A class of DNA nanocomputing elements, their preparation methods and applications

Abstract

The present invention belongs to the field of DNA computing nanotechnology, and specifically relates to a class of DNA nanocomputing elements, a preparation method thereof, and an application thereof in solving the Hamiltonian path problem. The DNA nanocomputing element uses a DNA nanostructure as a DNA computing element, and the designed DNA nanostructure has uniform dimensions. The recombined programmable sites provide precise node site design, and the rich extensible side chains can provide a stable and reliable physical connection relationship. Based on a class of DNA nanocomputing elements of the present invention, the rectangular origami plates with a connection relationship are coaxial. When applied to solve the Hamiltonian path problem, it can ensure that the polymer formed by connecting multiple rectangular origami plates still has good rigidity, so as to ensure that the calculation results can be stored in the correct order without errors, facilitating the subsequent observation of the calculation results. The DNA nanocomputing element provided by the present invention has parallel computing ability and storage ability.

Description

Technical Field

[0001] The present invention belongs to the field of DNA computing nanotechnology, and particularly relates to a class of DNA nano-computing elements, a preparation method thereof, and an application thereof in solving the Hamiltonian path problem. Background Art

[0002] In 1945, von Neumann established the architecture of electronic computers with the Turing machine (TM) as the computing model and semiconductors as the implementation materials. The following year, the first electronic computer based on the von Neumann architecture was successfully developed. Since the advent of electronic computers based on TM, they have developed at an astonishing speed according to Moore's law. To date, electronic computers have gone through four main stages: tube computers, transistor computers, integrated circuit computers, and large-scale integrated circuit computers. However, due to the reduction in size and the increasing influence of quantum effects, silicon-based chips are gradually reaching their physical limits, and the silicon-based electronic information industry is facing a major development bottleneck. Moreover, the long-standing problem of Non-deterministic Polynomial (NP), that is, the non-deterministic problem of polynomial complexity, cannot be solved on existing electronic computers due to the limitations of TM machines. Researchers have begun to attempt to establish a computing model beyond TM, such as bionic computing (such as neural networks, evolutionary computing, particle swarm optimization, and computing), optical computing, and quantum computing, which have been proposed in the literature. However, the computing power of bionic computing and optical computing is comparable to that of TM machines. An algorithm with a computational complexity of n under TM can be reduced to √n under the quantum computing model, which means that even quantum computers do not exceed TM in terms of computing power. The probe machine model proposed by Professor Xu Jin uses DNA for computing and perfectly solves the NP-complete problem. Therefore, it gives us inspiration to use DNA computing and, with the help of its huge advantage - parallel computing, solve large-scale NP-complete problems in a short time.

[0003] In 1994, Adleman demonstrated a solution to the Hamiltonian path problem through DNA hybridization, proving that an algorithm can be encoded into DNA and used to perform computational operations. Subsequently, the sticky model, self-assembly model, non-enumeration model, and parallel DNA model were successively proposed. Further, DNA-based chemical circuits were built, with DNA or ligands as signal inputs and generating output signals, such as various DNA logic gates, biological detection DNA circuits, etc. Currently, there are more and more DNA-based molecular computers, which can also be clinically used for disease diagnosis and biomarker detection, etc.

[0004] DNA nanotechnology has advantages such as precise addressability, good biocompatibility, and high stability. Many more complex nanostructures have been created using the Watson-Crick base pairing principle of DNA. Especially in 2006, a brand-new DNA self-assembly method proposed by Rothemund - DNA origami - is a major milestone in the fields of DNA nanotechnology and DNA self-assembly. Different from traditional DNA self-assembly techniques, DNA origami can quickly and precisely construct highly complex nano-patterns or structures in a "one-pot" method by base complementarity between a long single-stranded DNA and a series of designed short DNA fragments. Therefore, in the past ten-odd years, DNA origami has received extensive attention in the field of DNA nanotechnology. New two-dimensional and three-dimensional structures have been continuously constructed, the assembly efficiency has been further improved, and new application functions have been continuously developed. In 2006, Qian Lulu et al. constructed a DNA nanostructure in the shape of a Chinese map based on DNA origami, which is also a major leap in the application of DNA origami to the construction of complex two-dimensional structures. Among these objects, by chemically modifying the main strand with functional units, different sites can be precisely located and thus can be attached after the programmed self-assembly of the DNA "origami" structure. Advanced analysis at the single-molecule level can be carried out using atomic force microscopy (AFM) or electron microscopy.

[0005] Based on the previous research results of this research group, see the literature: Yu Yang, Su Shao, Chao Jie. Designing a solution to the Hamiltonian path problem based on "DNA origami". Science China, 2015, 11: 1226 - 1230, which discloses a technical solution that combines DNA origami and DNA computing, using a rectangular two-dimensional DNA nanostructure as a DNA computing carrier to solve the Hamiltonian path problem. Utilizing the programmable characteristics of all sites on the rectangular flat plate, markings are made on the surface of the rectangular flat plate, and the path information in the relationship graph is encoded into complementary relationships through sticky ends. Through intermolecular recognition and self-assembly, the solution to the specific Hamiltonian path problem can be calculated. However, when applying this rectangular two-dimensional DNA nanostructure computing element in the prior art, there are problems such as unclear calculation results, inability to be stably stored, and inability to be applied to more complex path solving; Summary of the Invention

[0006] To solve the above technical problems, through ingenious site design and precise sticky end design in the present invention, only when there are nodes represented by rectangular origami in the relationship graph and there are path relationships between different nodes, can the self-assembly of DNA molecules be used in a one-pot annealing to mutually recognize and connect the paths to form polymers, and the calculation results will be stably stored in these polymers.

[0007] In a first aspect, the present invention provides a class of DNA nanocomputing elements, which are composed of a rectangular two-dimensional DNA nanostructure and a labeling structure;

[0008] The rectangular two-dimensional DNA nanostructure is a two-dimensional DNA structure. Currently, most two-dimensional DNA structures are prepared by DNA origami technology, which was proposed by Rothemund of the California Institute of Technology in 2006. See the literature "Folding DNA to create nanoscale shapes and patterns. Nature, 2006, 440: 297-302". The rectangular two-dimensional DNA nanostructure of the present invention is assembled from a DNA template strand and DNA staple strands; using the DNA template strand as the main chain and an excess of DNA staple strands as the auxiliary chain, the main chain and the auxiliary chain hybridize and complement at specific positions to assemble the DNA nanostructure; the DNA template strand is preferably the M13mp18 phage genomic DNA sequence provided in GenBank: X02513.1; the present invention uses the rectangular two-dimensional DNA nanostructure as an information carrier considering the programmable characteristics of all sites of the rectangular plate;

[0009] As the auxiliary chain, some DNA staple strands at specific positions are selected as programmable auxiliary chains, i.e., functional DNA strands, and the remaining DNA staple strands are non-functional DNA staple strands;

[0010] A part of the functional DNA strands, hereinafter referred to as sticky DNA strands, after assembly, extend one or two sets of sticky end groups on one or both of the two short sides of the main body of the rectangular two-dimensional DNA nanostructure, and each set of sticky end groups consists of at least 2 sticky end strands evenly distributed on the short side of the main body of the rectangular two-dimensional DNA nanostructure. The base sequences of all sticky end strands within the same set of sticky end groups are different to ensure that the rectangular two-dimensional DNA nanostructure maintains a high specific recognition ability and self-assembly yield; the left side of the rectangular two-dimensional DNA nanostructure is defined as the in-degree of the node, and the right side is defined as the out-degree of the node to ensure that all the origami have the same directionality;

[0011] Preferably, each set of sticky end groups consists of 9 sticky end strands evenly distributed on the short side of the main body of the rectangular two-dimensional DNA nanostructure;

[0012] Preferably, in order to maximize the utilization of the path encoding of the rectangular two-dimensional DNA nanostructure while ensuring a high self-assembly efficiency of the rectangular two-dimensional DNA nanostructure, the size of the rectangular two-dimensional DNA nanostructure is set to a length of 90 nm and a width of 60 nm; when setting two sets of sticky end groups in the same direction, in order to ensure that the two sets of sticky end groups on the same rectangular two-dimensional DNA nanostructure have an equal competitive relationship, the 18 sticky end chains arranged in the same direction are split into two groups in the same group and cross-arranged, for example, as Figure 1 shown, the two sets of sticky end groups arranged in the in-degree direction are respectively denoted as the in-A sticky end group and the in-B sticky end group, and the two sets of sticky end groups arranged in the out-degree direction are respectively denoted as the out-A sticky end group and the out-B sticky end group; taking the two sets of sticky end groups in the in-degree direction as an example, the specific method of splitting into two groups in the same group and cross-arranging is: splitting the 9 sticky end chains of the in-A sticky end group into 3 small groups respectively denoted as the in-A1 group, the in-A2 group, and the in-A3 group, with 3 sticky end chains in each small group; the 9 sticky end chains of the in-B sticky end group are also split into 3 small groups respectively denoted as the in-B1 group, the in-B2 group, and the in-B3 group, with 3 sticky end chains in each small group, and the in-A1 group, the in-B1 group, the in-A2 group, the in-B2 group, the in-A3 group, and the in-B3 group are arranged in sequence in the in-degree direction.

[0013] At least one capture sequence for capturing the labeled structure extends from the surface of the main body of the rectangular two-dimensional DNA nanostructure after the assembly of the other part of the functional DNA strand, hereinafter referred to as the DNA capture strand; preferably, the capture sequence is a 15-nt nucleotide sequence added to the 5' end of the DNA staple strand, and more preferably, the capture sequence is 15-nt poly-A or poly-T, so as to precisely control the number and relative position of the labeled structure, such as biotin-ssDNA, on the surface of the DNA nanostructure; preferably, 12 capture sequences are set on the DNA nanostructure and anchored on the surface of the DNA nanostructure, and one labeled structure is captured by 3 capture sequences;

[0014] The labeled structure is not limited to gold nanoparticles-thiolated oligonucleotides, biotin-streptavidin, and fluorescent groups; the biotin-streptavidin is a complex formed by the reaction of a biotin-ssDNA complex and streptavidin SA; the labeled structure contains a sequence complementary to the capture sequence.

[0015] As a preferred embodiment, the preparation process of the above DNA nanocomputing element specifically includes the following steps:

[0016] (1) Mix the DNA template strand, non-functional DNA staple strand, and DNA functional strand in a buffer in proportion, anneal, and ultrafiltrate and purify to obtain a rectangular two-dimensional DNA nanostructure; (2) Mix the annealed product obtained in step (1) with the biotin-ssDNA complex in proportion and then anneal; (3) Mix the annealed product obtained in step (2) with SA in proportion and then anneal; (4) Ultrafiltrate and purify the annealed product obtained in step (3) to replace the buffer to obtain a DNA nanocomputing element.

[0017] Preferably, the annealing conditions in step (1) are: starting temperature 95°C, holding for 3 min, ending temperature 25°C, with a gradient of 1°C each, and staying at each gradient for 100 s.

[0018] Preferably, the molar ratio of the DNA template strand, non-functional DNA staple strand, and functional DNA strand in step (1) is 1:10:10.

[0019] Preferably, the annealing conditions in step (2) are: starting temperature 37°C, ending temperature 25°C, with a gradient of 1°C each, staying at each gradient for 10 min, and maintaining the annealing time for 2 h.

[0020] Preferably, the molar ratio of the annealed product to the biotin-ssDNA complex in step (2) is 1:50.

[0021] Preferably, the annealing conditions in step (3) are: maintaining at 37°C for 1 h;

[0022] Preferably, the molar ratio of the annealed product to SA at a single site in step (3) is 1:10;

[0023] Preferably, the buffer in steps (1) and (4) is 1×TAE / Mg 2+ ;

[0024] Preferably, the ultrafiltration step in steps (1) and (4) is to mix the obtained annealed product with 1×TAE / Mg 2+ buffer and add it to a 100 kDa ultrafiltration tube, and then centrifuge.

[0025] In a second aspect, the present invention also provides the application of the above DNA nanocomputing element in solving the Hamiltonian path problem, and the specific application includes the following steps:

[0026] Obtain preset conditions according to the Hamiltonian path problem to be solved, encode DNA sequences for the sticky ends of the rectangular two-dimensional DNA nanostructure according to the preset conditions, and label the rectangular two-dimensional DNA nanostructure with a labeling structure to construct a variety of different DNA nanocomputing elements; the preset conditions include the names of each node and the path relationships between each node, where using a labeling structure to label the rectangular two-dimensional DNA nanostructure represents node information; encode DNA sequences for the sticky end groups of the rectangular two-dimensional DNA nanostructure, and match and connect different rectangular two-dimensional DNA nanostructures representing nodes through the base complementary relationship of the sticky end groups, and this base complementary relationship of the sticky end groups represents the path relationship between nodes; mix and anneal a variety of different DNA nanocomputing elements constructed according to the preset conditions on a computing platform, dilute to a certain concentration, irradiate the resulting product with ultraviolet light, and then detect it with a detector to obtain the calculation result.

[0027] Preferably, the conditions for annealing are: starting temperature 45 °C, slowly annealing in a water bath to the end temperature 25 °C;

[0028] Preferably, each type of DNA nanocomputing element is mixed in equal proportion;

[0029] Preferably, dilute the obtained annealed product to 5 - 7 nM;

[0030] Preferably, after diluting the obtained annealed product, irradiate it with ultraviolet light with a wavelength of 254 nm, and the dose is 2.54 kJ / m 2 ;

[0031] Preferably, the detector is an atomic force microscope AFM.

[0032] As a preferred embodiment, in order to reduce the complexity of the path, multiple paths, that is, logical paths, existing in the Hamiltonian path problem relationship graph can share a physical path, and the physical path is the base complementary relationship of the sticky end group. This method is called the reuse of the physical path. This kind of reuse can reduce the workload of DNA sequence encoding of the sticky end group and solve complex path problems with fewer physical paths;

[0033] As a preferred embodiment, to solve larger-scale path problems, for example, when there are three or more path connection relationships, that is, logical paths, on a node, node splitting can be used to ensure that the physical path complexity of each node does not increase with the problem scale. The specific node splitting is as follows: construct and select two or more DNA nanocomputing elements with the same labeled node information but different encoded path information, and split the different logical paths contained in the same node.

[0034] As a preferred embodiment, the parity check method is adopted to ensure the accuracy of node information. Specifically, the parity check method is as follows: The specific sites on the rectangular two-dimensional DNA nanostructure for capturing the labeled structure are called information bits. Each rectangular two-dimensional DNA nanostructure is provided with n information bits, where n is a positive integer and an odd number; One more parity bit is added after the n-bit information bits to ensure that the number of 1s in all data is odd, which is the correct data, and the number of 1s is even, which is the wrong data. For example, for the Hamiltonian path problem of 7 nodes, the nodes are encoded by using three information bits plus one parity bit. The selection of the sites only needs to ensure that the high and low bits of the data reading are not incorrect. To further ensure the accuracy of the data, a case where all information bits are 0 is excluded. Therefore, all information bits start from 1.

[0035] In a third aspect, the present invention further provides a computing device based on the above DNA nanocomputing element. The computing device is used to solve the Hamiltonian problem. The device includes:

[0036] A programming module, configured to obtain preset conditions according to the Hamiltonian path problem to be solved, and perform DNA sequence encoding on the sticky ends of the rectangular two-dimensional DNA nanostructure according to the preset conditions and use a labeled structure to label the rectangular two-dimensional DNA nanostructure;

[0037] The DNA nanocomputing element database: used to store the DNA nanocomputing elements containing different coding information;

[0038] The computing platform, connected to the DNA nanocomputing element database and the detector, and providing appropriate reaction conditions for mixing and reacting the rectangular two-dimensional DNA nanostructure on the computing platform to obtain a reaction product.

[0039] Beneficial effects:

[0040] A class of DNA nanocomputing elements provided by the present invention utilize DNA nanostructures as DNA computing elements. The designed DNA nanostructures have uniform sizes, the reorganized programmable sites provide precise node site designs, and the rich extensible side chains can provide stable and reliable physical connection relationships. In the present invention, 15-nt sticky ends are grafted onto the two short sides of the rectangular two-dimensional DNA nanostructure, enabling the autonomous selection of a path for connection between rectangular plates. If connectable paths are still exposed at both ends of the rectangular plate, then it will continue to select and connect the corresponding paths until all the DNA nanocomputing elements are consumed. Based on a class of DNA nanocomputing elements of the present invention, the rectangular origami plates with connection relationships are coaxial. When applied to solve the Hamiltonian path problem, it can ensure that the polymer formed by connecting multiple rectangular origami plates still has good rigidity, thereby ensuring that the calculation results can be stored in the correct order without errors for subsequent observation of the calculation results. The DNA nanocomputing elements provided by the present invention have parallel computing capabilities and storage capabilities.

[0041] The rectangular two-dimensional DNA nanostructure mainly used in the present invention has good biocompatibility, and other required chemical and biological materials are also non-toxic to the human body. BRIEF DESCRIPTION OF THE DRAWINGS

[0042] Figure 1 Schematic diagram of the cross-arrangement of two sets of sticky end groups on the same rectangular two-dimensional DNA nanostructure of the present invention by splitting within the same group;

[0043] Figure 2 Schematic diagram of node information encoding in the specific embodiment of the present invention;

[0044] Figure 3a Schematic diagram of the physical path encoding of 7 nodes in the specific embodiment of the present invention;

[0045] Figure 3b Based on Figure 3a Schematic diagram of the calculation result finally obtained based on the path encoding;

[0046] Figure 4 Morphology characterization diagram of the DNA nanostructure constructed in the specific embodiment of the present invention;

[0047] Figure 5 Morphology characterization diagram of a partial rectangular plate DNA nanocomputing element containing node information;

[0048] Figure 6 Schematic diagram of the 7-node Hamiltonian path problem solved in the specific embodiment of the present invention;

[0049] Figure 7a Morphology characterization diagram of the rectangular plate polymer after irradiation in the specific embodiment of the present invention;

[0050] Figure 7b This is a characterization diagram of the correct results in the rectangular flat polymer in the specific embodiment of the present invention. Specific embodiment

[0051] To further illustrate the technical means and effects adopted by the present invention, the present invention will be further described below in conjunction with embodiments and drawings. It can be understood that the specific embodiments described herein are only used to explain the present invention, rather than limiting the present invention.

[0052] Description of raw materials and reagents used in the following preparation process and examples

[0053] 1. Tris (tris(hydroxymethyl)aminomethane), disodium ethylenediaminetetraacetate (EDTA), magnesium acetate, glacial acetic acid, and boric acid were all purchased from Sinopharm Chemical Reagent Co., Ltd.; 100 kDa ultrafiltration tubes were purchased from Pall China Co., Ltd.; the experimental water used was all ultrapure water.

[0054] 2. M13mp18 phage genomic DNA was purchased from BioDee Bio-Technology Co., Ltd., and its product number and specification were B3003-50 pmol; DNA staple strands, functional DNA strands, biotin-ssDNA strands, and streptavidin were all purchased from Shanghai Sangon Biotech Co., Ltd.

[0055] For those not specifying specific techniques or conditions in the examples, they shall be carried out according to the techniques or conditions described in the literature in this field or according to the product specifications. For those reagents or instruments without indicating the manufacturer, they are all conventional products that can be obtained through regular channels.

[0056] Specifically taking the solution of the path problem as shown in Figure 6 as an example, the preparation process of the DNA nanocomputing element described in the present invention and its application in solving the Hamiltonian path problem are specifically described;

[0057] According to the Hamiltonian path problem of 7 nodes as shown in Figure 6 obtain preset conditions, encode the DNA sequences of the sticky ends of the rectangular two-dimensional DNA nanostructure according to the preset conditions, and label the rectangular two-dimensional DNA nanostructure with a labeling structure to construct a variety of different DNA nanocomputing elements; the preset conditions of this embodiment include information of 7 nodes and the path relationships between each node;

[0058] Among them, a marking structure is adopted to mark the rectangular two-dimensional DNA nanostructure to represent node information. Specifically, in this embodiment, 12 DNA capture strands are designed to be anchored on the surface of the DNA nanostructure, and 3 strands capture one biotin-ssDNA molecule. They are Site 1: 148, 149, 125; Site 2: 140, 141, 118; Site 3: 10, 35, 36; Site 4: 19, 43, 44. Each rectangular plate has these four capture sites for biotin-ssDNA. Different nodes are encoded by controlling the addition of biotin-ssDNA. Among them, Sites 1-3 are information bits, and Site 1 is the low bit, Site 3 is the high bit, and Site 4 is the parity bit. For example, for node 0, the four-bit binary representation is 1000, and for node 2, it is 1101. The value of the current site is represented by the presence or absence of SA. If there is SA, it is represented as 1, and if there is no SA, it is represented as 0, as Figure 2 shown; the number of the biotin-SA complexes is 4. The nucleotide sequences of the DNA capture strands are as shown in SEQ ID NO:1-12, and the nucleotide sequence of biotin-ssDNA is as shown in SEQ ID NO:13;

[0059] The sticky end groups of the rectangular two-dimensional DNA nanostructure are encoded with DNA sequences, and the rectangular two-dimensional DNA nanostructures representing different nodes are matched and connected through the base complementary relationship of the sticky end groups. This base complementary relationship of the sticky end groups represents the path relationship between nodes. The node information and path information are encoded by means of physical path multiplexing and node splitting: Specifically, as Figure 3a shown, the 7 nodes are respectively Node 0, Node 1, Node 2, Node 3, Node 4, Node 5, and Node 6; Node 0 has three out-degrees. Node 0 is copied into Node 01 and Node 02, and then the paths 0→1 and 0→3 are assigned to Node 01, and the path 0→6 is assigned to Node 02. At the same time, the node coding information of Node 01 and Node 02 is the same. In fact, Node 0 has two origamis with different paths. Combining with the 7-node relationship diagram, 5 groups of physical paths are specifically designed. Each group of physical paths contains an in-degree and an out-degree, that is, it contains two groups of sticky end groups with complementary relationships. Therefore, the 5 groups of physical paths contain a total of 10 groups of sticky end groups. Each group of sticky end groups extends 9 sticky ends on the short side of the rectangular plate DNA nanostructure. It should be noted that in order to simplify the schematic diagram, Figure 3aIn the same group, each group of sticky ends is represented by a thick line for every 3 sticky die-forged chains. The thick lines with the same linear markings on the same side belong to a group of sticky ends. To form this sticky end structure, the specifically designed sticky DNA strand sequences are shown in SEQ ID NO: 14 - 73. The corresponding relationships of the sticky DNA strand sequences for forming 5 physical paths and a total of 10 groups of sticky ends are shown in the following table:

[0060]

[0061] According to the above preset conditions, respectively prepare Figure 3a A total of 10 rectangular flat DNA nano-computing elements containing different node information and path information as shown. Each rectangular flat DNA nano-computing element is operated according to the following steps:

[0062] (1) Mix the M13mp18 template strand, non-functional DNA staple strands, and DNA functional strands as shown; among them, the DNA sequence of the M13mp18 template strand is provided in GenBank: X02513.1, and the non-functional DNA staple strands refer to the staple strands used in the rectangular two-dimensional DNA nanostructure in the literature "Folding DNA to create nanoscale shapes and patterns. Nature, 2006, 440, 297 - 302"; the DNA functional strands include the above DNA capture strands and sticky DNA strands. The final concentrations of the template strand and other DNA short strands are 5 nM and 50 nM respectively;

[0063] (2) Use a gradient PCR instrument to slowly anneal the mixture. The annealing conditions are: starting temperature 95 °C, holding for 3 min, ending temperature 25 °C, with a 1 °C gradient for each step, and staying at each gradient for 100 s to obtain a rectangular two-dimensional DNA nanostructure;

[0064] (3) After the annealing program is completed, take out the rectangular two-dimensional DNA nanostructure sample and centrifuge it with a 100 kDa centrifuge tube to remove the excess DNA short strands; the centrifugation conditions are: add 300 μL of 1×TAE-Mg 2+ buffer to 100 μL of the sample, and centrifuge at 3000 rcf / min for 10 min, repeat centrifugation 3 times; the finally collected sample is observed for the morphology of the lamellar structure by atomic force microscopy (AFM). The results are as Figure 4 shown. The constructed DNA nanostructure is in a rectangular lamellar structure. The AFM characterization results show that the rectangular two-dimensional DNA nanostructure is about 90 nm long and about 60 nm wide, presenting a regular rectangular structure;

[0065] (4) Mix the purified rectangular two-dimensional DNA nanostructure sample and the biotin-ssDNA complex evenly at a molar ratio of 1:50, place them in a gradient PCR instrument, and slowly cool from 37 °C to 25 °C, with each 1 °C as a gradient and a residence time of 10 min for each gradient.

[0066] (5) After the annealing program is completed, take out the rectangular two-dimensional DNA nanostructure sample linked with biotin, centrifuge and separate it with a 100 kDa centrifuge tube to remove the excessive biotin-ssDNA short chains;

[0067] (6) Mix the DNA sample linked with biotin and SA at a molar ratio of 1:10 per unit site and anneal them. The annealing conditions are: keep at a constant temperature of 37 °C for 1 h.

[0068] (7) Then centrifuge with a 100 kDa centrifuge tube to remove the excess SA to obtain a purified rectangular flat DNA nanocomputing element encoding node and path information; use AFM to perform morphological characterization on the rectangular flat DNA nanocomputing element, and the results are as Figure 5 shown. There is corresponding node information on each rectangular flat DNA nanostructure, presenting regular site information. The node information of nodes 0, 2, 4, and 6 is shown in the figure.

[0069] (8) Mix the rectangular flat DNA nanocomputing elements with different encoded information evenly at an equimolar ratio, place them in a water bath for annealing, and slowly cool from 45 °C to 25 °C. After the annealing program is completed, dilute the obtained annealing product to 5 - 7 nM, and then irradiate it with ultraviolet light with a wavelength of 254 nm, and the dose is 2.54 kJ / m 2 . Use AFM for morphological characterization, and the results are as Figure 7a shown. The irradiated rectangular flat polymer will not be distorted anymore compared with the non-irradiated sample, and the whole polymer has a common axis; as Figure 7b shown is the characterization diagram of the correct result in the rectangular flat polymer, that is, corresponding to Figure 3b shown is the calculation result finally obtained based on Figure 3a the path encoding; all nodes are traversed once and only once, and the Hamiltonian solution is 0123456. It is observed that in some heptamers, each rectangular flat represents a different node and corresponds one by one to the nodes and paths in the figure. At the same time, other possible paths existing in the relationship diagram are also observed.

[0070] The applicant declares that the detailed method of the present invention is illustrated by the above embodiments, but the present invention is not limited to the above detailed method, that is, it does not mean that the present invention must rely on the above detailed method to be implemented. Those skilled in the art should understand that any improvement to the present invention, the equivalent substitution of each raw material of the product of the present invention, the addition of auxiliary components, the selection of specific methods, etc. all fall within the protection scope and the disclosure scope of the present invention.

Claims

1. A class of DNA nanocomputing elements, characterized in that, The DNA nanocomputing element is composed of a rectangular two-dimensional DNA nanostructure and a labeling structure; The rectangular two-dimensional DNA nanostructure is assembled from a DNA template strand and DNA staple strands; using the DNA template strand as the main chain and an excessive amount of DNA staple strands as the auxiliary chains, the rectangular two-dimensional DNA nanostructure is assembled by the hybridization and complementarity of the main chain and the auxiliary chains at specific positions; as the auxiliary chains, some of the DNA staple strands at specific positions are selected as programmable auxiliary chains, i.e., functional DNA strands, and the remaining DNA staple strands are non-functional DNA staple strands; The sticky DNA strands are part of the functional DNA strands, and after assembling the rectangular two-dimensional DNA nanostructure, the sticky DNA strands can extend one or two groups of sticky end groups on one or both of the two short sides of the main body, and each group of sticky end groups consists of at least 2 sticky end strands evenly distributed on the short side of the main body of the rectangular two-dimensional DNA nanostructure, and the base sequences of all the sticky end strands within the same group of sticky end groups are different; the left side of the rectangular two-dimensional DNA nanostructure is defined as the in-degree of the node, and the right side is defined as the out-degree of the node; The DNA capture strands are another part of the functional DNA strands, and after assembling, the DNA capture strands can extend at least one capture sequence on the surface of the main body of the rectangular two-dimensional DNA nanostructure, and the capture sequence is used to capture the labeling structure; The labeling structure contains a nucleotide sequence complementary to the capture sequence.

2. The class of DNA nanocomputing elements according to claim 1, characterized in that, The DNA template strand is the M13mp18 phage genomic DNA sequence provided in GenBank: X02513.1; the size of the rectangular two-dimensional DNA nanostructure is set to be 90 nm in length and 60 nm in width; when setting two groups of sticky end groups in the same direction, multiple sticky end strands set in the same direction are arranged in a split and cross-arranged manner in two groups; the capture sequence is a 15-nt poly-A or poly-T nucleotide sequence added at the 5'-end of the DNA staple strand.

3. The class of DNA nanocomputing elements according to claim 1, characterized in that, A total of 12 capture sequences are set on the surface of the rectangular two-dimensional DNA nanostructure to anchor the DNA nanostructure, and every 3 capture sequences capture one labeling structure; the labeling structure is biotin-streptavidin, and the biotin-streptavidin is a complex formed by the reaction of a biotin-ssDNA complex and streptavidin SA.

4. A class of DNA nanocomputing elements as claimed in claim 1, characterized in that, Each group of sticky end groups consists of 9 sticky end strands evenly distributed on the short side of the main body of the rectangular two-dimensional DNA nanostructure.

5. The preparation method of a class of DNA nanocomputing elements according to claim 1, characterized in that, The preparation method includes the following steps: Step (1): Mix the DNA template strand, non-functional DNA staple strands, and DNA functional strands in a buffer solution in proportion, anneal, and ultrafilter and purify to obtain the rectangular two-dimensional DNA nanostructure; Step (2): Mix the annealing product obtained in Step (1) with the biotin-ssDNA complex in proportion and then anneal; Step (3): Mix the annealing product obtained in Step (2) with SA in proportion and then anneal; Step (4): Ultrafilter and purify the annealing product obtained in Step (3) to replace the buffer solution to obtain the DNA nanocomputing element.

6. Application of a class of DNA nanocomputing elements described in any one of claims 1-4 in solving the Hamiltonian path problem. The specific application includes the following steps: obtaining preset conditions according to the Hamiltonian path problem to be solved, encoding the sticky ends of the rectangular two-dimensional DNA nanostructure with DNA sequences according to the preset conditions, and labeling the rectangular two-dimensional DNA nanostructure with a labeling structure to construct a variety of different DNA nanocomputing elements; the preset conditions include the names of each node and the path relationships between each node, where labeling the rectangular two-dimensional DNA nanostructure with a labeling structure represents node information; encoding the sticky end groups of the rectangular two-dimensional DNA nanostructure with DNA sequences, and matching and connecting the rectangular two-dimensional DNA nanostructures representing different nodes through the base complementary relationship of the sticky end groups, and this base complementary relationship of the sticky end groups represents the path relationship between nodes; detecting the product obtained by subjecting the variety of different DNA nanocomputing elements constructed according to the preset conditions to hybrid annealing on a computing platform, diluting to a certain concentration, and then irradiating with ultraviolet light using a detector to obtain the calculation result.

7. The application according to claim 6, characterized in that, The path relationship between different nodes, that is, the logical path, can share a physical path, and the physical path is the base complementary relationship of the sticky end groups.

8. The application according to claim 6, characterized in that, When there are three or more logical paths on a node, the method of node splitting is used to encode multiple logical paths of each node. The specific method of node splitting is as follows: constructing and selecting two or more DNA nanocomputing elements with the same labeled node information but different encoded path information, and splitting the different logical paths contained in the same node.

9. The application according to claim 6, characterized in that, The parity check method is used to ensure the accuracy of node information. Specifically, the parity check method is as follows: the specific site on the rectangular two-dimensional DNA nanostructure used to capture the labeling structure is called the information bit, and each rectangular two-dimensional DNA nanostructure is provided with n information bits, where n is a positive integer and an odd number; adding one more parity bit after the n-bit information bit to ensure that the number of 1s in all data is an odd number, which is the correct data, and an even number of 1s is the wrong data.

10. A computing device based on the DNA nanocomputing element described in any one of claims 1-4. The computing device is used to solve the Hamiltonian problem, and the device includes: A programming module for obtaining preset conditions according to the Hamiltonian path problem to be solved, encoding the sticky ends of the rectangular two-dimensional DNA nanostructure with DNA sequences according to the preset conditions, and labeling the rectangular two-dimensional DNA nanostructure with a labeling structure; The DNA nanocomputing element database: for storing the DNA nanocomputing elements containing different encoded information; The computing platform, connected to the DNA nanocomputing element database and the detector, and providing appropriate reaction conditions for mixing and reacting the rectangular two-dimensional DNA nanostructure on the computing platform to obtain a reaction product.

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