Nanometer groove array and manufacturing method and application thereof

Abstract

The invention relates to a nano groove array and a manufacturing method and application thereof, and the manufacturing method of the nano groove array comprises the following steps: taking single-stranded DNA (ssDNA) as a DNA brick, and carrying out self-assembly by adopting a complementary pairing principle of the ssDNA to obtain the nano groove array with a base and a side wall; wherein the width and the height of the nano groove array are fixed, the number of grooves is determined, and the nano groove array extends along a single direction; in the nano groove array, the width of the side wall and the width of the groove are equivalent to the diameter of two double-helix DNA (dsDNA). Compared with the prior art, the number of the grooves of the nano groove array is controllable, the width of the grooves is 4 nm, the period of the groove array is 8 nm, and the self-assembly defect can be greatly reduced.

Description

TECHNICAL FIELD

[0001] The present application relates to the technical field of nanomaterials, and in particular to a nanoslot array and a manufacturing method and application thereof. BACKGROUND

[0002] A nanoslot structure can construct parallel isolated regions in a very small space, and can be used for precisely arranging low-dimensional materials, and has important application value in the fields of semiconductor devices, nanophotonic devices, optoelectronics, and nanoelectromechanical dynamics. In the research and experiment stage, the preparation of a single nanoslot with controllable morphology can basically meet the needs of most tests, but when the target is upgraded to a nanoslot array with regular morphology and consistent period at the nanoscale, the preparation process puts forward very high requirements on the integrity of the design process, the stability of the equipment precision, and the reliability of the material performance.

[0003] To meet the array preparation requirements, one of the current mainstream technologies is extreme ultraviolet lithography technology based on semiconductor chip manufacturing. Although this technology can stably realize the manufacturing of a nanoslot array with a size of more than 10 nm, it faces key bottlenecks in the preparation of a nanoslot array with a size of less than 10 nm under the demand for “smaller size” in the post-moore era. In 2024, ASML Corporation of the Netherlands announced the realization of 8 nm dense line slot printing, which is a technological breakthrough, but is limited by the high cost of equipment, and cannot be commercialized until 2033, which makes it difficult to meet the rapid preparation requirements of a nanoslot array with a size of less than 10 nm for research and small and medium-sized industries at the current stage.

[0004] To break through the size and cost limitations of extreme ultraviolet lithography, DNA nanotechnology based on self-assembly principles has become a new research direction. This technology can realize a feature structure with a resolution of 2 nm through self-assembly of DNA bases in solution, and supports user self-design of 2D or 3D nanostructures: only need to add single-stranded DNA (ssDNA) with a specific sequence to the solution, and after annealing treatment, the preset DNA structure can be obtained. According to different design logics, the main template construction methods are divided into DNA tiles, DNA origami, and DNA bricks, among which the DNA brick nanotechnology can design uniformly spaced nanoslots to construct a nanoslot array in space; and the formed DNA nanoslot structure can be used as a synthesis template for oxides, graphene, plasmonic materials, polymers, and carbon nanotubes, etc., providing a new path for the preparation of nanomaterials in multiple fields.

[0005] Specifically, the DNA brick technology is originally used for designing a 3D structure with three mutually perpendicular extension axes, and the extension of the extension axis can be realized by designing a circular chain in the vertical section of the extension axis. At present, there are studies that have successfully constructed a groove array with a groove spacing of 10.6 nm by designing two extension axes. However, this strategy has obvious limitations in the preparation of planar nanogroove DNA templates: the construction method of taking two-axis direction extension is easy to cause assembly defects, and the number of nanogrooves cannot be accurately controlled, resulting in randomness in the length, width and edge defect number of the same batch of DNA templates, which directly affects the preparation yield and structure consistency of the planar nanogroove array, and is difficult to adapt to the application scenarios with strict precision requirements. SUMMARY

[0006] The purpose of the present application is to provide a nanogroove array and its manufacturing method and application, which utilizes the self-assembly of DNA bricks to form a nanogroove array with a groove spacing of less than 10 nm.

[0007] The purpose of the present application can be achieved by the following technical solutions: a manufacturing method of a nanogroove array, using ssDNA (single-stranded DNA) as a DNA brick, and using the complementary pairing principle of ssDNA for self-assembly to obtain a nanogroove array with a base and a sidewall. In the nanogroove array, the width and height are fixed, the number of grooves is determined, and the extension is along a single direction.

[0008] Preferably, in the nanogroove array, the width of the sidewall and the groove is 2 construction units, i.e. the width of the sidewall and the groove is equivalent to 2 dsDNA (double-stranded DNA) diameters.

[0009] The present application discloses a manufacturing method of a groove array with a spacing of 8 nm. The main construction material of this groove array is ssDNA (single-stranded DNA). In design, it is used as a DNA brick to build the required groove array, i.e. a DNA template. The present application uses 8 bases of ssDNA as a binding domain, and 8 bp (base pairs) of dsDNA (double-stranded DNA) as a construction unit, wherein the extension direction of the double-stranded DNA is parallel to the groove array.

[0010] Preferably, during self-assembly, the ssDNA with at least two binding domains is sequence-paired with other at least two ssDNA with matching binding domains.

[0011] Preferably, the ssDNA has 2-6 binding domains, and each binding domain has 8 bases.

[0012] Further preferably, according to the structural stability requirement, the 20-25% ssDNA contains 2 binding domains, the 60-65% ssDNA contains 4 binding domains, and the ~15% ssDNA contains 6 binding domains.

[0013] In the present application, the complementary pairing principle of ssDNA is adopted, and the ssDNA generally has 4 binding domains: the number of binding domains of a single ssDNA is appropriately increased or decreased according to the position of the structure, so as to improve the stability of the structure, and the minimum number is 2 and the maximum number is 6; each binding domain has 8 bases: the number of bases is reduced according to the requirement of reducing the overall spin. When annealing, the ssDNA with at least two binding domains is paired with other ssDNA with at least two matching binding domains. With the continuous pairing and stacking of the DNA bricks, a complex expected structure is gradually formed.

[0014] Preferably, when self-assembling, a buffer solution for stabilizing the solution environment, Mg 2+ for adjusting the charge of ssDNA, and Tris base (tris-hydroxymethyl aminomethane) for adjusting the pH of the solution are added to the aqueous solution containing all ssDNA, the ssDNA is denatured by first increasing the temperature, and then annealing is performed.

[0015] Further preferably, the pH of the solution is adjusted to be greater than 8.5, the ssDNA is denatured by first increasing the temperature to 80℃, and then annealing is performed.

[0016] Further preferably, after annealing, it is kept at 4℃.

[0017] In the present application, a buffer solution for stabilizing the solution environment, Mg 2+ for adjusting the charge of ssDNA, and Tris base (tris-hydroxymethyl aminomethane) for adjusting the pH of the solution are also added to the aqueous solution containing all DNA bricks. The assembly solution can be temperature-controlled using a PCR instrument, the ssDNA is denatured by first increasing the temperature, and then annealing is performed by gradient cooling or constant temperature, etc. After annealing is completed, the solution is kept at 4℃ for subsequent purification, characterization or other applications.

[0018] Preferably, the elongation direction of the double helix DNA is defined as the z-axis, the nanoslot array extends along the z-axis, and a corresponding periodic unit is designed on the double helix DNA, and the double helix DNA is repeatedly spliced in the z-axis direction by connecting the head and tail of the periodic unit.

[0019] In the application, the extension direction of the double helix DNA is parallel to the groove array, which is defined as the z-axis. The x-axis and y-axis perpendicular to the z-axis, the construction unit can be expanded in 6 directions along the 3 extension axes. The nano groove array of the design is extended along the z-axis, so the corresponding period unit needs to be designed, and the period unit is connected at the head and tail to realize repeated splicing in the z-axis direction, so as to obtain a long strip template with fixed x-axis and y-axis sizes and micron scale z-axis.

[0020] Preferably, the manufacturing method of the nano groove array comprises the following steps: (1) initially designing a DNA template with a nano groove array: using a construction unit to simulate the construction of a geometry with a base and a side wall in three-dimensional space to form a nano groove array, wherein the width of the side wall and the groove is 2 construction units respectively; (2) modifying the length of the DNA chain in the design: filling and connecting in the geometry in step (1) with 32 nt (base) as the main ssDNA chain length, wherein the DNA brick in the boundary area is modified to 48 nt ssDNA chain length, and the DNA brick in the side wall area is modified to less than 32 nt ssDNA chain length; (3) adjusting the template twist: deleting several bases of the DNA template to realize twist compensation in the long period; (4) sequence assignment: after the spatial position arrangement of the ssDNA is completed, according to the principle of base complementary pairing, all ssDNA are assigned with base sequences one by one, i.e. A, T, C, G; (5) DNA chain annealing: mixing all ssDNA in buffer, adjusting pH, heating, and then annealing to obtain a DNA template solution with a nano groove array.

[0021] In the application, the design structure of the DNA template has fixed sizes in the x-axis and y-axis directions and extends infinitely in the z-axis direction in three-dimensional space. When self-assembled, the design structure is connected at the head and tail in the z-axis direction, so that the target synthesized DNA template extends along the z-axis. The x-axis, y-axis and z-axis directions mentioned herein are only relative positions.

[0022] In the application, the base and the side wall with specific x-axis and y-axis sizes are constructed by using DNA bricks. The base is connected with multiple side walls, the overall width of the nano groove array is determined by the width of the base, the area containing the side wall can be changed by changing the width, the height of the nano groove array is determined by the base and the side wall together, the number of nano grooves is determined by the number of side walls on the base, and the spacing of the nano grooves is determined by the spacing of the side walls.

[0023] In the application, the width of the designed base can be adjusted appropriately according to the application scene, and the width is limited to contain a controllable number of side walls.

[0024] In the present application, the trench design requirements include: ① the minimum trench spacing is 4 nm; ② reduce the overall chiral spin of the template itself; ③ a single DNA template can accommodate 6 nanometer trenches; ④ the designed trench unit has unidirectional extensibility; ⑤ adjusting the length of the ssDNA at the edge of the structure can improve stability.

[0025] Further preferably, the base layer of the DNA template is 4 layers or more.

[0026] Further preferably, the side wall height of the DNA template is 4 layers.

[0027] Further preferably, when designing the base and the side wall in the x and y axis directions, double helix DNA is used as the construction unit, and the minimum feature size in the x and y axis directions is 2 nm. The designed side wall is composed of double helix DNA with a width of 2 layers and a height of 4 layers, so the side wall is 4 nm wide and 8 nm high. The designed base is 4 layers of double helix DNA, and the side wall is stacked on the base. Therefore, the height of the nanometer trench array is 16 nm.

[0028] Further preferably, in step (2), the 50% ssDNA brick in the side wall region is modified to have a ssDNA chain length of 16 nt.

[0029] Further preferably, in step (3), the twist compensation in the long period is achieved by deleting 1 base pair at the end of the DNA template.

[0030] In the present application, the overall twist design requirement is that there is no overall twist in the range of about 500 nm in length.

[0031] Further preferably, the design length of the DNA template is set to 63 bp (base pairs) or a multiple thereof.

[0032] More preferably, the design length of the DNA template is set to 63 bp.

[0033] Further preferably, in step (4), no consecutive bases or consecutive CGs appear in the ssDNA.

[0034] More preferably, in step (4), the consecutive bases or consecutive CGs refer to the number of consecutive bases or CGs being 5 or more. For example, AAAAA, TTTTTTT, CGCGCGCGCG.

[0035] Further preferably, in step (5), the buffer is a TE buffer containing Mg 2+ .

[0036] Further preferably, in step (5), Tris base is added to the buffer solution, and the pH is controlled to be greater than 8.5 by adjusting the concentration of the Tris base.

[0037] Further preferably, in step (5), the concentration of each ssDNA in the solution used for annealing is not less than 100 nM.

[0038] Further preferably, in step (5), the temperature is first raised to 80°C, and then annealing is performed, followed by stepwise cooling to 60°C, and then stepwise cooling to 25°C, and the entire annealing time is 40-200 h.

[0039] More preferably, when the temperature is stepwise cooled to 60°C, the temperature is maintained for >2 min per °C.

[0040] More preferably, when the temperature is stepwise cooled to 25°C, the temperature is maintained for >1 h per °C.

[0041] Further preferably, after step (5) in which the DNA template solution with a nanoslot array is obtained, a centrifugal filter is used to remove excess ssDNA.

[0042] More preferably, the pore size of the centrifugal filter is in the range of 10 kD-300 kD, and the relative centrifugal force is 2400 x g, and the centrifugation time is 2-5 min, and the centrifugation is performed at least 4 times.

[0043] More preferably, after centrifugation, the bottom of the centrifugal filter is washed with a buffer solution containing Mg 2+ , TE, and Tris.

[0044] More preferably, the bottom of the centrifugal filter is washed with the buffer solution to further disperse the DNA template, and the washing is performed at least 10 times each time.

[0045] Preferably, the method for manufacturing the nanoslot array comprises the following steps: 1) Preliminary design and construction of a DNA template with a nanoslot array: a geometric structure with a base and a side wall is constructed in a three-dimensional space using a building unit to form a slot array. The width of the side wall and the slot is 2 building units, about 4 nm.

[0046] 2) Modify the length of the DNA chain in the design: fill and connect in the above-mentioned geometric structure using 32 nt (bases) as the main ssDNA chain length. In addition, the DNA bricks located in the boundary region need to be modified to 48 nt ssDNA chains to ensure the stability of the structure. It should be noted that ssDNA chains with a length of less than 32 nt are used in the side wall region.

[0047] 3) Adjusting template twist: 8 bp building block will cause systematic overall twist of DNA template, which can be compensated by artificially deleting several bases in long period.

[0048] 4) Sequence assignment: after spatial arrangement of ssDNA, base sequence is assigned to all ssDNA according to base complementary pairing principle, i.e. A, T, C, G.

[0049] 5) DNA strand annealing: all ssDNA strands are mixed in specific buffer and adjusted to specific pH. Subsequent annealing treatment can obtain DNA template solution with specific nanoslot array.

[0050] 6) Remove excess ssDNA by centrifugal filter.

[0051] Further preferably, in step 1), the DNA template is periodically extended along the z-axis direction, which intuitively represents the extension of the nanoslot in the z-axis direction. During the self-assembly of DNA, the extension speed in the z-axis direction is faster than that in the x-axis and y-axis directions. Taking multi-directional extension (simultaneous extension in the x-axis and z-axis directions) is easy to cause various assembly defects caused by assembly misalignment, while single-directional periodic extension can avoid the occurrence of such defects.

[0052] Further preferably, the number of base layers of the DNA template is 4 or more, which can improve the rigidity of the overall structure and help resist systematic overall twist. The side wall height is adjusted according to the assembly material, and generally also uses 4 layers.

[0053] Further preferably, in step 2), 50% of the ssDNA bricks of the side wall use 16 nt ssDNA strands to prevent the formation of the side wall at a higher temperature in the annealing step, which can cause the side wall structure to be free and cause the side wall of the DNA template to be missing.

[0054] Further preferably, in step 3), the helical rotation period of double-stranded DNA (dsDNA) in nature is 10.5 bp. Therefore, the design length of the DNA template should be set to 63 bp or its multiples to meet the design of 8 bp building blocks, that is, 63 bp can be obtained by removing 1 bp in 8 building blocks, which is exactly equal to 6 rotation periods (6x10.5 bp = 63 bp), thereby weakening the overall spin effect of double-helical DNA.

[0055] Further preferably, in step 4), too many consecutive bases or consecutive CGs, such as AAAAA, TTTTTTT, CGCGCGCGCG, etc., should be avoided.

[0056] Further preferably, in step 5), the buffer system adopts a TE buffer containing Mg 2+ , and the pH>8.5 is controlled by adjusting the concentration of Tris base. In the final solution used for annealing, the concentration of each ssDNA is not less than 100 nM; during the annealing process, the temperature is first heated to 80℃, then gradually reduced to 60℃, and each ℃ is kept for >2 min, and then gradually reduced to 25℃, and each ℃ is kept for >1 h, and the whole annealing time is 40-200 h.

[0057] Further preferably, in step 6), the removal of the excess un-assembled ssDNA requires the use of a centrifugal filter with a pore size range of 10 kD-300 kD, a relative centrifugal force of 2400xg, centrifugation for 2-5 min, and at least 4 times of centrifugation, and the use of a buffer containing Mg 2+ , TE, and Tris for rinsing.

[0058] Further preferably, the bottom of the centrifuged residual solution is rinsed with a buffer each time, so that the DNA template is further dispersed, and the rinsing is performed at least 10 times each time.

[0059] A nano-groove array is prepared by the above manufacturing method, and the period of the nano-groove array is 8 nm, and the groove width is 4 nm.

[0060] The nano-groove array is applied to the field of semiconductor devices, nano-photonic devices, optoelectronics, or nano-quantum electrodynamics.

[0061] Compared with the prior art, the present application has the following beneficial effects: 1. The present application provides a manufacturing method of a groove array with a period of 8 nm and a groove width of 4 nm, which utilizes the self-assembly of DNA bricks to form a nano-groove array with an array pitch of less than 10 nm.

[0062] 2. The DNA template of the present application has the following significant features: the number of grooves of the nano-groove array is controllable; the groove width is 4 nm; the groove period is 8 nm; the design repeats the unit in a single extension axis direction; and the self-assembly defects can be greatly reduced.

[0063] 3. The present application can realize an array pitch of 8 nm, a groove width of 4 nm, and a sidewall width of 4 nm by controlling the size and shape of the nano-groove.

[0064] 4. The present application can precisely control the number of nano-grooves by setting addressable connection chains on the base and connecting the sidewall structure; and can construct neatly arranged nano-grooves by deleting specific position bases to weaken the overall twist.

[0065] 5. The present application uses periodic repeating units, takes a single expansion direction, and effectively controls assembly defects.

[0066] 6. The long axis size of the array of the present application reaches the micron scale, and the effective area of the array is large.

[0067] 7. The present application does not use organic volatile reagents and high-energy consumption equipment in the synthesis process, and is environmentally friendly.

[0068] 8. The present application has high synthesis efficiency, simple method, and is easy to scale up. BRIEF DESCRIPTION OF DRAWINGS

[0069] Figure 1 It is a schematic diagram of the mutual combination between ssDNA bricks and the binding domain in the present application.

[0070] Figure 2 It is a schematic diagram of the connection mode of ssDNA bricks in three-dimensional space.

[0071] Figure 3 It is a schematic diagram of the morphology of the DNA nanoslot.

[0072] Figure 4 It is a transmission electron microscope characterization diagram of the DNA nanoslot.

[0073] Figure 5 It is the DNA nanoslot spacing measured according to the electron microscope image.

[0074] Figure 6 It is the finite element modeling framework simulation before and after the length of the correction template unit.

[0075] Figure 7 It is a transmission electron microscope characterization diagram of the DNA nanoslot template after deleting 3 sidewalls. DETAILED DESCRIPTION

[0076] The present application will be described in detail below in combination with the drawings and specific embodiments. The present embodiment is implemented on the premise of the technical solution of the present application, and gives a detailed implementation manner and specific operation process, but the protection scope of the present application is not limited to the following examples.

[0077] A design and preparation method of a DNA nanoslot array, comprising the following steps: (1) Designing a DNA template with a nanoslot array: simulating and building a geometric structure with a base and a sidewall in three-dimensional space to form a nanoslot array; (2) Modify the length of DNA strand in the design: take 32 nt as the main ssDNA strand length, fill and connect in the geometry structure described in step (1), wherein the DNA bricks located in the boundary region are modified to 48 nt ssDNA strand length, and the DNA bricks located in the side wall region are modified to 32 nt or less ssDNA strand length; (3) Adjust the template twist: by deleting several bases of the DNA template, the twist compensation in the long period is realized; (4) Sequence assignment: after the spatial position arrangement of ssDNA is completed, according to the principle of base complementary pairing, all ssDNA are assigned with base sequences one by one, i.e. A, T, C, G; (5) DNA strand annealing: mix all ssDNA in buffer, adjust pH, heat, and then perform annealing treatment to obtain a DNA template solution with a nanoslot array.

[0078] As a preferred technical solution, when a DNA nanoslot array with an 8-nanometer period and a 4-nanometer width is needed to be designed and prepared, the base and the side wall are designed in the x and y axis directions, the double helix DNA is used as the construction unit, the minimum feature size of which in the x and y axis is 2 nm, the designed side wall is composed of 2-layer wide and 4-layer high double helix DNA, so the side wall is 4 nm wide and 8 nm high, the designed base is 4-layer double helix DNA, and the side wall is superimposed on the base, so the height of the nanoslot array is 16 nm, and the designed slot size is equivalent to 2-layer wide double helix DNA, so the slot width is 4 nm.

[0079] The following will be described in detail with specific examples.

[0080] Unless otherwise specified, the reagents, methods, instruments and equipment used in the present application are conventional reagents, methods, instruments and equipment in the art. Unless otherwise specified, the reagents and materials used in the following examples are commercially available.

[0081] Example 1 Design, preparation and characterization of a DNA nanoslot array with an 8-nanometer period and a 4-nanometer width Step 1: As Figure 1 , Figure 2As shown, by the design method of DNA bricks, the target structure was designed using nanobricks web software (https: / / yin.hms.harvard.edu / bricks / try / ), and then the DNA template with nanoslot features was generated. By cadnano open source software, the internal sequence of the DNA structure was edited to improve the overall stability, and the design length of the DNA template was modified to 63 bp. Under the requirement of sequence pairing required by the structure, the ssDNA therein was randomly assigned using cadnano.

[0082] Change the length of the local ssDNA: the boundary chain is changed to 48 nt, and the 50% side wall chain is changed to 16 nt; improve the overall twist of the designed structure: delete 1 base pair at the end of each layer on the template z axis. Make the edge structure stable, which is more conducive to obtaining the expected assembled structure, such as Figure 3 As shown.

[0083] The obtained ssDNA is dispersed in a low concentration of TE buffer, and the concentration is adjusted to 100 μM. 2 μL of each ssDNA is taken from all the ssDNA, mixed and dispersed in TE / Mg 2+ buffer, wherein the final concentration of each ssDNA is 100 nM. The mixed solution contains 1×TE, 40 mM Mg 2+ , and the pH is adjusted to 9.0 by Tris base.

[0084] The mixed solution is transferred to a PCR special centrifuge tube, and annealing assembly is carried out in a PCR instrument. The volume of the solution in a single PCR tube is 70 μL. Two-stage segmented cooling method is used for annealing operation. First, heat from room temperature to 80℃ at a speed of 6 ℃ / s, and keep for 15 min, and then carry out the first stage of rapid cooling (80℃→60℃): the temperature is reduced by an integer, each time by 1℃, the change rate is 0.1 ℃ / s, and each ℃ is kept for 6 min; then carry out the second stage of slow cooling (59℃→25℃): the temperature is reduced by an integer, each time by 1℃, the change rate is 0.1 ℃ / s, and each ℃ is kept for 3 h. When the annealing is completed, quickly reduce from 25℃ to 4℃ at a speed of 6 ℃ / s. At this time, the DNA nanoslot array is completed annealing assembly, and the assembly solution can be stored at 4℃ for a long time.

[0085] In order to reduce the excess ssDNA in the DNA template solution, a 30kD filter concentration centrifuge tube is used to purify the assembly solution. The DNA template in the centrifuge tube is washed and centrifuged several times using a washing solution of 1×TE, 40 mM Mg 2+ , pH 9.0, repeated 4 times. The purified DNA template solution can be stored at 4℃ for a long time.

[0086] The DNA templates were characterized using transmission electron microscopy (TEM).

[0087] The carbon support film copper mesh was subjected to glow discharge treatment to obtain hydrophilic properties. 4 μL of DNA template solution was added to the front surface of the copper mesh, deposited for 3 min, and then the excess solution was absorbed by clean filter paper close to the edge of the copper mesh. Then, 1% wt uranyl acetate solution was used for negative staining, generally 4 μL was added, deposited for 7 s, and then the excess solution was absorbed by clean filter paper close to the edge of the copper mesh. Then the copper mesh was placed on the sample rod and sent into the TEM vacuum chamber to obtain the morphology image.

[0088] Referring to Figure 4 The TEM characterization image of the assembled DNA nanoslot array. The black area between the arrays is the groove area highlighted after negative staining.

[0089] Referring to Figure 5 The size schematic diagram of the DNA nanoslot array. The width of the DNA template is basically constant, which is 53 nm, and extends along the z axis. The groove and sidewall widths are labeled, the average sidewall width is 4.01 nm; the average groove width is 4.21 nm; and the average array period is 8.22 nm.

[0090] Example 2 Finite element modeling framework simulation of DNA nanoslot array.

[0091] The overall twist correction effect of the DNA nanoslot was simulated by the open source web platform software Cando (https: / / cando-dna-origami.org / ). CanDo uses a mechanical model of DNA to predict the 3D shape and flexibility of programmed DNA, which assumes that the double helix structure is a uniform elastic rod with axial tensile, twist and bending stiffness. In the design, the length of a single template unit without twist correction is 64 bp, which is equivalent to 24 nm. In order to intuitively understand the twist of the DNA nanoslot array, 3 template units connected head to tail were selected as the simulated DNA structure. According to the requirements of the Cando operation manual, the free chains exposed on both ends were deleted. As shown in Figure 6 The thermal fluctuation three-view (RMSF) of the initial group (origin) with a length of 64 bp *3 was compared with the modified group (modified) with a length of 63 bp *3, and the results showed that the initial group was significantly twisted, and the twist characteristics of the modified group were reduced and did not occur overall twist. By comparing the simulation of this framework with the experimental characterization structure, it is confirmed that the designed DNA template structure can effectively prevent overall twist and can be used as a planar nanoslot array.

[0092] Example 3 Design, prepare, and characterize DNA groove array template containing 4 sidewalls (3 grooves).

[0093] Modify the target structure in Example 1 using open source software scadnano: delete 3 sidewalls, make it have 3 wider nanogrooves, and keep the remaining base sequence unchanged.

[0094] Based on the new structure, take 2 μL from the remaining ssDNA, mix, and disperse in TE / Mg 2+ buffer, where the final concentration of each ssDNA is adjusted to 100 nM. The mixed solution contains 1×TE, 40 mM Mg 2+ , and the pH is adjusted to 9.0 by Tris base.

[0095] Transfer the mixed solution to a PCR-specific centrifuge tube, and perform annealing assembly in a PCR instrument device, with a solution volume of 70 μL in a single PCR tube. Perform annealing operation using the same two-stage segmented cooling method as in Example 1. Purify the assembly solution using a 30 kD filter concentration centrifuge tube. When purifying, use 1×TE, 40 mM Mg 2+ , and pH 9.0 washing solution to wash and centrifuge the DNA template in the centrifuge tube multiple times, repeated 4 times. The purified DNA template solution can be stored for a long time at 4°C.

[0096] Use transmission electron microscopy (TEM) to characterize and analyze the DNA template, as shown in Figure 7 . The results show that the base width is consistent with the original template, and there are 4 sidewalls on the base, which are evenly spaced and separated by 3 wider nanogrooves.

[0097] The above description of the embodiments is to facilitate the understanding and use of the invention by those of ordinary skill in the art. Those skilled in the art can easily make various modifications to these embodiments, and apply the general principles described herein to other embodiments without having to go through creative labor. Therefore, the present invention is not limited to the above embodiments, and improvements and modifications made by those skilled in the art based on the disclosure of the present invention without departing from the scope of the present invention should be within the scope of protection of the present invention.

Claims

1. A method of manufacturing a nanogroove array, characterized by, The ssDNA is used as a DNA brick, and the self-assembly is performed according to the complementary pairing principle of the ssDNA to obtain a nanoslot array with a base and a side wall. The nanoslot array has a fixed width and height, a fixed number of grooves, and is elongated in a single direction.

2. The method of claim 1, wherein In a three-dimensional space, the design structure of the DNA template has a fixed size in the x-axis and y-axis directions and is infinitely extended in the z-axis direction. When self-assembled, the design structure is connected at the head and tail in the z-axis direction, so that the target synthesized DNA template is extended along the z-axis.

3. The method of claim 2, wherein The DNA brick is used to construct a base and a side wall with specific x-axis and y-axis sizes. The base is connected with multiple side walls. The overall width of the nanoslot array is determined by the width of the base. The area for accommodating the side wall is changed by changing the width. The height of the nanoslot array is determined by the base and the side wall. The number of nanoslots is determined by the number of side walls on the base. The spacing of the nanoslots is determined by the spacing of the side walls.

4. The method of claim 1, wherein When self-assembling, add buffer solution for stabilizing solution environment, Mg 2+ for adjusting charge of ssDNA, Tris base for adjusting pH of solution into aqueous solution containing all ssDNA, first raise temperature to make ssDNA denature, then anneal.

5. The method of claim 4, wherein The solution pH is adjusted to be greater than 8.5, and the ssDNA is denatured by being heated to 80℃, and then annealing is performed.

6. The method of claim 1, wherein The method comprises the following steps: (1) designing a DNA template with a nanoslot array: a geometric structure with a base and a side wall is simulated and built in a three-dimensional space to form a nanoslot array; (2) modifying the length of the DNA chain in the design: 32 nt is used as the main ssDNA chain length, and the geometric structure described in step (1) is filled and connected. The DNA brick in the boundary region is modified to have a 48 nt ssDNA chain length, and the DNA brick in the side wall region is modified to have a ssDNA chain length of less than 32 nt; (3) adjusting the template twist: the twist compensation in a long period is realized by deleting several bases of the DNA template; (4) sequence assignment: after the spatial position arrangement of the ssDNA is completed, the base sequences, i.e., A, T, C, and G, are assigned to all the ssDNA one by one according to the principle of base complementary pairing; (5) DNA chain annealing: all the ssDNA is mixed in a buffer solution, the pH is adjusted, the temperature is raised, and then annealing is performed to obtain a DNA template solution with a nanoslot array.

7. The method of claim 6, wherein When the base and the side wall are designed in the x-axis and y-axis directions, the double helix DNA is used as a construction unit, the minimum feature size of the double helix DNA in the x-axis and y-axis directions is 2 nm, the designed side wall is composed of 2 layers of double helix DNA in width and 4 layers of double helix DNA in height, so the side wall has a width of 4 nm and a height of 8 nm, the designed base is 4 layers of double helix DNA, and the side wall is stacked on the base. Therefore, the height of the nanoslot array is 16 nm.

8. The method of claim 6, wherein the nano-groove array is formed by a process comprising: In the step (2), 50% of the ssDNA bricks in the side wall region are modified to have a 16 nt ssDNA chain length. The design length of the DNA template is set to be 63 bp or a multiple of 63 bp.

9. A nanogroove array, characterized by, The nanoslot array is obtained by the manufacturing method of any one of claims 1-8, the groove width is 4 nm, and the period of the groove is 8 nm.

10. Use of the nanogroove array according to claim 9, characterized in that The nanoslot array is used in the fields of semiconductor devices, nanophotonic devices, optoelectronics, or nanoelectromechanical dynamics.

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