Surface modification in the gas phase

Abstract

A method for surface modification is provided. [Solution] A method is provided for forming a phosphorus-containing layer on a metallic surface, which is a metal surface or a metal oxide surface, the method comprising treating the metallic surface with a phosphoryl halide in the gas phase, wherein the phosphoryl halide forms the phosphorus-containing layer on the metallic surface. In some embodiments, a method is provided for forming a stable surface coating on an oxide surface.

Description

[Technical Field]

[0001] This disclosure relates to surface modification in the gas phase. [Background technology]

[0002] Array-based platforms are widely used in massively parallel technologies for detecting and analyzing biomolecules. Examples of these platforms include microarrays used in biosensing and pixel arrays used in single-molecule sequencing. Arrays typically consist of a collection of very small regions on the surface of a single device, each region capable of independently assaying a sample. These regions may have surface areas selectively modified to be more or less reactive to a desired chemical species. Certain assays require the use of corrosive solutions or other harsh conditions that may remove the desired surface functionality or damage the underlying material on the device surface. [Overview of the project]

[0003] The aspects of the technology disclosed herein relate to surface modification methods using one or more reagents in the gas phase. In some embodiments, the application relates to the use of gas-phase reagents in surface modification processes that conventionally use liquid-phase reagents. In some embodiments, the application provides a method for forming a phosphorus-containing layer on a substrate surface using a gas-phase reagent. In some embodiments, the application provides a method for forming a stable surface coating on a substrate surface. In some embodiments, the application provides a method for preparing selectively modified portions of a substrate surface.

[0004] In some embodiments, the present application provides a method for forming a phosphorus-containing layer on a metallic surface. In some embodiments, the method comprises treating the metallic surface with a phosphoryl halide in the gas phase, the phosphoryl halide forming a phosphorus-containing layer on the metallic surface. In some embodiments, the method further comprises treating the metallic surface with at least one chlorosilane in the gas phase, the at least one chlorosilane forming a coating layer on the metallic surface. In some embodiments, the at least one chlorosilane is a chlorosiloxane compound, an alkylchlorosilane compound, or both. In some embodiments, the at least one chlorosilane is a hexachlorodisiloxane, a hexyltrichlorosilane, or both. In some embodiments, at least a portion of the coating layer is formed on the phosphorus-containing layer.

[0005] In some embodiments, the present application provides a method for modifying the surface of a substrate. In some embodiments, the method comprises treating a substrate having a first surface portion and a second surface portion with a phosphoryl halide in the gas phase, wherein the phosphoryl halide preferentially forms a phosphorus-containing layer on the first surface portion. In some embodiments, the first surface portion and the second surface portion have different surface properties. In some embodiments, the method further comprises treating the substrate with at least one chlorosilane in the gas phase, wherein the at least one chlorosilane forms a coating layer on the first surface portion and the second surface portion. In some embodiments, the at least one chlorosilane is a chlorosiloxane compound, an alkylchlorosilane compound, or both. In some embodiments, the at least one chlorosilane is a hexachlorodisiloxane, a hexyltrichlorosilane, or both. In some embodiments, at least a portion of the coating layer is formed on the phosphorus-containing layer. In some embodiments, the method further comprises treating the substrate with a functionalizing agent containing a coupling moiety before treating the substrate with a phosphoryl halide in the gas phase, and the functionalizing agent functionalizes the second surface portion by preferentially bonding to the second surface portion. In some embodiments, the first surface portion is a metallic surface or a metal oxide surface. In certain embodiments, the first surface portion is a transition metal oxide surface. In some embodiments, the second surface portion is a transparent surface or a glass surface. In some embodiments, the second surface portion is a silica (SiO2) surface. In some embodiments, the first surface portion is a metallic surface or a plastic surface, and the second surface portion is a transparent surface or a glass surface (e.g., a silica surface).

[0006] In one embodiment, the phosphoryl halide preferentially forms a phosphorus-containing layer on the first surface portion with a selectivity of about 2 to about 60 times. In another embodiment, the phosphoryl halide preferentially forms a phosphorus-containing layer on the first surface portion with a selectivity of about 4 to about 40 times. In yet another embodiment, the phosphoryl halide preferentially forms a phosphorus-containing layer on the first surface portion with a selectivity of about 8 to about 20 times.

[0007] In some embodiments, the present application provides a method for functionalizing the surface of a sample well. In some embodiments, the method comprises treating a sample well having a metal oxide surface and a silica surface with a functionalizing agent comprising a coupling portion. In some embodiments, the functionalizing agent functionalizes the sample well surface by preferentially bonding to the silica surface. In some embodiments, the method further comprises treating the sample well with an organic reagent in the gas phase, the organic reagent preferentially forming an organic layer on the metal oxide surface. In some embodiments, the organic reagent is a phosphoryl halide, and the organic layer is a phosphorus-containing layer. In some embodiments, the organic reagent is an organic phosphoryl halide, and the organic layer is an organic phosphorus layer.

[0008] In some embodiments, the method further includes treating the sample well with one or more additional reagents that form coating layers on a metal oxide surface and a silica surface. In some embodiments, the coating layer on the metal oxide surface is formed on the sidewall of the sample well, and the coating layer on the silica surface is formed on the bottom surface of the sample well. In some embodiments, the one or more additional reagents are in the gas phase. In some embodiments, the one or more additional reagents are silanes, and the coating layer is a silane layer. In some embodiments, the one or more additional reagents are chlorosilanes. In some embodiments, one or more of the additional reagents are hexachlorodisiloxane, hexyltrichlorosilane, or both. In some embodiments, the method includes coupling a target molecule to the surface of the sample well by bringing the sample well into contact with the target molecule to be bound to a coupling portion. In some embodiments, the target molecule is a biomolecule such as a nucleic acid or polypeptide. In some embodiments, the polypeptide is a protein or a fragment thereof. In certain embodiments, the coupled polypeptide is a substrate for polypeptide sequencing, i.e., the polypeptide itself will be sequenced. Methods for sequencing and identifying proteins, polypeptides, and amino acids are disclosed in International Publication No. 2020 / 102741, which is incorporated herein by reference without limitation. In certain embodiments, the polypeptide is a polymerase. In certain embodiments, the polymerase is, for example, a nucleic acid polymerase for use in nucleic acid sequencing reactions. In such nucleic acid sequencing reactions, the coupled polypeptide itself is not sequenced, but rather facilitates the sequencing of the nucleic acid substrate. Methods for sequencing and identifying polynucleotides, nucleic acids, and nucleotides are disclosed in International Publication No. 2016 / 187580, which is incorporated herein by reference without limitation.

[0009] Accordingly, in some embodiments, the present application provides a method of surface modification using a phosphoryl halide. In some embodiments, the phosphoryl halide of the present application is of formula (I):

[0010] [ka]

[0011] It is the substance or a salt thereof, where X 1 Each of these examples is an independent halogen, R 1 This includes optionally substituted alkyl, optionally substituted heteroalkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted carbocyclyl, optionally substituted heterocyclyl, optionally substituted aryl, or optionally substituted heteroaryl, -OR O , or -N(R N )2, R O Each example independently consists of hydrogen, optionally substituted alkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted carbocyclyl, optionally substituted heterocyclyl, optionally substituted aryl, optionally substituted heteroaryl, optionally substituted acyl, or an oxygen protecting group, or optionally two R O However, together with the intervening atom, it forms a heterocycline that is optionally substituted, R N Each example independently is a hydrogen atom, an optionally substituted alkyl group, an optionally substituted alkenyl group, an optionally substituted alkynyl group, an optionally substituted carbocyclyl group, an optionally substituted heterocyclyl group, an optionally substituted aryl group, an optionally substituted heteroaryl group, an optionally substituted acyl group, or a nitrogen protecting group, or optionally two R groups. N However, together with the intervening atom, it forms a heterocycline in which substitutions are made at will.

[0012] Details of one embodiment of the present invention are shown herein. Other features, purposes, and effects of the present invention will become apparent from the detailed description, drawings, examples, and claims. [Brief explanation of the drawing]

[0013] The accompanying drawings, which constitute part of this specification, illustrate several embodiments of the present invention and, together with the description, are helpful in illustrating the principles of this disclosure. [Figure 1] Figures 1A to 1C show examples of workflows for surface modification according to the present invention. Figure 1A shows a workflow for preparing a selectively functionalized surface portion of a sample well. Figure 1B shows a workflow for coating the sample well surface using a backfill process. Figure 1C shows a workflow for coupling the target molecule to the functionalized surface. [Figure 2] Figure 2 shows an example of a sample well having a surface modified according to the present invention. [Figure 3] Figure 3 shows spectra obtained from X-ray photoelectron spectroscopy (XPS) and images obtained from wettability measurements, which, overall, demonstrate the high selectivity of gas-phase phosphonic acid dichloride for metal oxide surfaces. [Figure 4A] Figures 4A and 4B show spectra obtained from XPS and images obtained from atomic force microscopy (AFM). Figure 4A shows data obtained for the surface coating formed by treatment with liquid-phase hexylphosphonic acid. [Figure 4B] Figure 4B shows the data obtained for the surface coating formed by treatment with gas-phase phosphonic acid dichloride. [Figure 5] Figure 5 shows data obtained by XPS for the stability test of the silane surface coating. [Figure 6] Figure 6 shows a plot of the percentage of silicon relative to the percentage of phosphorus in the surface coating, as measured by XPS. [Figure 7]Figure 7 shows the spectra obtained by time-of-flight secondary ion mass spectrometry (TOF-SIMS) for a control experiment used to identify the characteristic fragment peaks of biotinylated silanes. [Figure 8] Figure 8 shows the relative amounts of fragment peaks detected by TOF-SIMS for surface coatings on silica and titanium dioxide surfaces. [Figure 9] Figure 9 shows the selectivity calculation values ​​identified from the data shown in Figure 8. [Figure 10] Figure 10 shows images and spectra obtained during the chemical and physical characterization of an array with a modified surface. [Modes for carrying out the invention]

[0014] Aspects of the present application relate to methods for using gas-phase reagents to prepare substrates having modified surfaces. In some embodiments, the present application provides methods for modifying metallic or plastic surfaces of a substrate. In some embodiments, the substrate is treated with a phosphoryl halide in the gas phase to form a phosphorus-containing layer on the metallic or plastic surface of the substrate. In some embodiments, the substrate is treated with at least one silane to form a coating layer on the metallic or plastic surface of the substrate. In some embodiments, at least a portion of the coating layer is formed on the phosphorus-containing layer. In some embodiments, the present application provides methods for preparing selectively modified surface portions of a substrate, such as a sample well, having surface portions with different surface properties.

[0015] The present invention relates to the discovery of a surface modification technique that, in some embodiments, can be used to modify the exposed surface of a substrate and, among several advantages, enables selective surface functionalization and imparts corrosion resistance and / or antifouling properties to the substrate surface. Previous techniques for forming phosphorus-containing layers on metallic or plastic surfaces involved the use of liquid-phase reagents. The inventors have recognized and understood that phosphorus-containing layers can be formed using the gas-phase reagents of the present invention, and that these gas-phase reagents significantly reduce preparation time and chemical waste compared to previous liquid-phase surface modification processes. Furthermore, the use of such reagents minimizes liquid-related process steps, thereby advantageously improving the scalability of the overall process and the consistency and uniformity of the coating.

[0016] In some embodiments, the present application relates to the discovery of a stable surface coating that can enable a substrate, such as an array, biosensor, or other assay device, to withstand corrosive solutions by protecting the underlying material of the exposed surface. For example, certain biological assays require the use of corrosive solutions or other harsh conditions (e.g., high-salt solutions, multi-solution washing, etc.) that may corrode the material of the substrate or remove functional chemical portions from the substrate surface. The inventors have recognized and understood that certain gas-phase reagents can be used to form a highly stable surface coating on a substrate surface. Accordingly, in some embodiments, the present application provides a method for modifying a substrate surface that stabilizes the substrate under harsh assay conditions and / or over a long period of time.

[0017] In some embodiments, the present invention relates to the discovery that a phosphorus-containing layer on a metallic or plastic surface enhances the formation of a silane coating layer on the metallic or plastic surface. Prior techniques for surface modification have used liquid phosphonic acid derivatives to passivate metallic surfaces for silane functionalization, based on the previous assumption that the passivation layer would inhibit the silane treatment and promote the functionalization of non-metallic surfaces. Surprisingly, the inventors have found that a phosphorus-containing layer formed using a phosphonic acid derivative enhances the silane treatment on metallic surfaces. Based on these findings, the inventors have developed a technique involving the functionalization of a non-metallic surface using a gas-phase phosphonic acid derivative before forming a phosphorus-containing layer on the metallic or plastic surface. The phosphorus-containing layer then functions as a base layer for the downstream silane treatment to form a silane coating layer on the metallic or plastic surface.

[0018] In addition to the advantages described above, surface coatings can provide a more favorable interface for reagents in biological assays, such as antifouling surface coatings that reduce or eliminate the adhesion of reagent components in biological reactions. Examples of surface coatings and surface modification processes that may be used in accordance with this application are described in U.S. Patent Application Publication No. 20180326412, entitled "SUBSTRATES HAVING MODIFIED SURFACE REACTIVITY AND ANTIFOULING PROPERTIES IN BIOLOGICAL REACTIONS," which is incorporated herein by reference in its entirety.

[0019] In particular, the present application provides a method for forming a phosphorus-containing layer on a metallic surface or a plastic surface, among several embodiments. In some embodiments, the metallic surface may include a metal, a metal oxide (e.g., oxide anion, hydroxide), or a combination thereof. In some embodiments, the metallic surface includes one or more transition metals. In some embodiments, the metallic oxide surface is a transition metal (i.e., transition metal) oxide surface. In some embodiments, the metallic oxide surface may include titanium oxide (e.g., titanium dioxide), aluminum oxide, zirconium oxide, iron oxide, tin oxide, and / or tantalum oxide. In one embodiment, the metallic surface is a titanium dioxide (TiO2) surface. In some embodiments, the plastic surface may include polyethylene, polypropylene, polystyrene, polycarbonate, polyvinyl chloride, polytetrafluoroethylene, or a combination thereof.

[0020] In some embodiments, metallic or plastic surfaces may be pre-treated or cleaned before or at the beginning of the surface modification processes described herein. In some embodiments, the surface may be subjected to a plasma cleaning process before or at the beginning of the surface modification processes. For example, in some embodiments, the surface may be exposed to plasma, radicals, excited species and / or atomic species. In some embodiments, the surface may be exposed to hydrogen, ammonia and / or mixtures thereof and heat-treated. In some embodiments, the surface may be treated with oxygen plasma, tetrabutylammonium hydroxide, potassium hydroxide in methanol, hydrogen peroxide in sulfuric acid (e.g., "Piranha" solution), nitric acid in sulfuric acid, hydrogen peroxide in ammonia, sulfuric acid, hydrofluoric acid, EDTA, or a combination of these treatment agents before a phosphorus-containing layer is formed on the surface. In some embodiments, the pre-treated surface has exposed hydroxide anionic functional groups and / or oxide anionic functional groups.

[0021] In the context of this specification, in some embodiments, the phosphorus-containing layer refers to an aggregate of phosphorus-containing molecules adsorbed onto a surface from a gas-phase precursor (e.g., a phosphoryl halide in the gas phase). In some embodiments, the phosphorus-containing molecules in this layer include an organic tail and a terminal phosphorus-containing portion, and the molecules are adsorbed onto the surface through the phosphorus-containing portion such that the organic tail is located distal to the surface compared to the phosphorus-containing portion. Examples of phosphorus-containing portions include, but are not limited to, phosphonates or phosphonic acids, phosphonits, phosphates, phosphites, phosphonamidates, phosphoramidates, and other such organophosphorus functional groups known in the art. Therefore, in some embodiments, the phosphorus-containing layer is an organic organophosphorus layer, such as an organic phosphonate layer. In some embodiments, the phosphorus-containing layer is a self-assembled monolayer (SAM), which is a relatively ordered aggregate of molecules spontaneously adsorbed from a gas-phase precursor reagent onto a metallic or plastic surface.

[0022] According to the present invention, a phosphorus-containing layer (e.g., an organic layer) is formed on a metallic or plastic surface by treating the surface with a gas-phase reagent (e.g., a gas-phase organic reagent such as phosphoryl halide). In some embodiments, the present invention provides a method for forming a stable surface coating on one or more surface portions of a substrate. In some embodiments, the stable surface coating refers to a surface coating comprising a phosphorus-containing layer and a coating layer (e.g., a silane coating layer).

[0023] In some embodiments, the coating layer is formed on the metallic or plastic surface after the phosphorus-containing layer has been formed on the metallic or plastic surface. In some embodiments, a portion of the coating layer is formed on the metallic or plastic surface, and another portion of the coating layer is formed on the phosphorus-containing layer. In some embodiments, a portion of the coating layer is formed on a first surface portion of the substrate (e.g., on the metallic or plastic surface), and another portion of the coating layer is formed on a second surface portion of the substrate (e.g., on the transparent or glass surface). In some embodiments, the coating layer is formed by treating the surface with at least one silane in the gas phase. In some embodiments, the at least one silane is a chlorosilane such as a chlorosiloxane, alkylchlorosilane, or a mixture thereof. In some embodiments, the at least one silane is adsorbed onto the metallic or plastic surface. In some embodiments, the at least one silane is adsorbed onto the phosphorus-containing layer. In some embodiments, the at least one silane is adsorbed onto the transparent or glass surface.

[0024] As used herein, a gas-phase reagent or a reagent in the gas phase means a molecule capable of undergoing a gas-to-solid deposition reaction, or a gas-to-solid precursor containing such a molecule. In some embodiments, gas means gas, vapor, and / or aerosol. In some embodiments, gas means a gaseous-phase substance (e.g., gas or vapor), or a substance otherwise dispersed in a volume that can be occupied by a gas, vapor, or aerosol (e.g., droplets sprayed or ejected into a volume). Gas-to-solid deposition means a reaction in which a molecule or material in the gas phase forms one or more solid-state deposition products. For example, in some embodiments, the gas-to-solid deposition reaction is a decomposition reaction. A gas-to-solid decomposition reaction may produce at least one gaseous-state product and at least one solid-state product. A gaseous-state product means a molecule whose physical state is gas, vapor, or aerosol. A solid-state product means an atom, ion, compound, molecule, or combination thereof whose physical state is solid. The solid-phase deposition products of precursors from gas to solid may contribute to the formation of solid materials such as phosphorus-containing layers and / or coating layers as described herein.

[0025] Accordingly, in some embodiments, the formation of a phosphorus-containing layer and / or coating layer may involve the chemical and / or physical transformation of one or more reagents in the gas phase. For example, during surface treatment with a phosphoryl halide in the gas phase, the phosphoryl halide may decompose into a gas-phase product in the form of a halide gas and a solid-phase product in the form of a phosphoryl layer on the surface (e.g., the phosphorus-containing layer). In some embodiments, the phosphorus-containing layer is formed through covalent bonding of phosphoryl to the surface. For example, the phosphorus atoms of the phosphoryl may be bonded by hydroxide anionic functional groups and / or oxide anionic functional groups on the surface, thereby covalently bonding the phosphoryl through phosphonate groups. In some embodiments, the phosphorus-containing layer is formed on the surface without forming covalent bonds and is held in place by van der Waals, hydrogen bonds or dipole forces, for example.

[0026] In some embodiments, the gaseous reagent of the present application may be suitable for chemical vapor deposition (CVD), atomic layer deposition (ALD), molecular beam epitaxy (MBE), physical vapor deposition (PVD), or any combination of these deposition processes. The gaseous reagent may be a CVD precursor and / or an ALD precursor. CVD and ALD are non-limiting examples of gas-to-solid deposition processes for depositing solid materials from gaseous precursors. Examples of CVD processes include, but are not limited to, metal-organic CVD (MOCVD), plasma-enhanced CVD (PECVD), microwave plasma-assisted CVD (MWCVD or MPCVD), hot filament CVD (HFCVD), photoinitiated CVD (PICVD), laser CVD (LCVD), vapor phase epitaxy (VPE), and ALD. The deposition of the gaseous reagent may involve a reaction of the reagent with the substrate or its receiving surface. In some embodiments, the substrate is exposed to multiple types of gaseous reagents simultaneously and / or sequentially to deposit a solid material such as the phosphorus-containing layer and / or coating layer described herein.

[0027] In some aspects, the present application provides a method and a composition for modifying a surface. As used herein, in some embodiments, the surface refers to the surface of a substrate or a solid support. In some embodiments, the substrate refers to a material, layer, or other structure having a surface such as a receiving surface that can support a deposited material such as the layer or coating described herein. In some embodiments, the receiving surface of the substrate may optionally have one or more features including nano-scale or micro-scale concave features such as an array of sample wells. In some embodiments, an array refers to an arrangement of elements such as sensors or sample wells in a planar manner. The array can be one-dimensional or two-dimensional. A one-dimensional array has a single row or column of elements in the first dimension and multiple rows or columns in the second dimension. The number of rows or columns in the first and second dimensions may be the same or different. In some embodiments, the array may have, for example, 10 2 pieces, 10 3 pieces, 104 pieces, 10 5 pieces, 10 6 pieces, or 10 7 It may contain individual sample wells.

[0028] In one embodiment, the sample well of the array has an upper opening, the upper opening extends into the substrate to a bottom surface distal to the upper opening, and the sample well has a side wall surface positioned between the upper opening and the bottom surface. In one embodiment, the side wall surface has a first surface portion, and the bottom surface has a second surface portion, and the phosphoryl halide preferentially forms a phosphorus-containing layer on the side wall surface of the sample well.

[0029] In some embodiments, the present application provides a method for modifying one or more surfaces of a sample well. In some embodiments, the sample well comprises a restricting region which may be a physical or chemical property of a substrate that enables the localization of a molecule or reaction of interest. In some embodiments, the restricting region may be a distinct region of the substrate surface to which the molecule of interest binds, for example, the bottom surface of the sample well having a polypeptide or nucleic acid coupled to such surface. In some embodiments, the sample well comprises a cavity or well created within a substrate or device and having a defined shape and volume. The sample well can be prepared using the techniques described in the art, for example, as disclosed in U.S. Patent Application No. 16 / 555,902, the contents of which are incorporated herein by reference without limitation.

[0030] Figure 1A schematically shows an example of functionalization of a sample well surface according to an embodiment of the present invention. A cross-sectional view of a sample well 100 is shown, which has a metallic surface 102 and a silica surface 104. For illustrative purposes, the sample well 100 is shown to be defined by a side wall extending from the top to the bottom, with the metallic surface 102 formed on the side wall and the top surface, and the silica surface 104 formed on the bottom surface.

[0031] It should be understood that in some embodiments, any of the feature portions (sidewalls, top surface, bottom surface) defining the sample well 100 may have different or additional surface properties. For example, in some embodiments, the sample well 100 is defined by a sidewall extending into the bottom material such that a portion of the silica surface 104 is formed on the sidewall and another portion of the silica surface 104 is formed on the bottom surface. In this configuration, the sidewall of the sample well 100 would include a silica surface 104 formed on the surface portion proximal to the bottom surface and a metallic surface 102 formed on the surface portion distal to the bottom surface.

[0032] In process (I), the sample well 100 is treated with a functionalizing agent 108 that preferentially binds to the silica surface 104 to form a functionalized surface 106. The functionalizing agent 108 includes a coupling portion that provides coupling functionality to the bottom surface of the sample well 100. As shown, in some embodiments, the functionalized surface 106 comprises the functionalizing agent 108 and a silane 110 that does not include the coupling portion. Thus, in some embodiments, the functionalized surface 106 is formed in process (I) by treating the sample well 100 with a mixture containing the functionalizing agent 108 and the silane 110. In some embodiments, the functionalizing agent 108 is a biotinylated silane (e.g., biotin-PEG-silane) and the silane 110 is a non-biotylated silane (e.g., PEG-silane).

[0033] In some embodiments, the functionalizing agents described herein include coupling moieties. In some embodiments, the coupling moieties are covalent coupling moieties. Examples of covalent coupling moieties include trans-cyclooctene (TCO) moieties, tetrazine moieties, azide moieties, alkyne moieties, aldehyde moieties, isocyanate moieties, N-hydroxysuccinimide moieties, thiol moieties, alkene moieties, dibenzocyclooctyl moieties, bicyclononyne moieties, and thiamine pyrophosphate moieties. Examples of functionalizing agents containing a covalent coupling moiety include, but are not limited to, azido-silanes and azido-organosilanes such as azido-PEG-silanes (e.g., azido-PEG3-silane, azido-PEG5-silane) and azido-alkylsilanes (e.g., azido-C11-silane). In some embodiments, the coupling moiety is a non-covalent coupling moiety. Examples of non-covalent coupling moieties include, but are not limited to, biotin moieties, avidin proteins, streptavidin proteins, lectin proteins, and SNAP tags. In some embodiments, the functionalizing agent includes a moiety that can preferentially bind to silica.

[0034] In one embodiment, the portion that can preferentially bond to silica is a silane. In one embodiment, the silane is monoethoxysilane, methoxysilane, diethoxysilane, trichlorosilane, or diethoxymethoxysilane. In one embodiment, the functionalizing agent includes a biotinylated silane.

[0035] In some embodiments, the functionalizing agent 108 and silane 110 may be provided in a ratio determined based on the desired density of coupling portions on the surface to be functionalized. For example, in some embodiments, the functionalized surface is formed using a mixture containing the functionalizing agent and silane in a molar ratio of at least 5 times excess silane relative to the functionalizing agent. In some embodiments, the mixture contains silane in an excess of about 5 to about 250 times relative to the functionalizing agent (e.g., about 5 to 100 times, about 5 to about 50 times, about 50 to about 250 times, about 100 to about 250 times, or about 50 to about 150 times excess silane relative to the functionalizing agent).

[0036] In process (II), the sample well 100 is treated with a phosphoryl halide in the gas phase to form a phosphorus-containing layer 112. As shown, in some embodiments, the phosphoryl halide preferentially forms the phosphorus-containing layer 112 on the metallic surface 102. In some embodiments, the phosphoryl halide preferentially forms the phosphorus-containing layer on the metallic surface (e.g., the first surface portion of the sample well) with a selectivity of about 2 to about 60 times. In some embodiments, the phosphoryl halide preferentially forms the phosphorus-containing layer on the metallic surface with a selectivity of about 4 to about 40 times. In some embodiments, the phosphoryl halide preferentially forms the phosphorus-containing layer on the metallic surface with a selectivity of about 8 to about 20 times.

[0037] In process (III), the sample well 100 is treated with at least one silane to form a coating layer 114 on the metallic surface 102 and the silica surface 104. In some embodiments, the coating layer 114 is formed by treating the sample well 100 with a single composition containing one or more silanes (e.g., one silane or a mixture of several silanes). For example, in some embodiments, the coating layer 114 is formed by co-deposition of two or more silanes. In some embodiments, the coating layer 114 is formed by backfill silanization, as shown in Figure 1B.

[0038] Figure 1B schematically illustrates an example of forming a coating layer 114 by backfill silane treatment. In process (i), the sample well 100 is treated with a first silane to form a partial coating layer 116 on the metallic surface 102 and the silica surface 104. In process (ii), the sample well 100 is treated with a second silane to form a coating layer 114. In some embodiments, the first and second silanes are the same. In some embodiments, the first and second silanes are different (e.g., different types of silanes).

[0039] In some embodiments, the coating layers described herein can be formed on a surface by deposition, co-deposition, or backfill silane treatment by treating the surface with one or more silanes in the gas phase. Silanes that can be used for gas-phase treatment include any silane compound having a preferred boiling point or flash point (e.g., less than about 200°C) in vacuum. Examples of silanes in the gas phase that can be used to form a coating layer include halogenated silanes (e.g., chlorosilanes such as chlorosiloxane, alkylchlorosilane, alkyldichlorosilane, alkyltrichlorosilane), alkoxysilanes (e.g., monoalkoxysilane, dialkoxysilane, and trialkoxysilane), silicon hydrides (e.g., hexylsilane), as well as azasilanes including both linear and cyclic forms (e.g., hexamethyldisilazane), and thiasilanes including both linear and cyclic forms (e.g., 2,2,4-trimethyl-1-thia-2-silacyclopentane). Specific examples of chlorosilanes include, but are not limited to, hexachlorodisiloxane, chloro(hexyl)dimethylsilane, hexyldichlorosilane, and hexyltrichlorosilane. Specific examples of alkoxysilanes include, but are not limited to, methoxytrimethylsilane, dimethoxydimethylsilane, and hexyltrimethoxysilane.

[0040] As generally shown in Figures 1A and 1B, in some embodiments, a portion of the coating layer 114 is formed on the silica surface 104 such that the coupling portion of the functionalizing agent 108 remains available for coupling the molecule of interest to the functionalized surface 106. Figure 1C shows an example in which the molecule of interest is coupled to the functionalized surface of the substrate. In process (IV), the molecule of interest 118, bound to the coupling portion of the functionalizing agent 108, is brought into contact with the sample well 100. Examples of molecules of interest are described herein and include, but are not limited to, nucleic acids and polypeptides (e.g., proteins and protein fragments for use in polypeptide sequencing reactions, and polymerization enzymes such as nucleic acid polymerases for use in nucleic acid sequencing reactions).

[0041] Figure 2 is a cross-sectional view showing an example of a sample well 100 having surface modification according to the present application. As shown in Figure 2, the sample well 100 is defined by an opening formed through a metal stack 202 disposed on a cladding layer 204 (e.g., SiO2). In some embodiments, the sample well 100 is one of several sample wells formed on the surface of a device (e.g., a device having an array of sample wells). The metal stack 202 may include one or more layers of metallic material (e.g., aluminum, titanium, copper). As shown in Figure 2, some embodiments of the metal stack 202 include an aluminum layer 206 disposed in close proximity to the top of the cladding layer 204 and a titanium nitride layer 208 on top of the aluminum layer 206.

[0042] The aluminum layer 206 may contain copper and / or silicon. In some embodiments, the aluminum layer 206 may contain less than about 2% copper and / or silicon and may have a thickness in the range of about 30 nm to 150 nm or any value within that range. In some embodiments, the aluminum layer is about 65 nm thick. The titanium nitride layer 208 may include a layer of titanium in contact with the aluminum layer 206 and may have a thickness in the range of about 1 nm to 150 nm or any value within that range. In some embodiments, the thickness of the titanium nitride layer 208 is about 80 nm thick.

[0043] To facilitate selective chemical functionalization for coupling the target molecule at the bottom of the sample well 100, the bottom surface of the sample well 100 should have a different composition from other surfaces (e.g., the side walls of the sample well 100). As shown in Figure 2, the bottom surface of the sample well may be the cladding layer material (e.g., exposed SiO2), and the side walls of the sample well 100 may be the spacer material 210. The spacer material 210 may contain one or more metal oxides (e.g., TiO2, Al2O3, SiO2, TiN, HfO2, ZrO2, and Ta2O5). The top surface of the array may contain one or more metal oxide materials formed by oxidation of the top surface of layer 208 (e.g., TiO2 formed by oxidation of TiN).

[0044] In some embodiments, it may be desirable that the exposed surfaces of the array (top surface of layer 208, sample well sidewalls, bottom surface) be substantially stable to certain types of solutions, including those used during surface modification and assays. For example, certain conditions used in an assay may include corrosive solutions or, in other cases, harsh processing conditions (e.g., high ionic strength aqueous solutions, multiple solution cycling, high or low pH solutions, etc.). According to the present invention, a surface modification process can be used to produce an array having one or more layers and / or coatings on its exposed surfaces that can be substantially stable when in contact with such solutions over a desired period of time.

[0045] As shown in Figure 2, a sample well 100 produced by the surface modification technique of the present invention may include a functionalizing agent 108 having a coupling portion 212 on the exposed surface of the cladding layer 204 defining the bottom surface of the sample well 100. The sample well 100 comprises a phosphorus-containing layer 112 formed on the upper surface of layer 208 and on the spacer material 210 (e.g., on the metallic surface of the sample well 100). As described herein, the phosphorus-containing layer 112 may be formed by treating the sample well 100 with a phosphoryl halide in the gas phase, which preferentially forms the phosphorus-containing layer 112 on the metallic surface of the sample well 100. As shown, the sample well 100 comprises a coating layer 114 formed on the exposed surface of the cladding layer 204 (e.g., the silica surface), as well as on the upper surface of layer 208 and on the spacer material 210 (e.g., the metallic surface). In some embodiments, the coating layer 114 is a silane coating layer. In some embodiments, the stable surface coating of the present invention comprises a phosphorus-containing layer 112 and a coating layer 114.

[0046] Phospholyl halide The method described herein uses phosphoryl halide as the gas-phase reagent. "Phosphoryl halide" is >P(=O)(X 1 An organic or inorganic compound containing a group, where X 1 is a halogen. Phosphoryl halides are phosphoryl monohalides, phosphoryl dihalides, or phosphoryl trihalides, as described below. In some embodiments, the phosphoryl halide is an organic phosphoryl halide. The term "organic" means that the reagent contains one or more carbon atoms (i.e., at least one carbon-containing group). In some embodiments, the phosphoryl halide is an organic phosphoryl halide, and the phosphorus-containing layer is an organic phosphorus layer.

[0047] The phosphoryl halide should have a molecular weight that aids in its use in the gas phase. In certain embodiments, the phosphoryl halide has a molecular weight of less than 500, 450, 400, 350, 300, 250, 200 or 150 g / mol. In certain embodiments, the phosphoryl halide has a molecular weight of about 132 g / mol to 260 g / mol. In certain embodiments, the phosphoryl halide has a molecular weight of about 132 g / mol to 232 g / mol. In certain embodiments, the phosphoryl halide has a molecular weight of about 203 g / mol to 232 g / mol.

[0048] Phosphoryl dihalide In certain embodiments, the phosphoryl halide is a phosphoryl dihalide (i.e., it has a -P(=O)(X 1 )2 group, where each instance of X 1 is independently a halogen). In certain embodiments, the phosphoryl dihalide is phosphoryl dichloride. In certain embodiments, the phosphoryl dihalide is an organic phosphoryl dihalide. In certain embodiments, the phosphoryl dihalide is an organic phosphoryl dichloride.

[0049] In certain embodiments, the phosphoryl dihalide is an organic phosphonic acid dihalide (-P(=O)(X 1 )2 is bonded to carbon). In certain embodiments, the phosphoryl dihalide is an organic phosphonic acid dichloride.

[0050] For example, in certain embodiments, the phosphoryl dihalide is of formula (I):

[0051]

Chemical formula

[0052] or a salt thereof, where each instance of X 1 is independently a halogen, and each instance of R 1This includes optionally substituted alkyl, optionally substituted heteroalkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted carbocyclyl, optionally substituted heterocyclyl, optionally substituted aryl, or optionally substituted heteroaryl, -OR O , or -N(R N )2, R O Each example independently consists of hydrogen, an optionally substituted alkyl, an optionally substituted alkenyl, an optionally substituted alkynyl, an optionally substituted carbocyclyl, an optionally substituted heterocyclyl, an optionally substituted aryl, an optionally substituted heteroaryl, an optionally substituted acyl, or an oxygen protecting group, or optionally two R O However, together with the intervening atom, it forms a heterocycline in which substitutions are made at will. R N Each example independently is either hydrogen, optionally substituted alkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted carbocyclyl, optionally substituted heterocyclyl, optionally substituted aryl, optionally substituted heteroaryl, optionally substituted acyl, or nitrogen protecting group, or optionally two R N However, together with the intervening atom, it forms a heterocycline in which substitutions are made at will.

[0053] As described in this specification, X 1 Each of these examples is independently a halogen. In one embodiment, X 1 Each example is independently -Cl or -Br. In one embodiment, X 1 Each example is -Cl. In one embodiment, X 1 Each example is -Specific.

[0054] In one embodiment, the compound of formula (I) is:

[0055] [ka]

[0056] It belongs to them. In one embodiment, R 1 R is an optionally substituted alkyl group. In one embodiment, R 1 C is replaced by an optional substitution. 1~10 It is alkyl. In one embodiment, R 1 C is replaced by an optional substitution. 1~8 It is alkyl. In one embodiment, R 1 is a non-substituted C 1~8 It is alkyl. In one embodiment, R 1 C is replaced by an optional substitution. 6~8 It is alkyl. In one embodiment, R 1 is a non-substituted C 6~8 It is alkyl. In one embodiment, R 1 R is an optionally substituted C6 alkyl. In one embodiment, R 1 is an unsubstituted C6 alkyl. In one embodiment, R 1 R is an optionally substituted C8 alkyl. In one embodiment, R 1 It is an unsubstituted C8 alkyl group.

[0057] In one embodiment, R 1 The compound is selected from the group consisting of methyl, ethyl, n-propyl, iso-propyl, n-butyl, iso-butyl, sec-butyl, or tert-butyl.

[0058] In one embodiment, R 1 teeth,

[0059] [ka]

[0060] It is selected from the group consisting of the following. For example, in one embodiment, the phosphoryl halide (e.g., a compound of formula (I)) is one of the compounds of the following formulas.

[0061]

Chem.

[0062] For example, in certain embodiments, the phosphoryl halide is

[0063]

Chem.

[0064] selected from the group consisting of. In certain embodiments, R 1 is -OR O In certain embodiments, R 1 is -OR O and R O is optionally substituted aryl. In certain embodiments, R 1 is -OR O and R O is optionally substituted phenyl. In certain embodiments, R 1 is -OR O and R O is unsubstituted phenyl. In certain embodiments, R 1 is -OR O and R O is phenyl substituted with halogen or -NO2.

[0065] For example, in certain embodiments, the phosphoryl halide is

[0066]

Chem.

[0067] selected from the group consisting of. Other examples of phosphoryl halides (i.e., compounds of formula (I)) include the following:

[0068]

Chem.

[0069] These are some examples, but are not limited to them. In one embodiment, R 1 is a polymer group (e.g., polyethylene glycol (PEG)). In one embodiment, R 1 It is a polyfluoroalkyl group.

[0070] In one embodiment, a phosphoryl halide (i.e., a compound of formula (I)) is formula (Ia):

[0071] [ka]

[0072] It is a substance or a salt thereof, where L 1 is an alkylene that has been optionally substituted. In one embodiment, a compound of formula (Ia) is:

[0073] [ka]

[0074] It belongs to them. In one embodiment, L 1 C is replaced by an optional substitution. 1~6 It is an alkylene. 1 is a non-substituted C 1~6 It is an alkylene. 1 This is one of the following formulas.

[0075] [ka]

[0076] For example, in one embodiment, a phosphoryl halide (i.e., a compound of formula (Ia)) is as follows:

[0077] [ka]

[0078] That is the case. Phosphoryl monohalide In one embodiment, phosphoryl halide is phosphoryl monohalide (i.e., >P(=O)(X 1 ) equipped with a base, where X 1 (is a halogen). In one embodiment, phosphoryl monohalide is a phosphoryl monolide. In one embodiment, phosphoryl monohalide is an organic phosphoryl monohalide. In one embodiment, phosphoryl monohalide is an organic phosphoryl monolide.

[0079] In one embodiment, phosphoryl monohalide is an organophosphinic acid dihalide (where >P(=O)(X 1 (2 is bonded to two different carbon groups). In one embodiment, the phosphoryl monohalide is an organophosphinic acid chloride.

[0080] In one embodiment, the phosphoryl halide is of formula (II):

[0081] [ka]

[0082] It is the substance or its salt, where, X 1 It is a halogen, R 2 Each example is an optionally substituted alkyl, optionally substituted heteroalkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted carbocyclyl, optionally substituted heterocyclyl, optionally substituted aryl, or optionally substituted heteroaryl, -OR O , or -N(RN )2 or Alternatively, you can choose two R's. 2 The group, together with the intervening atom, forms a heterocycline in which substitution is optional. R O Each example independently consists of hydrogen, an optionally substituted alkyl, an optionally substituted alkenyl, an optionally substituted alkynyl, an optionally substituted carbocyclyl, an optionally substituted heterocyclyl, an optionally substituted aryl, an optionally substituted heteroaryl, an optionally substituted acyl, or an oxygen protecting group, or optionally two R O However, together with the intervening atom, it forms a heterocycline in which substitutions are made at will. R N Each example independently is either hydrogen, optionally substituted alkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted carbocyclyl, optionally substituted heterocyclyl, optionally substituted aryl, optionally substituted heteroaryl, optionally substituted acyl, or nitrogen protecting group, or optionally two R N However, together with the intervening atom, it forms a heterocycline in which substitutions are made at will.

[0083] In one embodiment, X 1 is -Cl or -Br. In one embodiment, X is -Cl. In one embodiment, the phosphoryl halide is of the formula:

[0084] [ka]

[0085] It belongs to them. In one embodiment, R 2 Each example is an optionally substituted alkyl group independently. In one embodiment, R 2 Each example is independently replaced with C by choice. 1~10 It is alkyl. In one embodiment, R 2Each example of is, independently, optionally substituted C 1~6 alkyl. In certain embodiments, R 2 Each example of is, independently, unsubstituted C 1~6 alkyl.

[0086] In certain embodiments, R 2 Each example of is independently selected from the group consisting of methyl, ethyl, n-propyl, iso-propyl, n-butyl, iso-butyl, sec-butyl or tert-butyl.

[0087] In certain embodiments, R 2 Each example of is

[0088]

Chemical formula

[0089] independently selected from the group consisting of. For example, in certain embodiments, the phosphoryl halide (i.e., the compound of formula (II)) is as follows.

[0090]

Chemical formula

[0091] In certain embodiments, R 2 Each example of is, independently, -OR O and each example of R O is, independently, optionally substituted alkyl. In certain embodiments, R 2 Each example of is, independently, -OR O and each example of R O is, independently, optionally substituted C 1~10 alkyl. In certain embodiments, R 2 Each example of is, independently, -OR O and each example of R O is, independently, optionally substituted C 1~6 alkyl. In certain embodiments, R 2Each example is independent of -OR O And R O Each example is an independent, non-substitutive C 1~6 It is alkyl.

[0092] For example, in one embodiment, the phosphoryl halide (i.e., the compound of formula (II)) is one of the following:

[0093] [ka]

[0094] Phosphoryl trihalide In one embodiment, phosphoryl halide is phosphoryl trihalide (i.e., formula P(=O)(X 1 )3, where X 1 Each of these examples is independently a halogen. In one embodiment, the phosphoryl halide is phosphoryl trichloride. In another embodiment, the phosphoryl halide is phosphoryl tribromide.

[0095] definition chemical definition The specific functional groups and chemical terms are defined in more detail below. Chemical elements are defined according to the periodic table, CAS version, and the lid of the 75th edition of the Handbook of Chemistry and Physics. They are identified as such, and the specific functional groups are generally defined as described therein. In addition, the general principles of organic chemistry, as well as the specific functional parts and reactivity, are described in "Organic Chemistry "Thomas Sorrell, University Science Books, Sausalito, 1999," "Smith and March, March's Advanced Organic Chemistry, 5th edition, John Wiley & Sons, Inc., New York, 2001," "Larock, Comprehensive Organic Transformations, VCHPublishers, Inc., New York This is described in "York, 1989" and "Carruthers, Some Modern Methods of Organic Synthesis, 3rd edition, Cambridge University Press, Cambridge, 1987."

[0096] Unless otherwise specified, the structures shown herein also mean that the compounds differ only in the presence of one or more isotopically enriched atoms. For example, hydrogen substitution with deuterium or tritium, 18 F 19 Substitution of F, or 13 C or 14 By C 12 Compounds having the present structure, excluding the substitution of C, are within the scope of this disclosure. Such compounds are useful, for example, as analytical tools or probes in biological assays.

[0097] When a range of values ​​is listed, it is intended to encompass each value and sub-range within that range. For example, "C 1~6 "Alkyl" refers to C1, C2, C3, C4, C5, C6, C 1~6 , C 1~5 , C 1~4 , C 1~3 , C 1~2 , C 2~6 , C 2~5 , C 2~4 , C 2~3 , C 3~6 , C 3~5 , C 3~4 , C 4~6 , C4~5 and C 5~6 It is intended to include alkyl groups.

[0098] The term "aliphatic" refers to alkyl groups, alkenyl groups, alkynyl groups, and carbocyclic groups. Similarly, the term "heteroaliphatic" refers to heteroalkyl groups, heteroalkenyl groups, heteroalkynyl groups, and heterocyclic groups.

[0099] The term "alkyl" refers to a radical of a linear or branched saturated hydrocarbon group having 1 to 10 carbon atoms ("C"). 1~10 This refers to an alkyl group having 1 to 9 carbon atoms. In some embodiments, the alkyl group has 1 to 9 carbon atoms ("C"). 1~9 (alkyl). In some embodiments, the alkyl group has 1 to 8 carbon atoms ("C"). 1~8 Alkyl). In some embodiments, the alkyl group has 1 to 7 carbon atoms ("C"). 1~7 Alkyl). In some embodiments, the alkyl group has 1 to 6 carbon atoms ("C"). 1~6 Alkyl). In some embodiments, the alkyl group has 1 to 5 carbon atoms ("C"). 1~5 Alkyl). In some embodiments, the alkyl group has 1 to 4 carbon atoms ("C"). 1~4 Alkyl). In some embodiments, the alkyl group has 1 to 3 carbon atoms ("C"). 1~3 Alkyl). In some embodiments, the alkyl group has 1 to 2 carbon atoms ("C"). 1~2 Alkyl). In some embodiments, the alkyl group has one carbon atom ("C1 alkyl"). In some embodiments, the alkyl group has two to six carbon atoms ("C1 alkyl"). 2~6 Alkyl). C 1~6Examples of alkyl groups include methyl (C1), ethyl (C2), propyl (C3) (e.g., n-propyl, isopropyl), butyl (C4) (e.g., n-butyl, tert-butyl group, sec-butyl, iso-butyl), pentyl (C5) (e.g., n-pentyl, 3-pentanyl, amyl, neopentyl, 3-methyl-2-butanyl, tert-amyl), and hexyl (C6) (e.g., n-hexyl). Additional examples of alkyl groups include n-heptyl (C7) and n-octyl (C8). Unless otherwise specified, each example of an alkyl group is independently either unsubstituted ("unsubstituted alkyl") or substituted with one or more substituents (e.g., halogens such as F) ("substituted alkyl"). In some embodiments, the alkyl group is unsubstituted C 1~10 Alkyl (unsubstituted C) 1~6 Alkyl groups include, for example, -CH3(Me), unsubstituted ethyl (Et), unsubstituted propyl (Pr, e.g., unsubstituted n-propyl (n-Pr), unsubstituted isopropyl (i-Pr)), and unsubstituted butyl (Bu, e.g., unsubstituted n-butyl (n-Bu), unsubstituted tert-butyl (tert-Bu or t-Bu), unsubstituted sec-butyl (sec-Bu), and unsubstituted isobutyl (i-Bu)). In one embodiment, the alkyl group is substituted C 1~10 Alkyl (substituted C) 1~6 Examples include alkyl groups, such as -CF3 (Bn).

[0100] The term "haloalkyl" refers to a substituted alkyl group in which one or more hydrogen atoms are independently substituted by a halogen, such as fluoro, bromo, chloro, or iodine. In some embodiments, the haloalkyl moiety has 1 to 8 carbon atoms ("C"). 1~8 ("Haloalkyl"). In some embodiments, the haloalkyl portion has 1 to 6 carbon atoms ("C"). 1~6 ("Haloalkyl"). In some embodiments, the haloalkyl portion has 1 to 4 carbon atoms ("C"). 1~4 ("Haloalkyl"). In some embodiments, the haloalkyl portion has 1 to 3 carbon atoms ("C"). 1~3("Haloalkyl"). In some embodiments, the haloalkyl portion has 1 to 2 carbon atoms ("C"). 1~2 Haloalkyl groups are a type of alkyl group. Examples of haloalkyl groups include -CHF2, -CH2F, -CF3, -CH2CF3, -CF2CF3, -CF2CF2CF3, -CCl3, -CFCl2, and -CF2Cl.

[0101] "Hyperalkyl" refers to an alkyl group further comprising at least one heteroatom (e.g., 1, 2, 3, or 4 heteroatoms) selected from oxygen, nitrogen, or sulfur, located within the parent chain (i.e., inserted between adjacent carbon atoms of the parent chain) and / or at one or more terminal positions of the parent chain. In some embodiments, a heteroalkyl group is a saturated group having 1 to 10 carbon atoms and one or more heteroatoms in the parent chain ("heteroc"). 1~10 This refers to an alkyl group. In some embodiments, a heteroalkyl group is a saturated group having 1 to 9 carbon atoms and 1 or more heteroatoms in the parent chain (heteroC 1~9 This refers to an alkyl group. In some embodiments, a heteroalkyl group is a saturated group having 1 to 8 carbon atoms and 1 or more heteroatoms in the parent chain (heteroC 1~8 This refers to an alkyl group. In some embodiments, a heteroalkyl group is a saturated group having 1 to 7 carbon atoms and 1 or more heteroatoms in the parent chain (heteroC 1~7 This refers to an alkyl group. In some embodiments, a heteroalkyl group is a saturated group having 1 to 6 carbon atoms and 1 or more heteroatoms in the parent chain (heteroC 1~6 "Alkyl" refers to a saturated group having 1 to 5 carbon atoms and 1 or 2 heteroatoms in the parent chain ("heteroC"). In some embodiments, a heteroalkyl group is a saturated group having 1 to 5 carbon atoms and 1 or 2 heteroatoms in the parent chain ("heteroC"). 1~5 "Alkyl" refers to a saturated group having 1 to 4 carbon atoms and 1 or 2 heteroatoms in the parent chain ("heteroC"). In some embodiments, a heteroalkyl group is a saturated group having 1 to 4 carbon atoms and 1 or 2 heteroatoms in the parent chain ("heteroC"). 1~4 This refers to an alkyl group. In some embodiments, a heteroalkyl group is a saturated group having 1 to 3 carbon atoms and 1 heteroatom in the parent chain (heteroC 1~3This refers to an alkyl group. In some embodiments, a heteroalkyl group is a saturated group having 1-2 carbon atoms and 1 heteroatom in the parent chain (heteroC 1~2 "Alkyl" refers to a saturated group having one carbon atom and one heteroatom in the parent chain ("heteroC1 alkyl"). In some embodiments, a heteroalkyl group has two to six carbon atoms and one or two heteroatoms in the parent chain ("heteroC1 alkyl"). 2~6 This refers to "alkyl". Unless otherwise specified, each example of a heteroalkyl group is either unsubstituted ("unsubstituted heteroalkyl") or substituted with one or more substituents ("substituted heteroalkyl"). In some embodiments, the heteroalkyl group is an unsubstituted hetero-C 1~10 It is alkyl. In one embodiment, the heteroalkyl group is a substituted heteroC 1~10 It is alkyl.

[0102] The term "alkenyl" refers to a linear or branched hydrocarbon group radical having 2 to 10 carbon atoms and one or more carbon-carbon double bonds (e.g., one, two, three, or four double bonds). In some embodiments, the alkenyl group has 2 to 9 carbon atoms ("C"). 2~9 ("Alkenyl"). In some embodiments, the alkenyl group has 2 to 8 carbon atoms ("C"). 2~8 ("Alkenyl"). In some embodiments, the alkenyl group has 2 to 7 carbon atoms ("C"). 2~7 ("Alkenyl"). In some embodiments, the alkenyl group has 2 to 6 carbon atoms ("C"). 2~6 ("Alkenyl"). In some embodiments, the alkenyl group has 2 to 5 carbon atoms ("C"). 2~5 ("Alkenyl"). In some embodiments, the alkenyl group has 2 to 4 carbon atoms ("C"). 2~4 ("Alkenyl"). In some embodiments, the alkenyl group has 2-3 carbon atoms ("C"). 2~3"Alkenyl"). In some embodiments, the alkenyl group has two carbon atoms ("C2 alkenyl"). One or more carbon-carbon double bonds may be internal (e.g., 2-butenyl) or terminal (e.g., 1-butenyl). C 2~4 Examples of alkenyl groups include ethenyl (C2), 1-propenyl (C3), 2-propenyl (C3), 1-butenyl (C4), 2-butenyl (C4), and butadienyl (C4). 2~6 An example of an alkenyl group is the aforementioned C 2~4 Examples include alkenyl groups, as well as pentenyl (C5), pentadienyl (C5), hexenyl (C6), and the like. Additional examples of alkenyls include heptenyl (C7), octenyl (C8), and octatrienyl (C8). Unless otherwise specified, each example of an alkenyl group is either unsubstituted ("unsubstituted alkenyl") or substituted with one or more substituents ("substituted alkenyl"). In some embodiments, the alkenyl group is unsubstituted C 2~10 It is an alkenyl. In one embodiment, the alkenyl group is substituted with C 2~10 It is an alkenyl. In the alkenyl group, the stereochemistry is not defined by the C=C double bond (for example, -CH=CHCH3 or

[0103] [ka]

[0104] ) can be an (E)- or (Z)- double bond. The term "alkynyl" refers to a radical of a linear or branched hydrocarbon group having 2 to 10 carbon atoms and one or more carbon-carbon triple bonds (e.g., one, two, three, or four triple bonds). 2~10 This refers to an "alkynyl" group. In some embodiments, the alkynyl group has 2 to 9 carbon atoms ("C"). 2~9 ("Alkynyl"). In some embodiments, the alkynyl group has 2 to 8 carbon atoms ("C"). 2~8("Alkynyl"). In some embodiments, the alkynyl group has 2 to 7 carbon atoms ("C"). 2~7 ("Alkynyl"). In some embodiments, the alkynyl group has 2 to 6 carbon atoms ("C"). 2~6 ("Alkynyl"). In some embodiments, the alkynyl group has 2 to 5 carbon atoms ("C"). 2~5 ("Alkynyl"). In some embodiments, the alkynyl group has 2 to 4 carbon atoms ("C"). 2~4 ("Alkynyl"). In some embodiments, the alkynyl group has 2-3 carbon atoms ("C"). 2~3 "Alkynyl"). In some embodiments, the alkynyl group has two carbon atoms ("C2 alkynyl"). One or more carbon-carbon triple bonds may be internal (e.g., 2-butynyl) or terminal (e.g., 1-butynyl). C 2~4 Examples of alkynyl groups include, but are not limited to, ethynyl (C2), 1-propynyl (C3), 2-propynyl (C3), 1-butynyl (C4), and 2-butynyl (C4). 2~6 An example of an alkenyl group is the aforementioned C 2~4 Examples include alkynyl groups, as well as pentynyl (C5), hexynyl (C6), and the like. Additional examples of alkynyls include heptynyl (C7) and octinyl (C8). Unless otherwise specified, each example of an alkynyl group is independently either unsubstituted ("unsubstituted alkynyl") or substituted with one or more substituents ("substituted alkynyl"). In some embodiments, the alkynyl group is an unsubstituted C 2~10 It is an alkynyl group. 2~10 It is alkinyl.

[0105] The term "carbocyrill" or "carbocyclic" refers to a non-aromatic ring system having 3 to 14 ring carbon atoms ("C"). 3~14 A "carbocyclyl" group refers to a radical of a non-aromatic cyclic hydrocarbon group in which there are zero heteroatoms in the non-aromatic ring system. In some embodiments, the carbocyclyl group has 3 to 10 ring carbon atoms ("C").3~10 (Carbocyclyl). In some embodiments, the carbocyclyl group has 3 to 8 ring carbon atoms ("C"). 3~8 (Carbocyclyl). In some embodiments, the carbocyclyl group has 3 to 7 ring carbon atoms ("C3-7 carbocyclyl"). In some embodiments, the carbocyclyl group has 3 to 6 ring carbon atoms ("C 3~6 Carbocyclyl). In some embodiments, the carbocyclyl group has 4 to 6 ring carbon atoms ("C"). 4~6 Carbocyclyl). In some embodiments, the carbocyclyl group has 5-6 ring carbon atoms ("C"). 5~6 Carbocyclyl). In some embodiments, the carbocyclyl group has 5 to 10 ring carbon atoms ("C"). 5~10 Carbocyclyl). Exemplary C 3~6 Carbocyclyl groups include, but are not limited to, cyclopropyl (C3), cyclopropenyl (C3), cyclobutyl (C4), cyclobutenyl (C4), cyclopentyl (C5), cyclopentenyl (C5), cyclohexyl (C6), cyclohexenyl (C6), and cyclohexadienyl (C6). (Example C) 3~8 The carbocyric group is the aforementioned C 3~6 This includes, but is not limited to, carbocyclyl groups, as well as cycloheptyl (C7), cycloheptenyl (C7), cycloheptadienyl (C7), cycloheptatrienyl (C7), cyclooctyl (C8), cyclooctenyl (C8), bicyclo[2.2.1]heptanyl (C7), and bicyclo[2.2.2]octanyl (C8). Exemplary C 3~10 The carbocyric group is the aforementioned C 3~8 Carbocyclyl group, as well as cyclononyl (C9), cyclononenyl (C9), cyclodecyl (C9) 10 ), cyclodecenyl (C 10 ), octahydro-1H-indenyl (C9), decahydronaphthalenyl (C9) 10 ), spiro[4.5]decanil(C 10This includes, but is not limited to, the following. As the examples above show, in some embodiments, the carbocyclyl group is either monocyclic ("monocyclic carbocyclyl") or polycyclic (including fused ring systems such as bicyclic ("bicyclic carbocyclyl") or tricyclic ("tricyclic carbocyclyl") systems, bridging ring systems or spiro-ring systems), and may be saturated or contain one or more carbon-carbon double or triple bonds. "Carbocyclyl" also includes ring systems in which the carbocyclyl ring as defined above is fused with one or more aryl or heteroaryl groups, and the bond site is on the carbocyclyl ring, in which case the number of carbons still indicates the number of carbons in the carbocyclic system. Unless otherwise specified, each example of a carbocyclyl group is independently either unsubstituted ("unsubstituted carbocyclyl") or substituted with one or more substituents ("substituted carbocyclyl"). In one embodiment, the carbocyclyl group is an unsubstituted C 3~14 It is a carbocyclyl. In one embodiment, the carbocyclyl group is substituted with C 3~14 It is carbocyclyl.

[0106] In some embodiments, "carbocykrill" is a monocyclic saturated carbocykrill group having 3 to 14 ring carbon atoms ("C 3~14 It is a cycloalkyl group. In some embodiments, the cycloalkyl group has 3 to 10 ring carbon atoms ("C"). 3~10 ("Cycloalkyl"). In some embodiments, the cycloalkyl group has 3 to 8 ring carbon atoms ("C"). 3~8 ("Cycloalkyl"). In some embodiments, the cycloalkyl group has 3 to 6 ring carbon atoms ("C"). 3~6 ("Cycloalkyl"). In some embodiments, the cycloalkyl group has 4 to 6 ring carbon atoms ("C"). 4~6 ("Cycloalkyl"). In some embodiments, the cycloalkyl group has 5-6 ring carbon atoms ("C"). 5~6 ("Cycloalkyl"). In some embodiments, the cycloalkyl group has 5 to 10 ring carbon atoms ("C"). 5~10 Cycloalkyl). C5~6 Examples of cycloalkyl groups include cyclopentyl (C5) and cyclohexyl (C5). 3~6 Examples of cycloalkyl groups include, as mentioned earlier, C 5~6 Examples include cycloalkyl groups, as well as cyclopropyl (C3) and cyclobutyl (C4). 3~8 Examples of cycloalkyl groups include, as mentioned earlier, C 3~6 Examples include cycloalkyl groups, as well as cycloheptyl (C7) and cyclooctyl (C8). Unless otherwise specified, each example of a cycloalkyl group is independently either unsubstituted ("unsubstituted cycloalkyl") or substituted with one or more substituents ("substituted cycloalkyl"). In some embodiments, the cycloalkyl group is unsubstituted C 3~14 It is a cycloalkyl group. In one embodiment, the cycloalkyl group is an unsubstituted C 3~14 It is a cycloalkyl group.

[0107] The term “heterocyclyl” or “heterocyclic” refers to a radical of a 3- to 14-membered non-aromatic ring system having a ring carbon atom and 1 to 4 ring heteroatoms, where each heteroatom is independently selected from nitrogen, oxygen, and sulfur (“3- to 14-membered heterocyclyl”). In heterocyclyl groups containing one or more nitrogen atoms, the bond sites can be carbon atoms or nitrogen atoms, as long as the valence allows. Heterocyclyl groups may be monocyclic (“monocyclic heterocyclyl”) or polycyclic (including fused ring systems such as bicyclic (“bicyclic heterocyclyl”) or tricyclic (“tricyclic heterocyclyl”), bridging ring systems, or spirocyclic systems), may be saturated, or may contain one or more carbon-carbon double or triple bonds. Heterocyclyl polycyclic ring systems may contain one or more heteroatoms in one or both rings. "Heterocyclyl" also includes a ring system in which the heterocyclyl ring defined above is fused with one or more carbocyrill groups, with the bond site located either on the carbocyrill ring or the heterocyclyl ring, or a ring system in which the heterocyclyl ring defined above is fused with one or more aryl or heteroaryl groups, with the bond site located on the heterocyclyl ring, in which case the number of ring members still indicates the number of ring members in the heterocyclyl ring system. Unless otherwise specified, each example of a heterocyclyl group is independently either unsubstituted ("unsubstituted heterocyclyl") or substituted with one or more substituents ("substituted heterocyclyl"). In some embodiments, the heterocyclyl group is an unsubstituted 3- to 14-membered heterocyclyl. In some embodiments, the heterocyclyl group is a substituted 3- to 14-membered heterocyclyl.

[0108] In some embodiments, the heterocyclyl group is a 5-10 member non-aromatic ring system having a ring carbon atom and 1-4 ring heteroatoms, where each heteroatom is independently selected from nitrogen, oxygen, and sulfur ("5-10 member heterocyclyl"). In some embodiments, the heterocyclyl group is a 5-8 member non-aromatic ring system having a ring carbon atom and 1-4 ring heteroatoms, where each heteroatom is independently selected from nitrogen, oxygen, and sulfur ("5-8 member heterocyclyl"). In some embodiments, the heterocyclyl group is a 5-6 member non-aromatic ring system having a ring carbon atom and 1-4 ring heteroatoms, where each heteroatom is independently selected from nitrogen, oxygen, and sulfur ("5-6 member heterocyclyl"). In some embodiments, the 5-6 member heterocyclyl has 1-3 ring heteroatoms selected from nitrogen, oxygen, and sulfur. In some embodiments, the 5-6 member heterocyclyl has 1-2 ring heteroatoms selected from nitrogen, oxygen, and sulfur. In some embodiments, the 5-6 membered heterocyclil has one ring heteroatom selected from nitrogen, oxygen, and sulfur.

[0109] The term "aryl" refers to aromatic ring systems ("C 6~14 The term "aryl" refers to a monocyclic or polycyclic (e.g., bicyclic or tricyclic) 4n+2 aromatic ring system (e.g., having 6, 10, or 14 π electrons shared in a cyclic array) radical having 6 to 14 membered ring carbon atoms provided within it, and zero heteroatoms provided within the aromatic ring system. In some embodiments, the aryl group has 6 ring carbon atoms ("C6 aryl", e.g., phenyl). In some embodiments, the aryl group has 10 ring carbon atoms ("C6 aryl"). 10 "Aryl" (e.g., naphthyl such as 1-naphthyl and 2-naphthyl). In some embodiments, the aryl group has 14 ring carbon atoms ("C"). 14"Aryl" (e.g., anthracyl). "Aryl" also includes ring systems in which the aryl ring defined above is condensed with one or more carbocyrillic or heterocyclyl groups, and the bond radical or bond site is on the aryl ring, in which case the number of carbon atoms still indicates the number of carbon atoms in the aryl ring system. Unless otherwise specified, each example of an aryl group is independently either unsubstituted ("unsubstituted aryl") or substituted with one or more substituents ("substituted aryl"). In some embodiments, the aryl group is unsubstituted C 6~14 It is aryl. In one embodiment, the aryl group is substituted C 6~14 It is Ariel.

[0110] The term "heteroaryl" refers to a 5- to 14-membered monocyclic or polycyclic (e.g., bicyclic, tricyclic) 4n+2 aromatic ring system (e.g., having 6, 10, or 14 π electrons shared in a cyclic array) radical having a ring carbon atom and 1-4 ring heteroatoms provided within the aromatic ring system, where each heteroatom is independently selected from nitrogen, oxygen, and sulfur ("5- to 14-membered heteroaryl"). In heteroaryl groups containing one or more nitrogen atoms, the bond site can be a carbon atom or a nitrogen atom, as long as the valence allows. Heteroaryl polycyclic ring systems can contain one or more heteroatoms in one or both rings. "Heteroaryl" includes ring systems in which the heteroaryl ring as defined above is fused with one or more carbocyryl groups or heterocyclyl groups, and the bond site lies on the heteroaryl ring, in which case the number of ring members still indicates the number of ring members in the heteroaryl ring system. "Hyperaryl" also includes ring systems in which the heteroaryl ring defined above is fused with one or more aryl groups, and the bond site is on either the aryl ring or the heteroaryl ring, in which case the number of ring members indicates the number of ring members in the fused polycyclic (aryl / heteroaryl) ring system. In polycyclic heteroaryl groups in which one ring does not contain a heteroatom (e.g., indolyl, quinolinyl, carbazolyl), the bond site can be on either ring, i.e., on the ring containing a heteroatom (e.g., 2-indolyl) or on the ring without a heteroatom (e.g., 5-indolyl).

[0111] In some embodiments, the heteroaryl group is a 5-10 member aromatic ring system having a ring carbon atom provided to the aromatic ring system and 1-4 ring heteroatoms, where each heteroatom is independently selected from nitrogen, oxygen, and sulfur ("5-10 member heteroaryl"). In some embodiments, the heteroaryl group is a 5-8 member aromatic ring system having a ring carbon atom provided to the aromatic ring system and 1-4 ring heteroatoms, where each heteroatom is independently selected from nitrogen, oxygen, and sulfur ("5-8 member heteroaryl"). In some embodiments, the heteroaryl group is a 5-6 member aromatic ring system having a ring carbon atom provided to the aromatic ring system and 1-4 ring heteroatoms, where each heteroatom is independently selected from nitrogen, oxygen, and sulfur ("5-6 member heteroaryl"). In some embodiments, the 5-6 member heteroaryl has 1-3 ring heteroatoms selected from nitrogen, oxygen, and sulfur. In some embodiments, the 5-6 member heteroaryl has 1-2 ring heteroatoms selected from nitrogen, oxygen, and sulfur. In some embodiments, the 5- to 6-membered heteroaryl group has one ring heteroatom selected from nitrogen, oxygen, and sulfur. Unless otherwise specified, each example of a heteroaryl group is independently either unsubstituted ("unsubstituted heteroaryl") or substituted with one or more substituents ("substituted heteroaryl"). In some embodiments, the heteroaryl group is an unsubstituted 5- to 14-membered heteroaryl. In some embodiments, the heteroaryl group is a substituted 5- to 14-membered heteroaryl.

[0112] The term "unsaturated bond" refers to a double or triple bond. The terms "unsaturated" or "partially unsaturated" refer to a portion containing at least one double or triple bond. The term "saturated" refers to a portion that does not contain any double or triple bonds; that is, it contains only single bonds.

[0113] Adding the suffix "-ene" to a base group indicates that the group is a divalent part; for example, alkylene is the divalent part of alkyl. Unless otherwise specified, the groups are optionally substituted. The term "optionally substituted" means either substituted or unsubstituted. In some embodiments, alkyl groups, alkenyl groups, alkynyl groups, heteroalkyl groups, heteroalkenyl groups, heteroalkynyl groups, carbocykryl groups, heterocyclyl groups, aryl groups, and heteroaryl groups are optionally substituted. "Optionally substituted" refers to a group that may be substituted or unsubstituted (e.g., a "substituted" or "unsubstituted" alkyl group, a "substituted" or "unsubstituted" alkenyl group, a "substituted" or "unsubstituted" alkynyl group, a "substituted" or "unsubstituted" heteroalkyl group, a "substituted" or "unsubstituted" heteroalkenyl group, a "substituted" or "unsubstituted" heteroalkynyl group, a "substituted" or "unsubstituted" carbocyclyl group, a "substituted" or "unsubstituted" heterocyclyl group, a "substituted" or "unsubstituted" aryl group, or a "substituted" or "unsubstituted" heteroaryl group). Generally, the term "substituted" means that at least one hydrogen atom present in the group is replaced by an acceptable substituent, such as a substituent that, upon substitution, produces a stable compound, such as a compound that does not spontaneously undergo transformation by rearrangement, cyclization, elimination, or other reactions. Unless otherwise indicated, a “substituted” group has substituents at one or more substitutable positions of the group, and if two or more positions in a given structure are substituted, the substituents are either identical or different at each position. The term “substituted” is considered to include substitution by all acceptable substituents of an organic compound, and includes any substituents described herein that result in the formation of a stable compound. The present invention aims to achieve any such combination to obtain a stable compound. For the purposes of the present invention, heteroatoms such as nitrogen may have hydrogen substituents and / or any suitable substituents described herein that satisfy the valence of the heteroatom and result in the formation of a stable moiety. The present invention is not intended to be limited in any way by the exemplary substituents described herein.

[0114] Examples of carbon atom substituents include, but are not limited to, halogens, -CN, -NO2, -N3, -SO2H, -SO3H, -OH, and -OR aa , -ON(R bb )2, -N(R bb )2, -N(R bb )3 + X - , -N(OR cc )R bb -SH, -SR aa , -SSR cc -C(=O)R aa -CO2H, -CHO, -C(OR cc )3, -CO2R aa -OC(=O)R aa , -OCO2R aa -C(=O)N(R bb )2, -OC(=O)N(R bb )2, -NR bb C(=O)R aa , -NR bb CO2R aa , -NR bb C(=O)N(R bb )2, -C(=NR bb )R aa -C(=NR bb )OR aa -OC(=NR bb )R aa -OC(=NR bb )OR aa -C(=NR bb )N(R bb )2, -OC(=NR bb )N(R bb )2, -NR bb C(=NR bb )N(R bb )2, -C(=O)NR bb SO2R aa , -NR bb SO2R aa , -SO2N(R bb )2, -SO2R aa , -SO2OR aa , -OSO2R aa -S(=O)R aa -OS(=O)R aa , -Si(Raa )3、-OSi(R aa )3-C(=S)N(R bb )2、-C(=O)SR aa 、-C(=S)SR aa 、-SC(=S)SR aa 、-SC(=O)SR aa 、-OC(=O)SR aa 、-SC(=O)OR aa 、-SC(=O)R aa 、-P(=O)(R aa )2、-P(=O)(OR cc )2、-OP(=O)(R aa )2、-OP(=O)(OR cc )2、-P(=O)(N(R bb )2)2、-OP(=O)(N(R bb )2)2、-NR bb P(=O)(R aa )2、-NR bb P(=O)(OR cc )2、-NR bb P(=O)(N(R bb )2)2、-P(R cc )2、-P(OR cc )2、-P(R cc )3 + X - 、-P(OR cc )3 + X - 、-P(R cc )4、-P(OR cc )4、-OP(R cc )2、-OP(R cc )3 + X - 、-OP(OR cc )2、-OP(OR cc )3 + X - 、-OP(R cc )4、-OP(OR cc )4、-B(R aa )2、-B(OR cc )2、-BR aa (OR cc )、C 1~10 アルキル、C 1~10 ペルハロアルキル、C 2~10 アルケニル、C2~10 Alkinyl, HeteroC 1~10 Alkyl, hetero C 2~10 Alkenyl, HeteroC 2~10 Alkinyl, C 3~10 Carbocyclyl, 3-14 member heterocyclyl, C 6~14 The compounds include aryls and 5-14 member heteroaryls, where each alkyl, alkenyl, alkynyl, heteroalkyl, heteroalkenyl, heteroalkynyl, carbocykrill, heterocyclyl, aryl, and heteroaryl independently has 0, 1, 2, 3, 4, or 5 R dd It is substituted with a base, where X - Is it a counterion, or are the two geminal hydrogens on the carbon atom =O, =S, =NN(R) bb )2, =NNR bb C(=O)R aa ,=NNR bb C(=O)OR aa ,=NNR bb S(=O)2R aa ,=NR bb , or =NOR cc It is replaced by R aa Each example is independent of C 1~10 Alkyl, C 1~10 Perhaloalkyl, C 2~10 Alkenil, C 2~10 Alkinyl, HeteroC 1~10 Alkyl, hetero C 2~10 Alkenyl, HeteroC 2~10 Alkinyl, C 3~10 Carbocyclyl, 3-14 member heterocyclyl, C 6~14 Selected from aryls and 5-14 member heteroaryls, or two R aa The groups bond to form a 3-14 membered heterocyclyl or 5-14 membered heteroaryl ring, where each alkyl, alkenyl, alkynyl, heteroalkyl, heteroalkenyl, heteroalkynyl, carbocykrill, heterocyclyl, aryl, and heteroaryl independently has 0, 1, 2, 3, 4, or 5 R groups. dd It is substituted with R bb Each example independently represents hydrogen, -OH, and -OR.aa , -N(R cc )2, -CN, -C(=O)R aa -C(=O)N(R cc )2, -CO2R aa , -SO2R aa -C(=NR cc )OR aa -C(=NR cc )N(R cc )2, -SO2N(R cc )2, -SO2R cc , -SO2OR cc -SOR aa -C(=S)N(R cc )2, -C(=O)SR cc -C(=S)SR cc -P(=O)(R aa )2, -P(=O)(OR cc )2, -P(=O)(N(R cc )2)2, C 1~10 Alkyl, C 1~10 Perhaloalkyl, C 2~10 Alkenil, C 2~10 Alkinyl, HeteroC 1~10 Alkyl, hetero C 2~10 Alkenyl, HeteroC 2~10 Alkinyl, C 3~10 Carbocyclyl, 3-14 member heterocyclyl, C 6~14 Selected from aryls and 5-14 member heteroaryls, or two R bb The groups bond to form a 3-14 membered heterocyclyl or 5-14 membered heteroaryl ring, where each alkyl, alkenyl, alkynyl, heteroalkyl, heteroalkenyl, heteroalkynyl, carbocykrill, heterocyclyl, aryl, and heteroaryl independently has 0, 1, 2, 3, 4, or 5 R groups. dd It is substituted with a base, where X - R is a counterion, cc Each example independently involves hydrogen, C 1~10 Alkyl, C 1~10 Perhaloalkyl, C 2~10 Alkenil, C 2~10 Alkinyl, HeteroC 1~10 Alkyl, hetero C2~10 Alkenyl, HeteroC 2~10 Alkinyl, C 3~10 Carbocyclyl, 3-14 member heterocyclyl, C 6~14 Selected from aryls and 5-14 member heteroaryls, or two R cc The groups bond to form a 3-14 membered heterocyclyl or 5-14 membered heteroaryl ring, where each alkyl, alkenyl, alkynyl, heteroalkyl, heteroalkenyl, heteroalkynyl, carbocykrill, heterocyclyl, aryl, and heteroaryl independently has 0, 1, 2, 3, 4, or 5 R groups. dd It is substituted with R dd Each example is independently halogen, -CN, -NO2, -N3, -SO2H, -SO3H, -OH, -OR ee , -ON(R ff )2, -N(R ff )2, -N(R ff )3 + X - , -N(OR ee )R ff -SH, -SR ee , -SSR ee -C(=O)R ee -CO2H, -CO2R ee -OC(=O)R ee , -OCO2R ee -C(=O)N(R ff )2, -OC(=O)N(R ff )2, -NR ff C(=O)R ee , -NR ff CO2R ee , -NR ff C(=O)N(R ff )2, -C(=NR ff )OR ee -OC(=NR ff )R ee -OC(=NR ff )OR ee -C(=NR ff )N(R ff )2, -OC(=NR ff )N(R ff )2, -NR ff C(=NRff )N(R ff )2, -NR ff SO2R ee , -SO2N(R ff )2, -SO2R ee , -SO2OR ee , -OSO2R ee -S(=O)R ee , -Si(R ee )3, -OSi(R ee )3, -C(=S)N(R ff )2, -C(=O)SR ee -C(=S)SR ee -SC(=S)SR ee , -P(=O)(OR ee )2, -P(=O)(R ee )2, -OP(=O)(R ee )2, -OP(=O)(OR ee )2, C 1~6 Alkyl, C 1~6 Perhaloalkyl, C 2~6 Alkenil, C 2~6 Alkinyl, HeteroC 1~6 Alkyl, hetero C 2~6 Alkenyl, HeteroC 2~6 Alkinyl, C 3~10 Carbocyclyl, 3-10 member heterocyclyl, C 6~10 Selected from aryls and 5-10 membered heteroaryls, where each alkyl, alkenyl, alkynyl, heteroalkyl, heteroalkenyl, heteroalkynyl, carbocykrill, heterocyclyl, aryl, and heteroaryl independently has 0, 1, 2, 3, 4, or 5 R gg Substituted with or two Geminal R dd The substituents can bond to form either =O or =S. Yes, here, X - R is a counterion, ee Each example is independent of C 1~6 Alkyl, C 1~6 Perhaloalkyl, C 2~6 Alkenil, C 2~6 Alkinyl, HeteroC 1~6 Alkyl, hetero C 2~6 Alkenyl, HeteroC2~6 Alkinyl, C 3~10 Carbocyclyl, C 6~10 Selected from aryls, 3-10 membered heterocyclyls, and 3-10 membered heteroaryls, where each alkyl, alkenyl, alkynyl, heteroalkyl, heteroalkenyl, heteroalkynyl, carbocykrill, heterocyclyl, aryl, and heteroaryl independently has 0, 1, 2, 3, 4, or 5 R gg It is substituted with R ff Each example independently involves hydrogen, C 1~6 Alkyl, C 1~6 Perhaloalkyl, C 2~6 Alkenil, C 2~6 Alkinyl, HeteroC 1~6 Alkyl, hetero C 2~6 Alkenyl, HeteroC 2~6 Alkinyl, C 3~10 Carbocyclyl, 3-10 member heterocyclyl, C 6~10 Selected from aryls and 5-10 member heteroaryls, or two R ff The groups bond to form a 3-10 membered heterocyclyl or 5-10 membered heteroaryl ring, where each alkyl, alkenyl, alkynyl, heteroalkyl, heteroalkenyl, heteroalkynyl, carbocykrill, heterocyclyl, aryl, and heteroaryl independently has 0, 1, 2, 3, 4, or 5 R groups. gg Substituted with R gg Each of these examples is independent of halogen, -CN, -NO2, -N3, -SO2H, -SO3H, -OH, and -OC. 1~6 Alkyl, -ON(C 1~6 Alkyl)2,-N(C 1~6 Alkyl)2,-N(C 1~6 Alkyl)3 + X - , -NH(C 1~6 Alkyl)2 + X - -NH2(C 1~6 Alkyl) + X - , -NH3 + X - , -N(OC 1~6 Alkyl)(C1~6 Alkyl), -N(OH)(C 1~6 Alkyl), -NH(OH), -SH, -SC 1~6 Alkyl, -SS(C 1~6 Alkyl), -C(=O)(C 1~6 Alkyl), -CO2H, -CO2(C 1~6 Alkyl), -OC(=O)(C 1~6 Alkyl), -OCO2(C 1~6 Alkyl), -C(=O)NH2, -C(=O)N(C 1~6 Alkyl)2,-OC(=O)NH(C 1~6 Alkyl), -NHC(=O)(C 1~6 Alkyl), -N(C 1~6 Alkyl)C(=O)(C 1~6 Alkyl), -NHCO2(C 1~6 Alkyl), -NHC(=O)N(C 1~6 Alkyl)2,-NHC(=O)NH(C 1~6 Alkyl), -NHC(=O)NH2, -C(=NH)O(C 1~6 Alkyl), -OC(=NH)(C 1~6 Alkyl), -OC(=NH)OC 1~6 Alkyl, -C(=NH)N(C 1~6 Alkyl)2,-C(=NH)NH(C 1~6 Alkyl), -C(=NH)NH2, -OC(=NH)N(C 1~6 Alkyl)2,-OC(=NH)NH(C 1~6 Alkyl), -OC(=NH)NH2, -NHC(=NH)N(C 1~6 Alkyl)2, -NHC(=NH)NH2, -NHSO2(C 1~6 Alkyl), -SO2N(C 1~6 Alkyl)2,-SO2NH(C 1~6 Alkyl), -SO2NH2, -SO2(C 1~6 Alkyl), -SO2O(C 1~6 Alkyl), -OSO2(C 1~6 Alkyl), -SO(C 1~6 Alkyl), -Si(C 1~6 Alkyl)3,-OSi(C 1~6 Alkyl)3-C(=S)N(C 1~6Alkyl)2, C(=S)NH(C 1~6 Alkyl), C(=S)NH2, -C(=O)S(C 1~6 Alkyl), -C(=S)SC 1~6 Alkyl, -SC(=S)SC 1~6 Alkyl, -P(=O)(OC 1~6 Alkyl)2, -P(=O)(C 1~6 Alkyl)2, -OP(=O)(C 1~6 Alkyl)2, -OP(=O)(OC 1~6 Alkyl)2, C 1~6 Alkyl, C 1~6 Perhaloalkyl, C 2~6 Alkenil, C 2~6 Alkinyl, HeteroC 1~6 Alkyl, hetero C 2~6 Alkenyl, HeteroC 2~6 Alkinyl, C 3~10 Carbocyclyl, C 6~10 It is an aryl, a 3-10 member heterocyclyl, a 5-10 member heteroaryl, or two geminal Rs. gg Substituents can bond to form =O or =S, where X - It is a counterion.

[0115] The term "halo" or "halogen" refers to fluorine (fluoro, -F), chlorine (chloro, -Cl), bromine (bromo, -Br), or iodine (iod, -I). The term "acyl" is derived from the general formula -C(=O)R X1 , -C(=O)OR X1 -C(=O)-OC(=O)R X1 -C(=O)SR X1 -C(=O)N(R X1 )2, -C(=S)R X1 -C(=S)N(R X1 )2, -C(=S)O(R X1 ), -C(=S)S(R X1 ), -C(=NR X1 )R X1 -C(=NR X1 )OR X1 -C(=NR X1 )SR X1, and -C(=NR X1 )N(R X1 ) refers to a group having 2, where R X1 This includes hydrogen, halogens, substituted or unsubstituted hydroxyl groups, substituted or unsubstituted thiols, substituted or unsubstituted aminos, substituted or unsubstituted acyls, cyclic or acyclic substituted or unsubstituted branched or unbranched aliphatic groups, cyclic or acyclic substituted or unsubstituted branched or unbranched heteroaliphatic groups, cyclic or acyclic substituted or unsubstituted branched or unbranched alkyl groups, cyclic or acyclic substituted or unsubstituted branched or unbranched alkenyl groups, substituted or unsubstituted alkynyl groups, substituted or unsubstituted aryl groups, substituted or unsubstituted heteroaryl groups, and aliphatic oxy Aliphaticoxy, heteroaliphaticoxy, alkyloxy, heteroalkyloxy, aryloxy, heteroaryloxy, aliphatic thioxy, heteroaliphatic thioxy, alkyl thioxy, heteroalkyl thioxy, aryl thioxy, heteroaryl thioxy, mono- or di-aliphatic amino, mono- or di-heteroaliphatic amino, mono- or di-alkylamino, mono- or di-heteroalkylamino, mono- or di-arylamino, or mono- or di-heteroarylamino, or two R X1The groups combine to form a 5-6 membered heterocycle. Exemplary acyl groups include aldehydes (-CHO), carboxylic acids (-CO2H), ketones, acyl halides, esters, amides, imines, carbonates, carbamates, and urea. Acyl substituents include, but are not limited to, any substituents described herein that result in the formation of a stable moiety (e.g., aliphatic, alkyl, alkenyl, alkynyl, heteroaliphatic, heterocyclic, aryl, heteroaryl, acyl, oxo, imino, thiooxo, cyano, isocyano, amino, azide, nitro, hydroxyl, thiol, halo, aliphatic amino, heteroaliphatic amino, alkylamino, heteroalkylamino, arylamino, heteroarylamino, alkylaryl, arylalkyl, aliphatic oxy, heteroaliphatic oxy, alkyloxy, heteroalkyloxy, aryloxy, heteroaryloxy, aliphatic thiooxy, heteroaliphatic thiooxy, alkyl thiooxy, heteroalkyl thiooxy, aryl thiooxy, heteroaryl thiooxy, acyloxy, etc., each of which may be further substituted or unsubstituted).

[0116] The nitrogen atom may or may not be substituted, as long as its valence allows, and includes primary, secondary, tertiary, and quaternary nitrogen atoms. Exemplary nitrogen atom substituents include, but are not limited to, hydrogen, -OH, and -OR. aa , -N(R cc )2, -CN, -C(=O)R aa -C(=O)N(R cc )2, -CO2R aa , -SO2R aa -C(=NR bb )R aa -C(=NR cc )OR aa -C(=NR cc )N(R cc )2, -SO2N(R cc )2, -SO2R cc , -SO2OR cc -SOR aa -C(=S)N(R cc)2, -C(=O)SR cc -C(=S)SR cc , -P(=O)(OR cc )2, -P(=O)(R aa )2, -P(=O)(N(R cc )2)2, C 1~10 Alkyl, C 1~10 Perhaloalkyl, C 2~10 Alkenil, C 2~10 Alkinyl, HeteroC 1~10 Alkyl, hetero C 2~10 Alkenyl, HeteroC 2~10 Alkinyl, C 3~10 Carbocyclyl, 3-14 member heterocyclyl, C 6~14 It contains aryls and 5- to 14-membered heteroaryls, or two R atoms bonded to the N atom. cc The groups bond to form a 3-14 membered heterocyclyl or 5-14 membered heteroaryl ring, where each alkyl, alkenyl, alkynyl, heteroalkyl, heteroalkenyl, heteroalkynyl, carbocykrill, heterocyclyl, aryl, and heteroaryl independently has 0, 1, 2, 3, 4, or 5 R groups. dd It is substituted with R aa , R bb , R cc and R dd This is defined as above.

[0117] In one embodiment, the substituent present on the nitrogen atom is a nitrogen protecting group (also referred to herein as an "amino protecting group"). Nitrogen protecting groups include, but are not limited to, -OH and -OR groups. aa , -N(R cc )2, -C(=O)R aa -C(=O)N(R cc )2, -CO2R aa , -SO2R aa -C(=NR cc )R aa -C(=NR cc )OR aa -C(=NR cc )N(R cc )2, -SO2N(R cc )2, -SO2Rcc , -SO2OR cc -SOR aa -C(=S)N(R cc )2, -C(=O)SR cc -C(=S)SR cc , C 1~10 Alkyl (e.g., aralkyl, heteroaralkyl), C 2~10 Alkenil, C 2~10 Alkinyl, HeteroC 1~10 Alkyl, hetero C 2~10 Alkenyl, HeteroC 2~10 Alkinyl, C 3~10 Carbocyclyl, 3-14 member heterocyclyl, C 6~14 The group comprises aryl and 5-14 membered heteroaryl groups, where each alkyl, alkenyl, alkynyl, heteroalkyl, heteroalkenyl, heteroalkynyl, carbocykrill, heterocyclyl, aralkyl, aryl, and heteroaryl independently has 0, 1, 2, 3, 4, or 5 R groups. dd It is substituted with R aa , R bb , R cc and R dd This is as defined herein. Nitrogen protecting groups are well known in the art and include those described in detail in "Protecting Groups in Organic Synthetis, T.W. Greene and P.M. Wuts, 3rd edition, John Wiley & Sons, 1999," which is incorporated herein by reference.

[0118] In one embodiment, the substituent present on the oxygen atom is an oxygen protecting group (also referred to herein as a "hydroxyl protecting group"). Oxygen protecting groups include, but are not limited to, -R aa , -N(R bb )2, -C(=O)SR aa -C(=O)R aa , -CO2R aa -C(=O)N(R bb )2, -C(=NR bb )R aa -C(=NRbb )OR aa -C(=NR bb )N(R bb )2, -S(=O)R aa , -SO2R aa , -Si(R aa )3, -P(R cc )2, -P(R cc )3 + X - , -P(OR cc )2, -P(OR cc )3 + X - -P(=O)(R aa )2, -P(=O)(OR cc )2, and -P(=O)(N(R bb )2)2 is included, where X - , R aa , R bb , and R cc This is as defined herein. Oxygen protecting groups are well known in the art and include those described in detail in "Protecting Groups in Organic Synthetis," T.W. Green and P.M. Wutz, 3rd edition, John Wiley & Sons, 1999, which is incorporated herein by reference.

[0119] In one embodiment, the substituent present on the sulfur atom is a sulfur protecting group (also referred to herein as a "thiol protecting group"). Sulfur protecting groups include, but are not limited to, -R aa , -N(R bb )2, -C(=O)SR aa -C(=O)R aa , -CO2R aa -C(=O)N(R bb )2, -C(=NR bb )R aa -C(=NR bb )OR aa -C(=NR bb )N(R bb )2, -S(=O)R aa , -SO2R aa , -Si(R aa )3, -P(R cc)2, -P(R cc )3 + X - , -P(OR cc )2, -P(OR cc )3 + X - -P(=O)(R aa )2, -P(=O)(OR cc )2, and -P(=O)(N(R bb )2)2 is included, where R aa , R bb , and R cc The term is as defined herein. Sulfur protecting groups are well known in the art and include those described in detail in "Protecting Groups in Organic Synthetis," T.W. Green and P.M. Woods, 3rd edition, John Wiley & Sons, 1999, which is incorporated herein by reference.

[0120] As used herein, the phrase “at least one example” refers to one, two, three, four, or more examples, but also includes a range of examples, such as one to four, one to three, one to two, two to four, two to three, or three to four.

[0121] A "non-hydrogen group" refers to any group defined for a specific variable that is not hydrogen. These and other exemplary substituents are described in more detail in the Detailed Description, Examples and Claims. The present invention is not intended to be limited in any way by the above exemplary listing of substituents.

[0122] Other definitions As used herein, “protein,” “peptide,” or “polypeptide” includes polymers of amino acid residues linked to one another by peptide bonds. The term refers to proteins, polypeptides, and peptides of any size, structure, or function. Typically, a protein is at least three amino acid long. A protein may refer to an individual protein or a group of proteins. The proteins of the present invention preferably contain only natural amino acids, but non-natural amino acids (i.e., compounds that do not exist in nature but can be incorporated into polypeptide chains) and / or amino acid analogs known in the art may be used instead. Also, one or more amino acids in a protein may be modified by the addition of chemical components such as carbohydrate groups, hydroxyl groups, phosphate groups, farnesyl groups, isofarnesyl groups, fatty acid groups, linkers for conjugation or functionalization, or by other modifications. A protein may also be a single molecule or a multimolecular complex. A protein may be a fragment of a naturally occurring protein or peptide. A protein may be of natural origin, a recombinant, a synthetic, or any combination thereof.

[0123] The terms "polynucleotide," "nucleotide sequence," "nucleic acid," "nucleic acid molecule," "nucleic acid sequence," and "oligonucleotide" refer to a series of nucleotide bases (also called "nucleotides") in DNA and RNA, and mean any chain of two or more nucleotides. Polynucleotides may be single-stranded or double-stranded chimeric mixtures, derivatives thereof, or modified versions thereof. Oligonucleotides may be modified in the base portion, sugar portion, or phosphate backbone, for example, to improve molecular stability, their hybridization parameters, etc. Antisense oligonucleotides include 5-fluorouracil, 5-bromouracil, 5-chlorouracil, 5-iodouracil, hypoxanthine, xanthine, 4-acetylcytosine, 5-(carboxyhydroxylmethyl)uracil, 5-carboxymethylaminomethyl-2-thiouridine, 5-carboxymethylaminomethyluracil, dihydrouracil, beta-D-galactosylqueosine, inosine, N6-isopentenyladenine, 1-methylguanine, 1-methylinosine, 2,2-dimethylguanine, 2-methyladenine, 2-methylguanine, 3-methylcytosine, 5-methylcytosine, N6-adenine, 7-methylguanine, 5-methylaminomethyluracil, and 5-methoxyaminomethyl Modified base moieties may include, but are not limited to, groups such as thio-2-thiouracil, beta-D-mannosylqueosine, 5'-methoxycarboxymethyluracil, 5-methoxyuracil, 2-methylthio-N6-isopentenyladenine, wybutoxosine, pseudouracil, queosine, 2-thiocytosine, 5-methyl-2-thiouracil, 2-thiouracil, 4-thiouracil, 5-methyluracil, uracil-5-oxyacetate methyl ester, uracil-5-oxyacetate, 5-methyl-2-thiouracil, 3-(3-amino-3-N-2-carboxypropyl)uracil, thioguanine, and 2,6-diaminopurine. Nucleotide sequences typically contain genetic information that is used by cellular mechanisms to make proteins and enzymes.These terms include double-stranded or single-stranded genomic and cDNA, RNA, any synthetic and genetically engineered polynucleotides, and both sense and antisense polynucleotides. This includes single-stranded and double-stranded molecules, namely DNA-DNA, DNA-RNA and RNA-RNA hybrids, and "protein nucleic acids" (PNAs) formed by conjugating bases to an amino acid backbone. This also includes nucleic acids containing carbohydrates or lipids. Exemplary DNAs include single-stranded DNA (ssDNA), double-stranded DNA (dsDNA), plasmid DNA (pDNA), genomic DNA (gDNA), complementary DNA (cDNA), antisense DNA, chloroplast DNA (ctDNA or cpDNA), microsatellite DNA, mitochondrial DNA (mtDNA or mDNA), kinetoplast DNA (kDNA), proviruses, lysogenic bacteria, repetitive DNA, satellite DNA, and viral DNA. Exemplary RNAs include single-stranded RNA (ssRNA), double-stranded RNA (dsRNA), small interfering RNA (siRNA), messenger RNA (mRNA), precursor messenger RNA (pre-mRNA), small hairpin RNA or short hairpin RNA (shRNA), microRNA (miRNA), guide RNA (gRNA), transfer RNA (tRNA), antisense RNA (asRNA), heteronuclear RNA (hnRNA), coding RNA, non-coding RNA (ncRNA), long non-coding RNA (long ncRNA or lncRNA), satellite RNA, viral satellite RNA, signal recognition particle RNA, small cytoplasmic RNA, nuclear small RNA (snRNA), ribosomal RNA (rRNA), Piwi-interacting RNA (piRNA), polyinosinate, ribozymes, flexizymes, nucleolar small RNA (snoRNA), splice reader RNA, viral RNA, and viral satellite RNA. [Examples]

[0124] These and other aspects of the present invention will be further understood by considering the following embodiments. The following embodiments are intended to illustrate certain embodiments of the present invention, but are not intended to limit the scope defined by the claims.

[0125] Example 1. Gas-phase phosphonic acid dichloride for surface modification The selectivity of gas-phase phosphonic acid dichloride for surface modification was evaluated by contact angle measurement and X-ray photoelectron spectroscopy (XPS). Coupons with surface regions composed of TiO2 and SiO2 were treated with gas-phase octylphosphonic acid dichloride (PDC) by chemical vapor deposition. This was expected to form a surface coating with hydrophobic properties.

[0126] The results of the selectivity experiment are shown in Figure 3. The TiO2 side of the surface-modified coupon showed a high contact angle (approximately 99°) in the wettability measurement (Figure 3, left image) and a high phosphorus signal based on XPS measurement (Figure 3, left spectrum). The SiO2 side showed a relatively lower contact angle (approximately 30°) in the wettability measurement (Figure 3, right image), and no phosphorus signal was detected based on XPS measurement (Figure 3, right spectrum). These results indicate that vapor-phase deposition of PDC forms a coating on metal oxide surfaces with high selectivity compared to silica surfaces.

[0127] The surface coverage of coatings formed using liquid-phase hexylphosphonic acid (HPA) or gas-phase PDC was compared. Based on data from XPS and atomic force microscopy (AFM), the gas-phase PDC coating achieved better coverage in a more uniform form compared to the liquid-phase coating (Figures 4A-4B). The HPA-coated surface showed transparent spots approximately 2.7 nm high, roughly the size of two to three HPA molecules stacked end to end (Figure 4A). In comparison, the XPS and AFM data for the gas-phase PDC coating were consistent with the formation of a uniform monolayer with a more uniform surface distribution and coverage (Figure 4B).

[0128] Example 2. Stable silane surface coating for corrosive reaction conditions Corrosive reaction conditions during biological reactions (e.g., sequencing) can cause corrosion of the pore structure on the array and cleavage of the functionalized portion on the functionalized bottom surface. To provide a stable coating layer that can withstand such conditions, sample wells having a functionalized silica surface and metal oxide sidewalls were treated with silane using a hexachlorodisiloxane (HCDS) and silane crosslinked coating.

[0129] Using HCDS, a relatively thick silane surface coating was introduced without obstructing access to the functional moieties (terminal azide functional groups) at the bottom surface. Process time was optimized to ensure that the functional moieties were available for bonding or reaction, and stability was evaluated by comparing XPS of silane-treated substrates after immersion in strong acid for 24 hours with that of an untreated control. The results showed that the HCDS coating withstood harsh chemical conditions well (Figure 5). Different silane crosslinking coatings (aminopropyltrimethoxysilane or hexyltrichlorosilane) were applied to the HCDS coating. Both showed high durability and performance in biological sequencing reactions.

[0130] Example 3. Effect of pre-treatment with organophosphorus surface coating on silane treatment. A sample well array having metal oxide surface portions and silica surface portions can be used to monitor biological reactions by immobilizing the target molecule on the functionalized silica surface of individual sample wells. The pre-surface modification process for preparing the functionalized silica surface involves first passivating the metal oxide surface with an organophosphorus coating before functionalizing the silica surface by silane treatment. Such a process was based on the assumption that the organophosphorus coating would inhibit the silane treatment of the metal oxide surface, thereby promoting the selective functionalization of the silica surface.

[0131] After collecting data from XPS on coating compositions of metal oxide surfaces across multiple array process batches, a clear positive proportional correlation was observed between phosphorus ("percentage of P") derived from the organophosphorus coating and silicon ("percentage of Si") derived from the downstream biotinylated silane-treated coating (Figure 6). This observation indicated that increasing the amount of organophosphorus coating promoted silane treatment of the metal oxide surface, which contradicted the assumption that the organophosphorus coating inhibited silane treatment.

[0132] To further evaluate the possibility that the organophosphorus coating accelerated downstream silane treatment, sample well arrays with and without organophosphorus coating (formed by gas-phase PDC) were treated with trialkoxysilane in the gas phase, and the silane-treated metal oxide surfaces were analyzed by XPS. The results of the dual experiment are shown in Table 1.

[0133] [Table 1]

[0134] The XPS results in Table 1 show that the amount of silane on the organophosphorus-coated metal oxide surface was approximately 60–70% higher than that on the uncoated metal oxide surface. This data supports earlier observations that organophosphorus coatings accelerate silane treatment of downstream metal oxide surfaces.

[0135] The effect of pre-coating with organophosphorus on the surface selectivity of biotin-silane functionalization was quantitatively characterized by time-of-flight secondary ion mass spectrometry (TOF-SIMS). A control experiment was performed first to identify the characteristic fragment peaks of biotin-silane on both silica (SiO2) and metal oxide (TiO2) surfaces. Based on the spectra obtained during the control experiment (Figure 7), the distinctive fragment peaks of biotin were found on both the TiO2 and SiO2 surfaces, specifically in the CN region. - , CNO- C2H5O + It was identified as such.

[0136] Substrates having SiO2 and TiO2 surfaces with or without organophosphorus coatings were treated with biotin-PEG-silane, and the biotinylated silane-treated substrates were subjected to TOF-SIMS. The relative amounts of fragment peaks were quantified, and these results are shown in Figure 8. PO2 - The fragment peaks originating from the sample are shown to clarify that the replicated "Sample 1" and "Sample 2" were coated with gas-phase PDC before biotinylated silane treatment, while the replicated "Sample 3" and "Sample 4" were not coated with gas-phase PDC.

[0137] Further analysis of the data shown in Figure 8 quantified the selectivity of biotinylated silane treatment on the SiO2 surface compared to the TiO2 surface of the tested substrate. Figure 9 shows the unique fragment peak of biotin (C2H5O + , CNO - 5CN - The results of this analysis for ) are shown. Selectivity (SiO2 / TiO2) was calculated for each fragment peak using the formula shown in the upper panel of Figure 9.

[0138] The biotinylation selectivity of SiO2 / TiO2 calculated from fragment ions in both positive and negative modes consistently showed that substrates without organophosphorus coating on TiO2 (Samples 3 and 4) exhibited higher selectivity for biotinylation silane treatment of the SiO2 surface than substrates with organophosphorus coating on TiO2 (Samples 1 and 2). Based on experimental observations and data, it was concluded that pre-coating of metal oxide surfaces with organophosphorus coatings promotes the formation of silane-treated coatings on metal oxide surfaces, rather than inhibiting silane treatment as previously thought.

[0139] Example 4. Scalable surface modification in the gas phase After determining that organophosphorus coatings promote silane treatment of metal oxide surfaces, a surface modification process was devised for a sample well array, in which biotinylated silane treatment of the SiO2 surface was performed before gas-phase PDC coating of the TiO2 surface. Next, based on the theory that the organophosphorus coating formed on the TiO2 surface by PDC acts as a base layer to promote silane treatment, the array was treated with silane to form an upper layer on top of both surface compositions. Chemical and physical characterization of the treated array revealed hydrophobic behavior of the silane-treated surface with a contact angle of approximately 90°, complete coverage of the coating on the chip with a scaly morphology in AFM images, and high percentages of typical elements (Si and P) detected in XPS spectra (Figure 10).

[0140] The total process time was approximately 5 hours, including the initial plasma activation and intermittent rinsing and drying steps. After activation and rinsing / drying, the array was treated with a liquid-phase mixture of PEG-silane and biotin-PEG-silane for approximately 1 hour for bottom surface functionalization. After rinsing and drying, the array was treated with gas-phase octylphosphonic acid dichloride by chemical vapor deposition for approximately 30 minutes for sidewall surface pre-treatment. Next, the array surface was treated with HCDS and hexyltrichlorosilane by chemical vapor deposition for approximately 2.5 hours to form a stable silane coating layer on the sample well surface.

[0141] Equivalents and range In the claims, articles such as “a,” “an,” and “the” may mean one or more unless otherwise indicated or otherwise evident from the context. A claim or statement containing “or” between one or more elements of a group is considered satisfied unless otherwise indicated or otherwise evident from the context if one, two or more, or all of the elements of the group are present, used, or otherwise related to a given product or process. The present invention includes embodiments in which exactly one element of the group is present, used, or otherwise related to a given product or process. The present invention includes embodiments in which two or more, or all, of the elements of the group are present, used, or otherwise related to a given product or process.

[0142] Furthermore, the present invention encompasses all variations, combinations, and substitutions in which one or more limitations, elements, clauses, and descriptive terms from one or more of the listed claims are introduced into another claim. For example, any claim dependent on another claim may be modified to include one or more limitations found in other claims dependent on the same basic claim. Where elements are presented listed, for example in Markush group form, each subgroup of those elements is also disclosed, and any element(s) may be excluded from the group. In general, where the present invention or an aspect of the present invention is referred to as including certain elements and / or features, it should be understood that a certain embodiment or aspect of the present invention consists of or is essentially such elements and / or features. For brevity, these embodiments are not specifically shown in this specification by these words. Also note that the terms “including” and “contain” are intended to be open and allow for the inclusion of additional elements or steps. Where a scope is given, an endpoint is included. Furthermore, unless otherwise indicated, or unless otherwise evident from the context and the understanding of those skilled in the art, values ​​expressed as ranges may, unless the context explicitly indicates, be any specific value or subrange within the ranges described in different embodiments of the present invention, up to one-tenth of the lower limit unit of the range.

[0143] This application references various granted patents, published patent applications, journal articles, and other publications, all of which are incorporated herein by reference. In the event of any conflict between any of the references incorporated herein and this specification, this specification shall prevail. In addition, any particular embodiment of the present invention that is in the prior art may be expressly excluded from any one or more of the claims. Such embodiments may be excluded even if their exclusion is not expressly indicated herein, as they would be known to those skilled in the art. Any particular embodiment of the present invention may be excluded from any claim for any reason, whether or not it is related to the existence of the prior art.

[0144] Those skilled in the art will be able to recognize or confirm many equivalents to the specific embodiments described herein by means of routine experiments alone. The scope of the embodiments described herein is not intended to be limited to the above specification, but rather as set forth in the appended claims. Those skilled in the art will understand that various changes and modifications to this specification may be made without departing from the spirit or scope of the invention, as defined in the following claims.

Claims

1. A method for forming a phosphorus-containing layer on a metallic surface, the method being: A method comprising treating the metallic surface with a phosphoryl halide in the gas phase, wherein the phosphoryl halide forms the phosphorus-containing layer on the metallic surface.

2. The method according to claim 1, wherein the metallic surface is a metal surface or a metal oxide surface.

3. The method according to claim 1 or 2, wherein the metallic surface is a transition metal oxide surface.

4. The method according to any one of claims 1 to 3, wherein the metallic surface is a titanium oxide surface, an aluminum oxide surface, a zirconium oxide surface, an iron oxide surface, a tin oxide surface, or a tantalum oxide surface.

5. The aforementioned metallic surface is titanium dioxide (TiO ２ The method according to any one of claims 1 to 4, wherein the surface is...

6. The method according to any one of claims 1 to 5, wherein the phosphoryl halide is phosphoryl dihalide.

7. The method according to any one of claims 1 to 5, wherein the phosphoryl halide is an organic phosphoryl halide and the phosphorus-containing layer is an organic phosphorus layer.

8. The method according to claim 7, wherein the organophosphorus layer is an organophosphonate layer.

9. The method according to any one of claims 1 to 8, wherein the phosphoryl halide is an organic phosphoryl dihalide.

10. The method according to any one of claims 1 to 9, wherein the phosphoryl halide is an organic phosphoryl dichloride.

11. The method according to any one of claims 1 to 10, wherein the phosphoryl halide has a molecular weight of about 132 g / mol to 260 g / mol.

12. The method according to any one of claims 1 to 11, wherein the phosphoryl halide has a molecular weight of about 132 g / mol to 232 g / mol.

13. The method according to any one of claims 1 to 12, wherein the phosphoryl halide has a molecular weight of about 203 g / mol to 232 g / mol.

14. The phosphoryl halide is given by formula (I): 【Chemistry 1】 It is either the same as or its salt, X １ Each of these examples is independently a halogen, R １ This includes optionally substituted alkyl, optionally substituted heteroalkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted carbocyclyl, optionally substituted heterocyclyl, optionally substituted aryl, or optionally substituted heteroaryl, -OR Ｏ , or -N(R Ｎ ) ２ And, R Ｏ Each example independently consists of hydrogen, an optionally substituted alkyl, an optionally substituted alkenyl, an optionally substituted alkynyl, an optionally substituted carbocyclyl, an optionally substituted heterocyclyl, an optionally substituted aryl, an optionally substituted heteroaryl, an optionally substituted acyl, or an oxygen protecting group, or optionally two R Ｏ However, together with the intervening atom, it forms a heterocycline in which substitutions are made at will. R Ｎ Each example of which is, independently, hydrogen, optionally substituted alkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted carbocyclic, optionally substituted heterocyclic, optionally substituted aryl, optionally substituted heteroaryl, optionally substituted acyl, or a nitrogen protecting group, or optionally two R Ｎ together with intervening atoms form an optionally substituted heterocyclic, the method according to any one of claims 1 to 13.

15. R １ The method according to claim 14, wherein is an optionally substituted alkyl.

16. R １ C is replaced by an optional substitution. １～１０ The method according to claim 14 or 15, wherein the alkyl group is alkyl.

17. R １ C is replaced by an optional substitution. １～８ The method according to any one of claims 14 to 16, wherein the alkyl group is alkyl.

18. R １ is a non-substitutive C １～８ The method according to any one of claims 14 to 17, wherein the alkyl group is alkyl.

19. R １ C is replaced by an optional substitution. ６～８ The method according to any one of claims 14 to 17, wherein the alkyl group is alkyl.

20. R １ is a non-substitutive C ６～８ The method according to any one of claims 14 to 17, wherein the alkyl group is alkyl.

21. The compound of formula (I) is 【Chemistry 2】 A method according to any one of claims 14 to 17, selected from the group consisting of the following.

22. The phosphoryl halide is given by the following formula: 【Transformation 3】 It is either the same as or its salt, L １ The method according to claim 14, wherein is an optionally substituted alkylene.

23. L １ C is replaced by an optional substitution. １～６ The method according to claim 22, wherein the material is alkylene.

24. L １ is an unsubstituted C １～６ The method according to claim 22 or 23, wherein the material is alkylene.

25. The phosphoryl halide is given by formula (II): 【Chemistry 4】 It is either the same as or its salt, X １ It is a halogen, R ２ Each example is an optionally substituted alkyl, optionally substituted heteroalkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted carbocyclyl, optionally substituted heterocyclyl, optionally substituted aryl, or optionally substituted heteroaryl, -OR Ｏ , or -N(R Ｎ ) ２ Either or, at your discretion, two R's ２ The group, together with the intervening atom, forms a heterocycline in which substitution is optional. R Ｏ Each example independently consists of hydrogen, an optionally substituted alkyl, an optionally substituted alkenyl, an optionally substituted alkynyl, an optionally substituted carbocyclyl, an optionally substituted heterocyclyl, an optionally substituted aryl, an optionally substituted heteroaryl, an optionally substituted acyl, or an oxygen protecting group, or optionally two R Ｏ However, together with the intervening atom, it forms a heterocycline in which substitutions are made at will. R Ｎ Each example independently consists of hydrogen, an optionally substituted alkyl, an optionally substituted alkenyl, an optionally substituted alkynyl, an optionally substituted carbocyclyl, an optionally substituted heterocyclyl, an optionally substituted aryl, an optionally substituted heteroaryl, an optionally substituted acyl, or a nitrogen protecting group, or optionally two R Ｎ The method according to any one of claims 1 to 5, wherein the intervening atoms together form a heterocycline that is optionally substituted.

26. X １ The method according to any one of claims 14 to 25, wherein each example is Cl.

27. The method according to any one of claims 1 to 5, wherein the phosphoryl halide is a phosphoryl trihalide.

28. The method according to claim 27, wherein the phosphoryl halide is phosphoryl trichloride.

29. The method according to any one of claims 1 to 28, further comprising treating the metallic surface with at least one chlorosilane in the gas phase, wherein the at least one chlorosilane forms a coating layer on the metallic surface.

30. The method according to claim 29, wherein the at least one chlorosilane is a chlorosiloxane compound, an alkylchlorosilane compound, or both thereof.

31. The method according to claim 30, wherein the at least one chlorosilane is hexachlorodisiloxane, hexyltrichlorosilane, or both thereof.

32. The method according to any one of claims 29 to 31, wherein at least a portion of the coating layer is formed on the phosphorus-containing layer.

33. A method for modifying the surface of a substrate, wherein the method is A method comprising treating a substrate having a first surface portion and a second surface portion with a phosphoryl halide in the gas phase, wherein the phosphoryl halide preferentially forms a phosphorus-containing layer on the first surface portion, and the first surface portion and the second surface portion have different surface properties.

34. The method according to claim 33, wherein the first surface portion is a metal surface or a metal oxide surface.

35. The method according to claim 33 or 34, wherein the first surface portion is a transition metal oxide surface.

36. The method according to any one of claims 33 to 35, wherein the first surface portion is a titanium oxide surface, an aluminum oxide surface, a zirconium oxide surface, an iron oxide surface, a tin oxide surface, or a tantalum oxide surface.

37. The first surface portion is titanium dioxide (TiO ２ The method according to any one of claims 33 to 36, wherein the surface is...

38. The method according to any one of claims 33 to 37, wherein the second surface portion is a transparent surface or a glass surface.

39. The second surface portion is silica (SiO ２ The method according to any one of claims 33 to 38, wherein the surface is...

40. The method according to any one of claims 33 to 39, wherein the phosphoryl halide preferentially forms the phosphorus-containing layer on the first surface portion with a selectivity of about 2 to about 60 times.

41. The method according to any one of claims 33 to 40, wherein the phosphoryl halide preferentially forms the phosphorus-containing layer on the first surface portion with a selectivity of about 4 to about 40 times.

42. The method according to any one of claims 33 to 41, wherein the phosphoryl halide preferentially forms the phosphorus-containing layer on the first surface portion with a selectivity of about 8 to about 20 times.

43. The method according to any one of claims 33 to 42, further comprising treating the substrate with at least one chlorosilane in the gas phase, wherein the at least one chlorosilane forms a coating layer on a first surface portion and a second surface portion.

44. The method according to claim 43, wherein the at least one chlorosilane is a chlorosiloxane compound, an alkylchlorosilane compound, or both thereof.

45. The method according to claim 44, wherein the at least one chlorosilane is hexachlorodisiloxane, hexyltrichlorosilane, or both thereof.

46. The method according to any one of claims 43 to 45, wherein at least a portion of the coating layer is formed on the phosphorus-containing layer.

47. The method according to any one of claims 33 to 46, wherein the phosphoryl halide is an organic phosphoryl halide and the phosphorus-containing layer is an organic phosphorus layer.

48. The method according to claim 47, wherein the organophosphorus layer is an organic phosphonate layer.

49. The method according to any one of claims 33 to 48, further comprising treating the substrate with a functionalizing agent having a coupling portion before treating the substrate with the phosphoryl halide in the gas phase, wherein the functionalizing agent functionalizes the second surface portion by preferentially bonding to the second surface portion.

50. The method according to claim 49, wherein the coupling portion is a covalent coupling portion or a non-covalent coupling portion.

51. The method according to claim 50, wherein the covalent coupling portion is a trans-cyclooctene (TCO) portion, a tetrazine portion, an azide portion, an alkyne portion, an aldehyde portion, an isocyanate portion, an N-hydroxysuccinimide portion, a thiol portion, an alkene portion, a dibenzocyclooctyl portion, a bicyclononine portion, or a thiamine pyrophosphate portion.

52. The method according to claim 50, wherein the non-covalent coupling portion is a biotin portion, an avidin protein, a streptavidin protein, a lectin protein, or a SNAP tag.

53. The method according to any one of claims 49 to 52, wherein the functionalizing agent includes a portion that can preferentially bond to silica.

54. The method according to claim 53, wherein the portion that can preferentially bond to the silica is silane.

55. The method according to claim 54, wherein the silane is monoethoxysilane, methoxysilane, diethoxysilane, trichlorosilane, or diethoxymethoxysilane.

56. The method according to any one of claims 49 to 55, wherein the functionalizing agent comprises biotinylated silane.

57. The method according to any one of claims 33 to 56, wherein the substrate comprises an array of sample wells.

58. The method according to claim 57, wherein the sample well of the array has an upper opening, the upper opening extends into the substrate to a bottom surface distal to the upper opening, and the sample well has a side wall surface disposed between the upper opening and the bottom surface.

59. The method according to claim 58, wherein the side wall surface comprises a first surface portion, the bottom surface comprises a second surface portion, and the phosphoryl halide preferentially forms the phosphorus-containing layer on the side wall surface of the sample well.

60. A method for functionalizing the surface of a sample well, (a) Metal oxide surface and silica (SiO ２ A step of treating a sample well having a surface with a functionalizing agent containing a coupling portion, wherein the functionalizing agent functionalizes the surface of the sample well by preferentially bonding to the silica surface, (b) A step of treating the sample well of (a) with an organic reagent in the gas phase, wherein the organic reagent preferentially forms an organic layer on the metal oxide surface. Methods that include...

61. (c) The method according to claim 60, further comprising the step of treating the sample well with one or more additional reagents that form a coating layer on the metal oxide surface and the silica surface.

62. The method according to claim 60 or 61, wherein the coating layer on the metal oxide surface is formed on the side wall of the sample well, and the coating layer on the silica surface is formed on the bottom surface of the sample well.

63. The metal oxide surface is titanium dioxide (TiO ２ The method according to any one of claims 60 to 62, wherein the surface is...

64. The method according to any one of claims 60 to 63, wherein the functionalizing agent includes a portion that can preferentially bond to silica.

65. The method according to claim 64, wherein the portion that can preferentially bond to the silica is silane.

66. The method according to claim 65, wherein the silane is monoethoxysilane, methoxysilane, diethoxysilane, trichlorosilane, or diethoxymethoxysilane.

67. The method according to any one of claims 60 to 66, wherein the coupling portion is a covalent coupling portion or a non-covalent coupling portion.

68. The method according to claim 67, wherein the covalent coupling portion is a trans-cyclooctene (TCO) portion, a tetrazine portion, an azide portion, an alkyne portion, an aldehyde portion, an isocyanate portion, an N-hydroxysuccinimide portion, a thiol portion, an alkene portion, a dibenzocyclooctyl portion, a bicyclononine portion, or a thiamine pyrophosphate portion.

69. The method according to claim 67, wherein the non-covalent coupling portion is a biotin portion, an avidin protein, a streptavidin protein, a lectin protein, or a snap tag.

70. The method according to any one of claims 60 to 69, wherein the functionalizing agent comprises biotinylated silane.

71. The method according to any one of claims 60 to 70, wherein the organic reagent is an organic phosphoryl halide and the organic layer is an organic phosphorus layer.

72. The method according to any one of claims 61 to 71, wherein the one or more additional reagents are in the gas phase.

73. The method according to any one of claims 61 to 72, wherein the one or more additional reagents are silanes, and the coating layer is a silane layer.

74. The method according to claim 72, wherein the one or more additional reagents are chlorosilanes.

75. The method according to claim 73 or 74, wherein one or more of the additional reagents are hexachlorodisiloxane, hexyltrichlorosilane, or both thereof.

76. The method according to any one of claims 60 to 75, further comprising the step of bringing a molecule of interest to bond to the coupling portion into contact with the sample well, thereby coupling the molecule of interest to the surface of the sample well.

77. The method according to claim 76, wherein the target molecule is a biomolecule.

78. The method according to claim 77, wherein the biomolecule is a nucleic acid or polypeptide.

79. The method according to claim 78, wherein the polypeptide is a protein or a fragment thereof.

80. The method according to claim 78, wherein the polypeptide is a polymerase.

81. The method according to claim 80, wherein the polymerase is a nucleic acid polymerase.

82. The method according to claim 79, wherein the polypeptide is a substrate for a polypeptide sequencing reaction.

83. The method according to any one of claims 79 to 81, wherein the polypeptide is not a substrate for a polypeptide sequencing reaction.

84. The method according to claim 83, wherein the polypeptide promotes a nucleic acid sequencing reaction.

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