon region at a preset position around the drain electrode connected to the capacitor, the second electrode is the drain electrode connected to the capacitor, and the insulating medium is a silicon dioxide layer located between the first electrode and the second electrode.

5. The spatial omics detection tool according to claim 3, characterized in that: The field effect tube is an NMOS tube.

6. The spatial omics detection tool according to any one of claims 1 to 5, characterized in that: The spatial omics detection tool also includes a probe card, which is used to encapsulate the microelectrode array and the electrical structure at the site; the probe card includes n probes and a PCB circuit; one end of each of the probes is electrically connected to the external pad corresponding to an electrode at the detection site, and the other end of each of the probes leads the electrical connection of the external pad to the packaging pin row on the outer surface through the PCB circuit, and the control system is electrically connected to the packaging pin row to control the power-on state of the electrode at the detection site.

7. The spatial omics detection tool according to any one of claims 1 to 5, characterized in that: n is greater than or equal to 5000.

8. The spatial omics detection tool according to any one of claims 1 to 5, characterized in that: The distance between two adjacent detection site electrodes is less than or equal to 10 microns, or the distance between two adjacent detection site electrodes is less than or equal to 5 microns.

9. A spatial omics detection method, characterized in that: The detection method is performed using the spatial omics detection tool according to any one of claims 1 to 8, and the detection method comprises: Controlling the liquid exchange microchannel in the microfluidic liquid exchange system to pass the label raw material to the detection site electrode in the microelectrode array; controlling the detection site electrode through which the label raw material is passed to be energized by the control system, so as to electrochemically synthesize the label nucleic acid sequence from the label raw material at the detection site electrode; The control system controls the detection site electrode to be powered off; after perforating the cell membrane of the tissue slice to be detected, the tissue slice to be detected is transferred to the detection site electrode; Performing DNA unbinding and DNA cutting on the tissue slice to be detected transferred to the detection site electrode to obtain multiple DNA fragments, so that each DNA fragment obtained after cutting is captured by the label nucleic acid sequence corresponding to the detection site electrode; The tag nucleic acid sequence of the DNA fragment captured by the detection site electrode is subjected to library preparation, so that the DNA fragment captured by the tag nucleic acid sequence replicates the tag structure of the corresponding tag nucleic acid sequence, and the DNA fragment with the tag structure is collected and processed into a structure that can be sequenced to obtain a transcript; The DNA fragments with the tag structure in the transcripts obtained by library preparation and collection are subjected to high-throughput sequencing to obtain the spatial position information of the DNA fragments in the tissue section to be detected.

10. A method for preparing a spatial omics detection tool, characterized in that: The preparation method is used to prepare the spatial omics detection tool described in claim 2, and the preparation method comprises: Providing a substrate having a first oxide layer disposed on its upper surface; Disposing a first metal layer on the first oxide layer; Patterning the first metal layer; Covering the gaps between and the upper surface of the patterned first metal layers with a second oxide layer; Patterning the second oxide layer above the patterned first metal layer to expose a portion of the first metal layer below; providing a second metal layer in the gap between the patterned second oxide layers, wherein the second metal layer forms a corresponding electrical connection with the first metal layer below; Disposing a third metal layer on the surface of the second metal layer, wherein the third metal layer covers the second metal layer and the second oxide layer exposed on the surface; The third metal layer is patterned to obtain a microelectrode array and an external pad electrically connected to the second metal layer, wherein the microelectrode array includes n detection site electrodes arranged in an array, and each detection site electrode is electrically connected to the first metal layer and a corresponding external pad via the second metal layer below; the detection site electrode is used to electrochemically synthesize a label nucleic acid sequence for a label raw material passed into the detection site electrode after power is supplied through the external pad, and the label nucleic acid sequence is used for capturing DNA and preparing a library in spatial omics technology.

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