SARS-CoV-2 spike glycoprotein-binding nucleic acid molecule, sensor for detecting SARS-CoV-2, SARS-CoV-2 detection reagent, method for detecting SARS-CoV-2, and inactivator for SARS-CoV-2 virus

Abstract

Provided is a binding nucleic acid molecule that can specifically bind to SARS-CoV-2 spike glycoprotein, and that can bind strongly and is released slowly.　This SARS-CoV-2 spike glycoprotein-binding nucleic acid molecule is characterized by containing a polynucleotide of either (a) or (b). (a) A polynucleotide comprising a base sequence of SEQ ID NOs: 1-7 or a partial sequence of a base sequence of SEQ ID NOs: 1-7. (b) A polynucleotide comprising a base sequence having at least 80% identity with the base sequence of (a), and binding to the SARS-CoV-2 spike glycoprotein.

Description

[Technical field]

[0001] The present invention relates to a SARS-CoV-2 spike glycoprotein-binding nucleic acid molecule, a sensor for detecting SARS-CoV-2, a reagent for detecting SARS-CoV-2, a method for detecting SARS-CoV-2, and an agent for inactivating the SARS-CoV-2 virus. [Background technology]

[0002] Recently, the global pandemic caused by the novel coronavirus SARS-CoV-2 (also known as 2019-nCoV) has become a problem, and efforts are underway to clarify the mechanism of infection by SARS-CoV-2 and to develop treatments and vaccines.

[0003] Therefore, in order to detect SARS-CoV-2, attempts have been made to obtain aptamers that target proteins that constitute SARS-CoV-2 (Non-Patent Document 1). [Prior art documents] [Non-patent literature]

[0004] [Non-Patent Document 1] Song Y et al, Anal. Chem. (2020), 92, 9895-9900. Summary of the Invention [Problem to be solved by the invention]

[0005] However, there is a demand for aptamers that can specifically bind to SARS-CoV-2, have high binding affinity, and exhibit slow dissociation.

[0006] Therefore, the present invention aims to provide a binding nucleic acid molecule that can specifically bind to the SARS-CoV-2 spike glycoprotein, which is involved in binding to angiotensin-converting enzyme 2 (ACE2), the target of SARS-CoV-2, and has high binding affinity and slow dissociation. [Means for solving the problem]

[0007] The SARS-CoV-2 spike glycoprotein-binding nucleic acid molecule (hereinafter also referred to as "nucleic acid molecule") of the present invention is characterized in that it comprises any one of the following polynucleotides (a), (b), (c), and (d): (a) a polynucleotide consisting of the nucleotide sequence of any one of SEQ ID NOs: 1 to 7 or a partial sequence of the nucleotide sequence of any one of SEQ ID NOs: 1 to 7; (b) a polynucleotide having a nucleotide sequence having 80% or more identity to the nucleotide sequence of (a), and binding to the SARS-CoV-2 spike glycoprotein; (c) a polynucleotide that has a nucleotide sequence complementary to a polynucleotide that hybridizes under stringent conditions to the polynucleotide having the nucleotide sequence of (a), and that binds to the SARS-CoV-2 spike glycoprotein; (d) a polynucleotide that is composed of the base sequence of (a) in which one or several bases are deleted, substituted, inserted and / or added, and that binds to the SARS-CoV-2 spike glycoprotein; SEQ ID NO: 1: GGTATGTCTCCGCCACTGAAATCCG T GCC T AA T C T CACCCCACGGAA TT CA T GGCAAAGCCGAGG T G T C TT G T A TT C; SEQ ID NO: 2: GGTATGTCTCCGCCACTGAAATC T AA T C T CACA TT G T AAGCAAAGGAGAA T AAGCAAGCCGAGG T G T C TT G T A TT C; Phone 3:GGTATGTCTCCGCCACTGAAATCCC T GACCGC T ACKNOWLEDGMENTS T C T CAG T GCAGA T GCAAAGCCGAGG T G T C TT G T A TT C; Phone 4:GGTATGTCTCCGCCACTGAAATC TT G T CCCC T SECOND T AGG TT CCGAC T GACAAG T GCAAAGCCGAGG T G T C TT G T A TT C; Photo 5:GGTTTAGCCCTCGACTCCAAATG T GCAC T C T CCCGG T A T CCC T SECOND T C T CACCCGA T ACCG TT THOUGH T G T GC T GCCAA T AC; Page 6:GGTTTAGCCCTCGACTCCAAATGACA T GAGCCAGG T GCA T C TT GAACG T CA T AGA T ACCG TT THOUGH T G T GC T GCCAA T AC; Photo 7:GGTTTAGCCCTCGACTCCAAATGT GC T GA T AC T CGG T A T CCC T AA T C T CACCCGA T ACCG TT GA T G T GC T GCCAA T AC.

[0008] The sensor for detecting SARS-CoV-2 of the present invention is characterized by containing the SARS-CoV-2 spike glycoprotein-binding nucleic acid molecule of the present invention.

[0009] The SARS-CoV-2 detection reagent of the present invention is characterized by containing the SARS-CoV-2 spike glycoprotein-binding nucleic acid molecule of the present invention.

[0010] The method for detecting SARS-CoV-2 of the present invention includes the steps of contacting a sample with a nucleic acid molecule and detecting the SARS-CoV-2 spike glycoprotein in the sample, the nucleic acid molecule is the SARS-CoV-2 spike glycoprotein-binding nucleic acid molecule of the invention, The detection step is characterized in that the SARS-CoV-2 spike glycoprotein in the sample is bound to the nucleic acid molecule, and SARS-CoV-2 in the sample is detected through the binding. Effect of the Invention

[0011] The SARS-CoV-2 spike glycoprotein-binding nucleic acid molecule of the present invention can specifically bind to the SARS-CoV-2 spike glycoprotein, has high binding strength, and exhibits slow dissociation, and therefore, the SARS-CoV-2 spike glycoprotein-binding nucleic acid molecule of the present invention can detect SARS-CoV-2 with high accuracy based on the presence or absence of binding to the SARS-CoV-2 spike glycoprotein in a sample. [Brief description of the drawings]

[0012]

Figure 1

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Figure 8

[0013] (1) SARS-CoV-2 spike glycoprotein-binding nucleic acid molecule The SARS-CoV-2 spike glycoprotein-binding nucleic acid molecule of the present invention is characterized in that it comprises any one of the following polynucleotides (a), (b), (c), and (d): (a) a polynucleotide consisting of the nucleotide sequence of any one of SEQ ID NOs: 1 to 7 or a partial sequence of the nucleotide sequence of any one of SEQ ID NOs: 1 to 7; (b) a polynucleotide having a nucleotide sequence having 80% or more identity to the nucleotide sequence of (a), and binding to the SARS-CoV-2 spike glycoprotein; (c) a polynucleotide that has a nucleotide sequence complementary to a polynucleotide that hybridizes under stringent conditions to the polynucleotide having the nucleotide sequence of (a), and that binds to the SARS-CoV-2 spike glycoprotein; (d) a polynucleotide that is composed of the base sequence of (a) in which one or several bases are deleted, substituted, inserted and / or added, and that binds to the SARS-CoV-2 spike glycoprotein;

[0014] In the present invention, "SARS-CoV-2" means severe acute respiratory syndrome coronavirus 2. Infection caused by SARS-CoV-2 is also called COVID-19. SARS-CoV-2 is a virus classified in the genus Betacoronavirus. Coronaviruses including SARS-CoV-2 contain an envelope composed of an envelope protein, a membrane protein, a spike glycoprotein, etc., and a single-stranded RNA, which is a genomic RNA, is covered by the envelope. The amino acids on the surface of the spike glycoprotein may or may not be glycosylated.

[0015] As described above, the nucleic acid molecule of the present invention can bind to the SARS-CoV-2 spike glycoprotein, which is a protein constituting SARS-CoV-2. The SARS-CoV-2 spike glycoprotein is involved in binding to angiotensin-converting enzyme 2 (ACE2), which is the target of SARS-CoV-2. A schematic diagram of the sequence of the SARS-CoV-2 spike glycoprotein is shown in FIG. 7. The nucleic acid molecule of the present invention may bind to at least one of the RBD domain, S1+S2 domain, S1 domain, and a trimer of the S1+S2 domain (also called S-trimer), which constitute the SARS-CoV-2 spike glycoprotein. It may also bind to all of these, or to other regions of the SARS-CoV-2 spike glycoprotein. The binding affinity of the nucleic acid molecule of the present invention to the RBD domain, S1+S2 domain, S1 domain, and trimer can be confirmed by using, for example, the four types of commercially available reagents shown in Table 1 described in Example 1 below as targets. The nucleic acid molecule of the present invention is also referred to hereinafter as an aptamer, for example.

[0016] Information on the amino acid sequence of the SARS-CoV-2 spike glycoprotein includes, for example, the following amino acid sequence (SEQ ID NO: 15) registered under the accession number P0DTC2 in UniProt (http: / / www.uniprot.org / ). The domains of the SARS-CoV-2 spike glycoprotein are represented by the amino acid sequence numbers of the accession number P0DTC2, RBD domain: 319-541 (SEQ ID NO: 16), S1+S2 domain: 13-1273 (SEQ ID NO: 17), and S1 domain: 13-685 (SEQ ID NO: 18). The SARS-CoV-2 spike glycoprotein and each of its domains may have one or more mutations (e.g., deletions, substitutions, insertions, and / or additions) introduced into each amino acid sequence within the scope of binding to the nucleic acid molecule of the present invention.

[0017] SARS-CoV-2 spike glycoprotein (SEQ ID NO: 15)

[0018] RBD domain of SARS-CoV-2 spike glycoprotein (SEQ ID NO: 16) RVQPTESIVRFPNITNLCPFGEVFNATRFASVYAWNRKRISNCVADYSVLYNSASFSTFKCYGVSPTKLNDLCFTNVYADSFVIRGDEVRQIAPGQTGKIADYNYKLPDDF TGCVIAWNSNNLDSKVGGNYNYLYRLFRKSNLKPFERDISTEIYQAGSTPCNGVEGFNCYFPLQSYGFQPTNGVGYQPYRVVVLSFELLHAPATVCGPKKSTNLVKNKCVNF

[0019] S1+S2 domain of SARS-CoV-2 spike glycoprotein (SEQ ID NO: 17)

[0020] S1 domain of SARS-CoV-2 spike glycoprotein (SEQ ID NO: 18) SQCVNLTTRTQLPPAYTNSFTRGVYYPDKVFRSSVLHSTQDLFLPFFSNVTWFHAIHVSGTNGTKRFDNPVLPFNDGVYFASTEKSNIIRGWIFGTTLDSKTQSLLIVNNATNVVIKVCEFQFCNDPFLGVYYHKNNKSWMESEFRVYSSANNCTFEYVSQPFLMDLE GKQGNFKNLREFVFKNIDGYFKIYSKHTPINLVRDLPQGFSALEPLVDLPIGINITRFQTLLALHRSYLTPGDSSSGWTAGAAAYYVGYLQPRTFLLKYNENGTITDAVDCALDPLSETKCTLKSFTVEKGIYQTSNFRVQPTESIVRFPNITNLCPFGEVFNATRFA SVYAWNRKRISNCVADYSVLYNSASFSTFKCYGVSPTKLNDLCFTNVYADSFVIRGDEVRQIAPGQTGKIADYNYKLPDDFTGCVIAWNSNNLDSKVGGNYNYLYRLFRKSNLKPFERDISTEIYQAGSTPCNGVEGFNCYFPLQSYGFQPTNGVGYQPYRVVVLSFE LLHAPATVCGPKKSTNLVKNKCVNFNFNGLTGTGVLTESNKKFLPFQQFGRDIADTTDAVRDPQTLEILDITPCSFGGVSVITPGTNTSNQVAVLYQDVNCTEVPVAIHADQLTPTWRVYSTGSNVFQTRAGCLIGAEHVNNSYECDIPIGAGICASYQTQTNSPRRAR

[0021] In the present invention, "binding to SARS-CoV-2 spike glycoprotein" also means, for example, "having binding ability to SARS-CoV-2 spike glycoprotein" or "having binding activity to SARS-CoV-2 spike glycoprotein". The binding between the nucleic acid molecule of the present invention and the SARS-CoV-2 spike glycoprotein can be determined, for example, by surface plasmon resonance (SPR) analysis. For the analysis, for example, ProteON (trade name, BioRad) can be used. Since the nucleic acid molecule of the present invention binds to the SARS-CoV-2 spike glycoprotein, it can be used, for example, to detect SARS-CoV-2.

[0022] The nucleic acid molecule of the present invention may be, for example, a molecule that consists of the polynucleotide of (a) or (b), or a molecule that contains the polynucleotide.The nucleic acid molecule of the present invention may be, for example, composed of DNA, RNA, or DNA and RNA.When the nucleic acid molecule of the present invention is composed of DNA, the nucleic acid molecule of the present invention may be, for example, a DNA molecule or a DNA aptamer.

[0023] The polynucleotide (a) may be, for example, a polynucleotide containing the base sequence of SEQ ID NO: 1 to 7, or may be a polynucleotide consisting of the base sequence of SEQ ID NO: 1 to 7. The polynucleotide (a) may be a nucleotide containing a partial sequence of the base sequence of SEQ ID NO: 1 to 7, or may be a polynucleotide consisting of the partial sequence. The partial sequence is not particularly limited, and may be, for example, a sequence in which at least one of the 5'-end and 3'-end sequences in the base sequence of SEQ ID NO: 1 to 7 is deleted, or a sequence in which the sequence in the middle region is deleted. The polynucleotides of SEQ ID NO: 1 to 7 are shown below. In addition, modified bases are underlined in SEQ ID NO: 1 to 7 and the base sequences of SEQ ID NO: 8 to 13 described later. Modified bases will be described later.

[0024] SARS-CoV-2 spike glycoprotein-binding nucleic acid molecule 1 (SEQ ID NO:1) GGTATGTCTCCGCCACTGAAATCCG T GCC T AA T C T CACCCCACGGAA TT CA T GGCAAAGCCGAGG T G T C TT G T A TT C SARS-CoV-2 spike glycoprotein-binding nucleic acid molecule 2 (SEQ ID NO:2) GGTATGTCTCGCCACTGAAATC T AA T C T CACA TT G T AAGCAAAGGAGAA T AAGCAAGCCGAGG T G T C TT G T A TT C SARS-CoV-2 spike glycoprotein-binding nucleic acid molecule 3 (SEQ ID NO:3) GGTATGTCTCGCCACTGAAATCCC T GACCGC T GACCAAA T C T CAG T GCAGA T GCAAAGCCGAGG T G T C TT G T A TT C SARS-CoV-2 spike glycoprotein-binding nucleic acid molecule 4 (SEQ ID NO: 4) GGTATGTCTCGCCACTGAAATC TT G T CCCC T AA T AGG TT CCGAC T GACAAG T GCAAAGCCGAGG T G T C TT G TA TT C SARS-CoV-2 Spike glycoprotein-binding nucleic acid molecule 5 (SEQ ID NO: 5) GGTTTAGCCCTCGACTCCAAATG T GCAC T C T CCCGG T A T CCC T AA T C T CACCCGA T ACCG TT GA T G T GC T GCCAA T AC SARS-CoV-2 Spike glycoprotein-binding nucleic acid molecule 6 (SEQ ID NO: 6) GGTTTAGCCCTCGACTCCAAATGACA T GAGCCAGG T GCA T C TT GAACG T CA T AGA T ACCG TT GA T G T GC T GCCAA T AC SARS-CoV-2 Spike glycoprotein-binding nucleic acid molecule 7 (SEQ ID NO: 7) GGTTTAGCCCTCGACTCCAAATG T GC T GA T AC T CGG T A T CCC T AA T C T CACCCGA T ACCG TT GA T G T GC T GCCAA T AC

[0025] The partial sequence is not particularly limited, and for example, a partial sequence of the base sequence of SEQ ID NO: 1 is the base sequence of SEQ ID NO: 8, a partial sequence of the base sequence of SEQ ID NO: 2 is the base sequence of SEQ ID NO: 9, a partial sequence of the base sequence of SEQ ID NO: 3 is the base sequence of SEQ ID NO: 10, a partial sequence of the base sequence of SEQ ID NO: 5 is the base sequence of SEQ ID NO: 11, a partial sequence of the base sequence of SEQ ID NO: 6 is the base sequence of SEQ ID NO: 12, or a partial sequence of the base sequence of SEQ ID NO: 7 is the base sequence of SEQ ID NO: 13.

[0026] SARS-CoV-2 spike glycoprotein-binding nucleic acid molecule 8 (SEQ ID NO:8) CCACTGAAATCCG T GCC T AA T C T CACCCCACGGAA TT CA T GG SARS-CoV-2 spike glycoprotein-binding nucleic acid molecule 9 (SEQ ID NO: 9) TCCGCCACTGAAATC T AA T C T CACA TT G T AAGCAAAGGAGAA T AA SARS-CoV-2 spike glycoprotein-binding nucleic acid molecule 10 (SEQ ID NO: 10) CCACTGAAATCCC T GACCGC T GACCAAA T C T CAG T GCAGA T SARS-CoV-2 spike glycoprotein-binding nucleic acid molecule 11 (SEQ ID NO: 11) TGTGCACTCTCCCGG T A T CCC T AA T C T CACCCGA T ACC SARS-CoV-2 spike glycoprotein-binding nucleic acid molecule 12 (SEQ ID NO: 12) TGACATGAGCCAGG T GCA T C TT GAACG T CA T AGA T ACCG TT GA T G T GC T G SARS-CoV-2 spike glycoprotein-binding nucleic acid molecule 13 (SEQ ID NO: 13) GCTGATACTCGG T A T CCC T AA T C T CACCCGA T ACCG

[0027] In (b), the "identity" is not particularly limited, and may be within the range in which the polynucleotide of (b) binds to the SARS-CoV-2 spike glycoprotein. The identity is, for example, 80% or more, preferably 85% or more, more preferably 90% or more, even more preferably 95% or more, 96% or more, 97% or more, particularly preferably 98% or more, and most preferably 99% or more. The identity can be calculated using default parameters, for example, using analysis software such as BLAST or FASTA (hereinafter the same).

[0028] The polynucleotide of (b) may be, for example, the polynucleotide of (b1) below. In this case, the nucleic acid molecule of the present invention may be, for example, a molecule consisting of the polynucleotide of (b1) above, or a molecule containing the polynucleotide. In (b1) below, "containing any of the base sequences of SEQ ID NOs: 8 to 13" can also be interpreted as "containing any of the base sequences of SEQ ID NOs: 8 to 13". (b1) A polynucleotide that has a base sequence having 80% or more identity to the base sequence of (a), contains any of the base sequences of SEQ ID NOs: 8 to 13, and binds to the SARS-CoV-2 spike glycoprotein.

[0029] The polynucleotide in the nucleic acid molecule of the present invention may be, for example, the polynucleotide (b2) below. In this case, the nucleic acid molecule of the present invention may be, for example, a molecule consisting of the polynucleotide (b2) above, or a molecule containing the polynucleotide. (b2) A polynucleotide that has a base sequence having 80% or more identity to the base sequence of (a), is capable of forming a secondary structure represented by any of the following formulas (2) to (14), and binds to the SARS-CoV-2 spike glycoprotein: [ka] [ka] [ka] [ka] [ka] [ka] [ka] [ka] [ka] [ka] [ka] [ka] [ka]

[0030] Specifically, the polynucleotide (b2) is a polynucleotide that consists of a base sequence having 80% or more identity to the base sequences of SEQ ID NOs: 1 to 13, is capable of forming the secondary structures represented by formulas (2) to (14), respectively, and binds to the SARS-CoV-2 spike glycoprotein.

[0031] In the polynucleotides (b1) and (b2) above, the identity can be determined by referring to the explanation for the identity in the polynucleotide (b), for example.

[0032] The correspondence between the base sequences of SEQ ID NOs: 1 to 13 and the secondary structures represented by the formulas (2) to (14) is shown below. A polynucleotide consisting of the base sequence of SEQ ID NO: 1 is capable of forming, for example, a secondary structure represented by the above formula (2). A polynucleotide consisting of the base sequence of SEQ ID NO: 2 is capable of forming, for example, the secondary structure represented by the above formula (3). A polynucleotide consisting of the base sequence of SEQ ID NO: 3 is capable of forming, for example, the secondary structure represented by the above formula (4). A polynucleotide consisting of the base sequence of SEQ ID NO: 4 is capable of forming, for example, the secondary structure represented by the above formula (5). A polynucleotide consisting of the base sequence of SEQ ID NO: 5 is capable of forming, for example, the secondary structure represented by the above formula (6). A polynucleotide consisting of the base sequence of SEQ ID NO: 6 is capable of forming, for example, the secondary structure represented by the above formula (7). A polynucleotide consisting of the base sequence of SEQ ID NO: 7 is capable of forming, for example, the secondary structure represented by the above formula (8). A polynucleotide consisting of the base sequence of SEQ ID NO: 8 is capable of forming, for example, the secondary structure represented by the above formula (9). A polynucleotide consisting of the base sequence of SEQ ID NO: 9 is capable of forming, for example, the secondary structure represented by the above formula (10). A polynucleotide consisting of the base sequence of SEQ ID NO: 10 is capable of forming, for example, the secondary structure represented by the above formula (11). A polynucleotide consisting of the base sequence of SEQ ID NO: 11 is capable of forming, for example, the secondary structure represented by the above formula (12). A polynucleotide consisting of the base sequence of SEQ ID NO: 12 is capable of forming, for example, the secondary structure represented by the above formula (13). A polynucleotide consisting of the base sequence of SEQ ID NO: 13 is capable of forming, for example, the secondary structure represented by the above formula (14).

[0033] Furthermore, the polynucleotide (b2) is specifically a polynucleotide that consists of a base sequence having 80% or more identity to the base sequences of SEQ ID NOs: 1 to 3 and 5 to 7, is capable of forming a secondary structure represented by formulas (9) to (14) corresponding to SEQ ID NOs: 8 to 13, which are partial sequences of the base sequences of SEQ ID NOs: 1 to 3 and 5 to 7, respectively, and binds to the SARS-CoV-2 spike glycoprotein.

[0034] In (b2), "capable of forming a secondary structure" means, for example, that the polynucleotide of (b2) is capable of forming a stem structure and a loop structure in the formula. The stem structure and the loop structure will be described later. In the stem structure, the opposing base pairs may be a combination of bases capable of forming hydrogen bonds. Examples of the combination of bases capable of forming hydrogen bonds include AT, TA, AU, UA, GC, and CG. The "secondary structure" may be, for example, a secondary structure that is actually formed, or a secondary structure obtained by simulation using known means.

[0035] The polynucleotide in the nucleic acid molecule of the present invention may be, for example, the polynucleotide (c) below. In this case, the nucleic acid molecule of the present invention may be, for example, a molecule consisting of the polynucleotide (c) below, or a molecule containing the polynucleotide (c) below. (c) a polynucleotide that has a nucleotide sequence complementary to a polynucleotide that hybridizes under stringent conditions to the polynucleotide having the nucleotide sequence of (a) and binds to the SARS-CoV-2 spike glycoprotein.

[0036] In the above (c), the "hybridizable polynucleotide" is, for example, a polynucleotide that is completely or partially complementary to the polynucleotide of the above (a), and may be capable of binding to the SARS-CoV-2 spike glycoprotein. The hybridization can be detected, for example, by various hybridization assays. The hybridization assay is not particularly limited, and may be, for example, a hybridization assay such as that described in "Molecular Cloning: A Laboratory Manual 2nd Edition" edited by Sambrook et al. nd Alternatively, the method described in, for example, "Cold Spring Harbor Laboratory Press (1989)" can be adopted.

[0037] In the above (c), the "stringent conditions" may be, for example, low stringency conditions, medium stringency conditions, or high stringency conditions. The "low stringency conditions" are, for example, 5×SSC, 5×Denhardt's solution, 0.5% SDS, 50% formamide, and 32° C. The "medium stringency conditions" are, for example, 5×SSC, 5×Denhardt's solution, 0.5% SDS, 50% formamide, and 42° C. The "high stringency conditions" are, for example, 5×SSC, 5×Denhardt's solution, 0.5% SDS, 50% formamide, and 50° C. Those skilled in the art can set the degree of stringency by appropriately selecting conditions such as temperature, salt concentration, probe concentration and length, ionic strength, and time. The "stringent conditions" are, for example, those described in the aforementioned "Molecular Cloning: A Laboratory Manual 2nd Edition" edited by Sambrook et al. nd Alternatively, the conditions described in, for example, "Cold Spring Harbor Laboratory Press (1989)" Ed. may be employed.

[0038] The polynucleotide in the nucleic acid molecule of the present invention may be, for example, the polynucleotide (d) below. In this case, the nucleic acid molecule of the present invention may be, for example, a molecule consisting of the polynucleotide (d) below or a molecule containing the polynucleotide (d) below. (d) a polynucleotide that binds to the SARS-CoV-2 spike glycoprotein and that is composed of a base sequence in which one or several bases are deleted, substituted, inserted and / or added in the base sequence of (a)

[0039] In (d), "one or several" may be within the range in which the polynucleotide of (d) binds to the SARS-CoV-2 spike glycoprotein. The "one or several" may be, for example, 1 to 16, 1 to 12, 1 to 10, 1 to 8, 1 to 7, 1 to 5, 1 to 3, 1 to 2, or 1 in the base sequence of (a). In the present invention, the numerical range of the number of bases and the number of sequences, etc., discloses, for example, all positive integers within that range. That is, for example, the description "1 to 5 bases" means the disclosure of all of "1, 2, 3, 4, and 5 bases" (hereinafter the same).

[0040] The polynucleotide of (d) may be, for example, the polynucleotide of (d1) below. In this case, the nucleic acid molecule of the present invention may be, for example, a molecule consisting of the polynucleotide of (d1) above, or a molecule containing the polynucleotide. In (d1) below, "containing any of the base sequences of SEQ ID NOs: 8 to 13" can also be interpreted as "containing any of the base sequences of SEQ ID NOs: 8 to 13." (d1) A polynucleotide that is a base sequence in which one or several bases are deleted, substituted, inserted and / or added in the base sequence of (a), contains any of the base sequences of SEQ ID NOs: 8 to 13, and binds to the SARS-CoV-2 spike glycoprotein.

[0041] The polynucleotide (d) may be, for example, the polynucleotide (d2) below. In this case, the nucleic acid molecule of the present invention may be, for example, a molecule consisting of the polynucleotide (d2) or a molecule containing the polynucleotide. (d2) A polynucleotide having a base sequence in which one or several bases are deleted, substituted, inserted and / or added in the base sequence of (a), capable of forming a secondary structure represented by any one of formulas (2) to (14), and capable of binding to the SARS-CoV-2 spike glycoprotein.

[0042] The "one or several" in the polynucleotides (d1) and (d2) above can refer to the same explanation as for the "one or several" in the polynucleotide (d) above.

[0043] In the nucleic acid molecule of the present invention, the constituent unit of the polynucleotide is, for example, a nucleotide residue, such as a deoxyribonucleotide residue or a ribonucleotide residue. As described below, the polynucleotide may be, for example, a DNA consisting of deoxyribonucleotide residues, a DNA containing deoxyribonucleotide residues and ribonucleotide residues, or may further contain non-nucleotide residues.

[0044] The nucleic acid molecule of the present invention may be, for example, a molecule consisting of any one of the polynucleotides (a) to (d) above, or may be a molecule containing any one of the polynucleotides (a) to (d) above. In the latter case, the nucleic acid molecule of the present invention may contain, for example, two or more of the polynucleotides (a) to (d) above, as described below. The two or more polynucleotides (a) to (d) above may have the same sequence or different sequences. In the latter case, the nucleic acid molecule of the present invention may further have, for example, a linker and / or an additional sequence. Here, the linker is, for example, a sequence between polynucleotides, and the additional sequence is, for example, a sequence added to the end.

[0045] For example, when the nucleic acid molecule of the present invention contains multiple polynucleotides, it is preferable that the multiple polynucleotide sequences are linked to form a single-stranded polynucleotide. For example, the multiple polynucleotide sequences may be directly linked to each other, or may be indirectly linked to each other via a linker. It is preferable that the polynucleotide sequences are directly or indirectly linked at each end. When the nucleic acid molecule of the present invention contains multiple polynucleotide sequences, the number of the sequences is not particularly limited, and is, for example, 2 or more, 2 to 20, 2 to 10, 2, or 3.

[0046] The length of the linker is not particularly limited, and may be, for example, 1 to 200 bases, 1 to 24 bases, 1 to 20 bases, 3 to 12 bases, or 5 to 9 bases. The constituent unit of the linker is, for example, a nucleotide residue, such as a deoxyribonucleotide residue or a ribonucleotide residue. The linker is not particularly limited, and may be, for example, a polynucleotide such as DNA consisting of deoxyribonucleotide residues or DNA containing ribonucleotide residues. Specific examples of the linker include polydeoxythymine (poly dT), polydeoxyadenine (poly dA), and poly dAdT, which is a repeat sequence of A and T, and preferably poly dT or poly dAdT.

[0047] In the nucleic acid molecule of the present invention, the polynucleotide is preferably a single-stranded polynucleotide. The single-stranded polynucleotide is preferably capable of forming, for example, a stem structure and a loop structure by self-annealing. The polynucleotide is preferably capable of forming, for example, a stem-loop structure, an internal loop structure, and / or a bulge structure.

[0048] The nucleic acid molecule of the present invention may be, for example, double-stranded. In the case of double-stranded, for example, one single-stranded polynucleotide comprises any one of the polynucleotides (a) to (d), and the other single-stranded polynucleotide is not limited. The other single-stranded polynucleotide may be, for example, a polynucleotide comprising a base sequence complementary to any one of the polynucleotides (a) to (d). When the nucleic acid molecule of the present invention is double-stranded, it is preferable to dissociate it into single-stranded polynucleotides by, for example, denaturation prior to use. In addition, it is preferable that the dissociated single-stranded polynucleotide of any one of the (a) to (d) forms a stem structure and a loop structure, for example, as described above.

[0049] In the present invention, "capable of forming a stem structure and a loop structure" includes, for example, the actual formation of a stem structure and a loop structure, as well as the ability to form a stem structure and a loop structure under certain conditions even if the stem structure and the loop structure are not formed. "Capable of forming a stem structure and a loop structure" includes, for example, both cases confirmed experimentally and cases predicted by simulations such as a computer.

[0050] The constituent unit of the nucleic acid molecule of the present invention is, for example, a nucleotide residue. The length of the nucleic acid molecule is not particularly limited. The lower limit of the length of the nucleic acid molecule is, for example, 15 bases, 35 bases, 55 bases, or 75 bases. The upper limit of the length of the nucleic acid molecule is, for example, 1000 bases, 200 bases, 100 bases, 90 bases, or 80 bases. The length of the nucleic acid molecule ranges, for example, from 15 to 1000 bases, from 35 to 200 bases, from 55 to 90 bases, or from 75 to 80 bases.

[0051] Examples of the nucleotide residue include deoxyribonucleotide residues and ribonucleotide residues. Examples of the nucleic acid molecule of the present invention include DNA composed only of deoxyribonucleotide residues, DNA containing one or several ribonucleotide residues, etc. In the latter case, "one or several" is not particularly limited, and may be, for example, 1 to 91, 1 to 30, 1 to 15, 1 to 7, 1 to 3, or 1 or 2 in the polynucleotide.

[0052] The polynucleotide may contain a natural base or a modified base as a base in a nucleotide residue. The natural base (non-artificial base) is not particularly limited, and examples thereof include a purine base having a purine skeleton, a pyrimidine base having a pyrimidine skeleton, and the like. The purine base is not particularly limited, and examples thereof include adenine (a) and guanine (g). The pyrimidine base is not particularly limited, and examples thereof include cytosine (c), thymine (t), uracil (u), and the like.

[0053] When the polynucleotide has the modified base, the site and the number are not particularly limited. When the nucleic acid molecule of the present invention has the modified base, for example, in the polynucleotides of SEQ ID NOs: 1 to 13, a part or all of the underlined thymine is a modified base. When the underlined thymine is a modified base, the modified base is preferably a modified thymine in which the thymine base is modified.

[0054] The modified base is, for example, a base modified with a modifying group. The base modified with the modifying group (modified base) is, for example, the natural base. Examples of the natural base include a purine base and a pyrimidine base. The modified base is not particularly limited, and examples thereof include a modified adenine, a modified guanine, a modified cytosine, a modified thymine, and a modified uracil.

[0055] The modified base may be, for example, a modified base directly modified with the modifying group, or a modified base indirectly modified with the modifying group. In the latter case, for example, the modified base may be modified with the modifying group via a linker. The linker is not particularly limited.

[0056] The modified site of the modified base by the modifying group is not particularly limited. When the base is a pyrimidine base, the modified site of the pyrimidine base may be, for example, the 5th and 6th positions of the pyrimidine backbone, and the 5th position is preferred. When the 5th position of the pyrimidine base is modified, since thymine has a methyl group at the 5th carbon, for example, the modifying group may be directly or indirectly bonded to the 5th carbon, or the modifying group may be directly or indirectly bonded to the carbon of the methyl group bonded to the 5th carbon. When "=O" is bonded to the 4th carbon in the pyrimidine backbone and a group other than "-CH3" or "-H" is bonded to the 5th carbon, it can be called a modified thymine or modified uracil.

[0057] The modified thymine base is a nucleotide residue represented by the following formula (1) (hereinafter, also referred to as "NG7"). [ka]

[0058] In this case, for example, the nucleotide triphosphate represented by the formula (1) can be used as a monomer molecule in the synthesis of the polynucleotide. In the synthesis of the polynucleotide, for example, the monomer molecule is bonded to another nucleotide triphosphate through a phosphodiester bond. The monomer can be produced by a known method, for example, see International Publication No. WO 2018 / 052063.

[0059] When the nucleotide residue represented by formula (1) is contained as the modified thymine base, the nucleotide residue containing the modified thymine base in the polynucleotide is represented, for example, by the following formula (1A). [ka]

[0060] In the polynucleotide, the underlined thymine base in the base sequence of SEQ ID NO: 1 to 13 is preferably a modified thymine base represented by the formula (1). In this case, it can also be said that the underlined nucleotide residue in the polynucleotide consisting of the base sequence of SEQ ID NO: 1 to 13 is a nucleotide residue represented by the formula (1A).

[0061] The polynucleotide may contain, for example, only one type of modified base, or may contain two or more types of modified bases.

[0062] The nucleic acid molecule of the present invention may contain, for example, a modified nucleotide. The modified nucleotide may be a nucleotide having the modified base described above, a nucleotide having a modified sugar in which the sugar residue is modified, or a nucleotide having both the modified base and the modified sugar.

[0063] The sugar residue is not particularly limited and may be, for example, a deoxyribose residue or a ribose residue. The modification site in the sugar residue is not particularly limited and may be, for example, the 2'-position or the 4'-position of the sugar residue, either one or both of which may be modified. The modification group in the modified sugar may be, for example, a methyl group, a fluoro group, an amino group, a thio group, etc.

[0064] In the modified nucleotide residue, when the base is a pyrimidine base, for example, the 2'-position and / or 4'-position of the sugar residue is preferably modified. Specific examples of the modified nucleotide residue include 2'-methylated uracil nucleotide residues, 2'-methylated cytosine nucleotide residues, 2'-fluorolated uracil nucleotide residues, 2'-fluorolated cytosine nucleotide residues, 2'-aminolated uracil nucleotide residues, 2'-aminolated cytosine nucleotide residues, 2'-thiolated uracil nucleotide residues, 2'-thiolated cytosine nucleotide residues, and the like, each of which is modified at the 2'-position of a deoxyribose residue or ribose residue.

[0065] The number of modified bases is not particularly limited. In the polynucleotide, the number of modified bases is, for example, 1 or more. In the polynucleotide, the number of modified bases is, for example, 1 to 80, 1 to 70, 1 to 50, 1 to 40, 1 to 30, 1 to 20, or 1 to 10, and all bases may be the modified bases. The number of modified bases may be, for example, the number of any one type of modified base, or the total number of two or more types of modified bases. The number of modified bases in the entire length of the nucleic acid molecule containing the polynucleotide is also not particularly limited, and is, for example, 1 to 80, 1 to 50, or 1 to 20, and is preferably the same as the above-mentioned range.

[0066] In the polynucleotide, the ratio of the modified bases is not particularly limited. The ratio of the modified bases is, for example, 1 / 100 or more, 1 / 40 or more, 1 / 20 or more, 1 / 10 or more, 1 / 4 or more, or 1 / 3 or more of the total number of bases in the polynucleotide. The ratio of the modified bases in the entire length of the nucleic acid molecule containing the polynucleotide is also not particularly limited and is the same as the above-mentioned range. Here, the total number of bases is, for example, the sum of the number of natural bases and the number of the modified bases in the polynucleotide. The ratio of the modified bases is expressed as a fraction, and the total number of bases and the number of modified bases that satisfy this are each positive integers.

[0067] When the modified base in the polynucleotide is the modified thymine, the number of the modified thymines is not particularly limited. In the polynucleotide, for example, natural thymine can be substituted with the modified thymine. In the polynucleotide, the number of the modified thymines is, for example, 1 or more. In the polynucleotide, the number of the modified thymines is, for example, 1 to 80, 1 to 70, 1 to 50, 1 to 40, 1 to 30, 1 to 20, or 1 to 10, and all the thymines may be the modified thymines.

[0068] In the polynucleotide, the ratio of the modified thymines is not particularly limited and is, for example, 1 / 100 or more, 1 / 40 or more, 1 / 20 or more, 1 / 10 or more, 1 / 4 or more, or 1 / 3 or more of the total number of the natural thymines and the modified thymines.

[0069] The nucleic acid molecule of the present invention may contain, for example, one or several artificial nucleic acid monomer residues. The "one or several" is not particularly limited, and may be, for example, 1 to 100, 1 to 50, 1 to 30, or 1 to 10 in the polynucleotide. Examples of the artificial nucleic acid monomer residue include PNA (peptide nucleic acid), LNA (locked nucleic acid), and ENA (2'-O, 4'-C-Ethylene-bridged Nucleic Acids). The nucleic acid in the monomer residue is, for example, the same as described above.

[0070] The nucleic acid molecule of the present invention is preferably, for example, nuclease resistant. The nucleic acid molecule of the present invention preferably has, for example, the modified nucleotide residue and / or the artificial nucleic acid monomer residue for nuclease resistance. For example, the nucleic acid molecule of the present invention may have several tens of kDa of PEG (polyethylene glycol) or deoxythymidine bound to the 5'-end or 3'-end for nuclease resistance.

[0071] The nucleic acid molecule of the present invention may further have, for example, an additional sequence. For example, the additional sequence is preferably bound to at least one of the 5'-end and 3'-end of the nucleic acid molecule, more preferably to the 3'-end. The additional sequence is not particularly limited. The length of the additional sequence is not particularly limited, and is, for example, 1 to 200 bases long, 1 to 50 bases long, 1 to 25 bases long, or 18 to 24 bases long. The constituent unit of the additional sequence is, for example, a nucleotide residue, such as a deoxyribonucleotide residue or a ribonucleotide residue. The additional sequence is not particularly limited, and is, for example, a polynucleotide such as DNA consisting of deoxyribonucleotide residues or DNA containing ribonucleotide residues. Specific examples of the additional sequence include polydT, polydA, and the like.

[0072] The nucleic acid molecule of the present invention can be used, for example, by immobilizing it on a carrier. The nucleic acid molecule of the present invention is preferably immobilized, for example, at either the 5'-end or the 3'-end, more preferably at the 3'-end. When immobilizing the nucleic acid molecule of the present invention, for example, the nucleic acid molecule may be directly or indirectly immobilized on the carrier. In the latter case, it is preferable to immobilize it via the additional sequence, for example.

[0073] The nucleic acid molecule of the present invention may further have, for example, a labeling substance, and specifically, the labeling substance may be bound to the nucleic acid molecule. The nucleic acid molecule bound to the labeling substance may be, for example, the nucleic acid sensor of the present invention. The labeling substance may be bound to at least one of the 5' end and the 3' end of the nucleic acid molecule. The labeling with the labeling substance may be, for example, binding or chemical modification. The labeling substance is not particularly limited, and examples thereof include enzymes, fluorescent substances, dyes, isotopes, drugs, toxins, and antibiotics. Examples of the enzyme include luciferase and NanoLuc luciferase. Examples of the fluorescent substance include fluorophores such as pyrene, TAMRA, fluorescein, Cy3 dye, Cy5 dye, FAM dye, rhodamine dye, Texas Red dye, JOE, MAX, HEX, and TYE, and examples of the dye include Alexa dyes such as Alexa488 and Alexa647. The labeling substance may be linked directly to the nucleic acid molecule, or indirectly via a linker. The linker is not particularly limited and may be, for example, a polynucleotide linker.

[0074] The method for producing the nucleic acid molecule of the present invention is not particularly limited, and the molecule can be synthesized by, for example, a nucleic acid synthesis method using chemical synthesis, a genetic engineering technique, or a known method.

[0075] As described above, the nucleic acid molecule of the present invention exhibits binding to the SARS-CoV-2 spike glycoprotein. Therefore, the use of the nucleic acid molecule of the present invention is not particularly limited as long as it utilizes the binding to the SARS-CoV-2 spike glycoprotein. The nucleic acid molecule of the present invention can be used in various methods, for example, in place of an antibody against the SARS-CoV-2 spike glycoprotein.

[0076] The nucleic acid molecule of the present invention can detect the SARS-CoV-2 spike glycoprotein. This allows detection of SARS-CoV-2. The detection method of the SARS-CoV-2 spike glycoprotein and SARS-CoV-2 is not particularly limited, and can be performed, for example, by detecting the binding between the SARS-CoV-2 spike glycoprotein and the nucleic acid molecule with reference to the detection method described below.

[0077] (2) SARS-CoV-2 detection sensor As described above, the detection sensor of the present invention is a sensor for detecting SARS-CoV-2, and is characterized by including the nucleic acid molecule of the present invention. The detection sensor of the present invention is not particularly limited as long as it includes the nucleic acid molecule of the present invention. By using the detection sensor of the present invention, for example, by binding the nucleic acid molecule to the SARS-CoV-2 spike glycoprotein, the SARS-CoV-2 can be detected as described above.

[0078] The detection sensor of the present invention further includes, for example, a carrier, and the nucleic acid molecule is disposed on the carrier. The nucleic acid molecule is preferably immobilized on the carrier. The nucleic acid molecule is immobilized on the carrier, for example, as described above. The method of using the detection sensor of the present invention is not particularly limited, and the nucleic acid molecule of the present invention and the detection method of the present invention can be used.

[0079] (3) Detection reagents and kits The detection reagent of the present invention is characterized by containing the SARS-CoV-2 spike glycoprotein-binding nucleic acid molecule of the present invention. The detection reagent of the present invention is not limited in any other configuration as long as it contains the nucleic acid molecule of the present invention. By using the detection reagent of the present invention, for example, the detection of the SARS-CoV-2 can be performed as described above.

[0080] The detection reagent of the present invention may include the sensor of the present invention as the nucleic acid molecule of the present invention, for example. The detection reagent of the present invention may further include a labeling substance, and the labeling substance may be bound to the nucleic acid molecule. For the labeling substance, for example, the explanation for the nucleic acid molecule of the present invention can be used. Furthermore, the detection reagent of the present invention may include a carrier, and the nucleic acid molecule may be immobilized on the carrier. For the carrier, for example, the explanation for the nucleic acid molecule of the present invention can be used.

[0081] The detection reagent of the present invention may contain, for example, other components in addition to the nucleic acid molecule of the present invention. The components include, for example, a non-specific adsorbent, the carrier, a buffer solution, and an instruction manual. The non-specific adsorbent includes, for example, dextran, tRNA (transfer RNA), salmon sperm DNA, and the like. The tRNA can be, for example, tRNA from E. coli MRE 600 (Merck Life Sciences, Cat No.: 10109541001), etc. The salmon sperm DNA can be, for example, deoxyribonucleic acid, low molecular weight from salmon sperm (Sigma-Aldrich, Cat No.: 31149-10G-F), etc.

[0082] In the detection reagent of the present invention, for example, the nucleic acid molecule and other components such as the buffer solution may be contained in separate containers, or may be contained in the same container, either mixed or unmixed. When the nucleic acid molecule and the other components are contained in separate containers, the detection reagent of the present invention can also be called a detection kit.

[0083] (4) Detection method The detection method of the present invention, as described above, includes a step of contacting a sample with a nucleic acid molecule and detecting the SARS-CoV-2 spike glycoprotein in the sample, the nucleic acid molecule being the SARS-CoV-2 spike glycoprotein-binding nucleic acid molecule of the present invention, and the detection step is characterized in that the SARS-CoV-2 spike glycoprotein in the sample is bound to the nucleic acid molecule, and SARS-CoV-2 in the sample is detected by the binding. The detection method of the present invention is characterized in that the nucleic acid molecule of the present invention is used, and other steps and conditions, etc. are not particularly limited. In addition, the detection method of the present invention may use the SARS-CoV-2 detection sensor of the present invention or the detection reagent or detection kit of the present invention as the nucleic acid molecule of the present invention.

[0084] According to the present invention, since the nucleic acid molecule of the present invention specifically binds to the SARS-CoV-2 spike glycoprotein, for example, by detecting the binding between the SARS-CoV-2 spike glycoprotein and the nucleic acid molecule, it is possible to specifically detect the SARS-CoV-2 spike glycoprotein and SARS-CoV-2 in a sample. Specifically, since the detection method of the present invention can detect, for example, the presence or absence of the SARS-CoV-2 spike glycoprotein or the amount of the SARS-CoV-2 spike glycoprotein in a sample, it can be said that it is also possible to perform qualitative or quantitative analysis. In addition, since the detection method of the present invention can perform, for example, qualitative or quantitative analysis, it can also be called an analytical method.

[0085] In the present invention, the sample is not particularly limited. Examples of the sample include aerosol, saliva, urine, plasma, and serum. The aerosol refers to, for example, a mixture of minute liquid or solid particles suspended in gas and the surrounding gas. When the sample is an aerosol, for example, gas that may contain the minute liquid or solid particles can be used as the sample.

[0086] The sample may be, for example, a gas sample (including the aerosol), a liquid sample, or a solid sample. The sample is preferably a liquid sample, for example, because it is easy to contact with the nucleic acid molecule and easy to handle. In the case of the gas sample and solid sample, for example, a mixed liquid, an extract, a dissolution solution, etc. may be prepared using a solvent, and this may be used. The solvent is not particularly limited, and examples thereof include water, physiological saline, a buffer solution, etc.

[0087] The detection step includes, for example, a contact step of contacting the sample with the nucleic acid molecule to bind the SARS-CoV-2 spike glycoprotein in the sample to the nucleic acid molecule, and a binding detection step of detecting the binding between the SARS-CoV-2 spike glycoprotein and the nucleic acid molecule. The detection step further includes, for example, a step of detecting the presence or absence or amount of the SARS-CoV-2 spike glycoprotein in the sample based on the result of the binding detection step.

[0088] In the contact step, the method of contacting the sample with the nucleic acid molecule is not particularly limited. The contact between the sample and the nucleic acid molecule is preferably performed in a liquid, for example. The liquid is not particularly limited, and examples thereof include water, physiological saline, and buffer solutions.

[0089] In the contact step, the contact conditions between the sample and the nucleic acid molecule are not particularly limited. The contact temperature is, for example, 4 to 37° C. or 18 to 25° C., and the contact time is, for example, 10 to 120 minutes or 30 to 60 minutes.

[0090] In the contact step, the nucleic acid molecule may be, for example, an immobilized nucleic acid molecule immobilized on a carrier, or an unimmobilized free nucleic acid molecule. In the latter case, the nucleic acid molecule is contacted with the sample, for example, in a container. The nucleic acid molecule is preferably an immobilized nucleic acid molecule, for example, because it is easy to handle. The carrier is not particularly limited, and examples thereof include a substrate, beads, a container, etc., and the container includes, for example, a microplate, a tube, etc. The immobilization of the nucleic acid molecule is, for example, as described above.

[0091] As described above, the binding detection step is a step of detecting the binding between the SARS-CoV-2 spike glycoprotein and the nucleic acid molecule in the sample. By detecting the presence or absence of the binding between the two, for example, the presence or absence of the SARS-CoV-2 spike glycoprotein in the sample can be detected (qualitative), and by detecting the degree of binding between the two (amount of binding), for example, the amount of the SARS-CoV-2 spike glycoprotein in the sample can be detected (quantitative).

[0092] If no binding between the SARS-CoV-2 spike glycoprotein and the nucleic acid molecule is detected, it can be determined that SARS-CoV-2 spike glycoprotein and SARS-CoV-2 are not present in the sample, and if the binding is detected, it can be determined that SARS-CoV-2 spike glycoprotein and SARS-CoV-2 are present in the sample.

[0093] The method for detecting the binding between the SARS-CoV-2 spike glycoprotein and the nucleic acid molecule is not particularly limited. For example, a conventionally known method for detecting binding between substances can be used, and a specific example is the above-mentioned SPR. In addition, the binding may be, for example, detection of a complex between the SARS-CoV-2 spike glycoprotein and the nucleic acid molecule.

[0094] (5) SARS-CoV-2 virus inactivation agent As described above, the inactivating agent for SARS-CoV-2 virus of the present invention is characterized by containing the SARS-CoV-2 spike glycoprotein-binding nucleic acid molecule of the present invention. The inactivating agent of the present invention is characterized by using the nucleic acid molecule of the present invention, and other configurations are not particularly limited.

[0095] "Activity" refers to, for example, the function of SARS-CoV-2 spike glycoprotein and SARS-CoV-2, specifically, for example, the function of binding to a target (e.g., angiotensin-converting enzyme 2 (ACE2)). It may also refer to, for example, the infectivity and toxicity of SARS-CoV-2. "Inactivation" refers to, for example, inhibiting the function of SARS-CoV-2 spike glycoprotein and SARS-CoV-2. "Inactivation" may completely or partially suppress the activity.

[0096] According to the present invention, since the nucleic acid molecule of the present invention specifically binds to the SARS-CoV-2 spike glycoprotein, for example, the SARS-CoV-2 spike glycoprotein and SARS-CoV-2 can be inactivated by the binding of the nucleic acid molecule to the SARS-CoV-2 spike glycoprotein. The inactivation or inactivation can also be referred to as, for example, "neutralization." Therefore, the inactivating agent of the present invention can also be referred to as a neutralizing agent. EXAMPLES

[0097] Next, examples of the present invention will be described. However, the present invention is not limited to the following examples. Commercially available reagents were used according to their protocols unless otherwise indicated.

[0098] [Example 1] The binding ability of the SARS-CoV-2 spike glycoprotein-binding nucleic acid molecule of the present invention to the SARS-CoV-2 spike glycoprotein was confirmed by SPR.

[0099] (1) Aptamers Polynucleotides consisting of the base sequences of SEQ ID NOs: 1 to 7 (SARS-CoV-2 spike glycoprotein-binding nucleic acid molecules 1 to 7 (hereinafter also referred to as "binding nucleic acid molecules 1 to 7", respectively)) were synthesized and used as DNA aptamers of the present embodiment. In the SARS-CoV-2 spike glycoprotein-binding nucleic acid molecules 1 to 7, the underlined thymine nucleotide residue in the base sequences of SEQ ID NOs: 1 to 7 was replaced with NG7 represented by formula (1).

[0100] The SARS-CoV-2 spike glycoprotein-binding nucleic acid molecules 1 to 7 were added with 20-base-long polydeoxyadenine (poly dA) at their 3' ends, and used as poly dA-added aptamers for SPR, which will be described later.

[0101] (2) Sample Four types of commercially available reagents shown in Table 1 below were used in the following tests. In the table, the targets are as follows: "RBD domain" refers to the RBD domain (hereinafter also referred to as RBD) constituting the SARS-CoV-2 spike glycoprotein; "S1+S2 domain" refers to the S1+S2 domain (hereinafter also referred to as S1S2) constituting the SARS-CoV-2 spike glycoprotein; "S1 domain" refers to the S1 domain (hereinafter also referred to as S1) constituting the SARS-CoV-2 spike glycoprotein; and "trimer" refers to the trimer of the S1+S2 domain (hereinafter also referred to as trimer). The amino acid sequence of each target was prepared based on the amino acid sequence registered under the following NCBI accession numbers, and has the amino acid sequence of SEQ ID NOs: 15 to 18 described above. RBD domain: YP_009724390.1 S1+S2 domain: YP_009724390.1 S1 domain: QHD43416.1 Trimer: QHD43416.1

[0102] [Table 1]

[0103] Each of the reagents was dissolved in sterilized distilled water to a concentration of 1 mg / mL, and the solution was used as a sample. In the binding analysis described below, SB1T buffer was used to dilute each of the samples. The composition of the SB1T buffer was 40 mmol / L HEPES (pH 7.4), 125 mmol / L NaCl, 1 mmol / L MgCl 2、 The solution contained 5 mmol / L KCl and 0.01% Tween® 20.

[0104] (3) Analysis of binding ability by SPR For the analysis of binding ability, ProteOn XPR36 (BioRad) was used according to the manufacturer's instructions.

[0105] First, a streptavidin-immobilized chip (product name: ProteOn NLC Sensor Chip, BioRad) was set in ProteON XPR36 as a sensor chip dedicated to ProteON. 2.5 μmol / L of biotinylated polydT was injected into the flow cell of the sensor chip using ultrapure water (DDW) and allowed to bind until the signal intensity (RU: Resonance Unit) was saturated. The biotinylated polydT was prepared by biotinylating the 5' end of a 20-base-long deoxythymidine. Then, 200 nmol / L of the binding nucleic acid molecules 1 to 7 with the polydA added thereto were injected into the flow cell of the chip using SB1T buffer at a flow rate of 25 μL / min for 80 seconds, respectively, and allowed to bind until the signal intensity was saturated. Next, 400 nmol / L of each sample was injected using SB1T buffer at a flow rate of 50 μL / min for 120 seconds, followed by washing for 300 seconds with SB1T buffer under the same conditions. The start of sample injection was set as 0 seconds, and the signal intensity after sample injection was measured. The SPR was performed under conditions of 25°C.

[0106] Furthermore, as a control, the signal intensity was measured in the same manner, except that a sample containing BSA (Sigma, catalog number: #A7906) was used instead of each sample shown in Table 1 above.

[0107] The results are shown in FIG. 1. FIG. 1(A)-(G) are graphs showing the binding of the binding nucleic acid molecules 1-7 to each of the samples at 400 nmol / L. In each graph in FIG. 1, the horizontal axis indicates the time (seconds) elapsed after the start of injection of the sample, and the vertical axis indicates the signal intensity (RU). As shown in FIG. 1(A), (B), and (E)-(G), the binding nucleic acid molecules 1, 2, 5-7 showed binding to RBD, S1S2, S1, and trimer in particular in each of the samples. Also, as shown in FIG. 1(C) and (D), the binding nucleic acid molecules 3 and 4 showed binding to S1S2, S1, and trimer in particular in each of the samples. On the other hand, the signal intensity of the binding nucleic acid molecules 1-7 was almost 0 for the control (BSA), and they did not show binding. From these results, it was found that the binding nucleic acid molecules 1 to 7 bind to each of the samples with excellent specificity, and that the binding can be detected by measuring the signal intensity.

[0108] As described above, the binding ability of the binding nucleic acid molecule of the present invention to the SARS-CoV-2 spike glycoprotein was confirmed by SPR.

[0109] [Example 2] The binding ability of the SARS-CoV-2 spike glycoprotein-binding nucleic acid molecule of the present invention to the SARS-CoV-2 spike glycoprotein was confirmed by SPR in a solution containing dextran, which has the effect of preventing non-specific binding of nucleic acids. Kinetic parameters were also determined.

[0110] (1) Aptamers In addition to the polynucleotides consisting of the base sequences of SEQ ID NOs: 1 to 7, polynucleotides consisting of the base sequences of SEQ ID NOs: 8 to 13 (SARS-CoV-2 spike glycoprotein-binding nucleic acid molecules 8 to 13 (hereinafter also referred to as binding nucleic acid molecules 8 to 13, respectively)) were synthesized in the same manner as in Example 1.

[0111] In addition, a similar experiment was performed using a control nucleic acid molecule (SEQ ID NO: 14) consisting of the base sequence of SEQ ID NO: 14 below instead of the binding nucleic acid molecules 1 to 13. The base sequence of SEQ ID NO: 14 below is the base sequence described in Non-Patent Document 1 (Song Y et al, Anal. Chem. (2020), 92, 9895-9900.).

[0112] Control nucleic acid molecule (SEQ ID NO: 14) CAGCACCGACCTTGTGCTTTGGGAGTGCTGGTCCAAGGGCGTTAATGGACA

[0113] (2) Sample As samples, the RBD and the trimer shown in Table 1 of Example 1 were used.

[0114] (3) Analysis of binding ability by SPR The analysis of binding ability by SPR was carried out in the same manner as in Example 1. The concentrations of the samples were 400, 200, 100, 50, and 25 nmol / L for the RBD, and 200, 100, 50, 25, and 12.5 nmol / L for the trimer, respectively.

[0115] Further, an experiment was carried out in the same manner as in Example 1, except that a buffer containing dextran (SB1T buffer containing dextran) was used instead of the SB1T buffer. The composition of the SB1T buffer containing dextran was 40 mmol / L HEPES (pH 7.4), 125 mmol / L NaCl, 1 mmol / L MgCl 2、The solution consisted of 5 mmol / L KCl, 0.01% Tween® 20, and 0.1 mmol / L dextran.

[0116] The results are shown in Figures 2 to 6. Figures 2 to 5 are graphs showing the binding affinity of the binding nucleic acid molecules 1, 2, 5 to 9, and 11 to 13 to the RBD, where (A) to (K) show the results without dextran, and (L) to (V) show the results with dextran. In each graph of Figures 2 to 5, the horizontal axis shows the elapsed time (seconds) after the start of injection of the sample, and the vertical axis shows the signal intensity (RU). In each graph of Figures 2 to 6, the black dotted line shows the actual measured value, and the gray solid line shows the fitting curve calculated based on the actual measured value. As shown in (A) to (J) and (L) to (U) of Figures 2 to 5, the binding nucleic acid molecules 1, 2, 5 to 9, and 11 to 13 showed high binding affinity to the RBD in the absence and presence of dextran. As shown in Figure 3(K) and Figure 5(V), the control nucleic acid molecule (SEQ ID NO: 14) showed binding to the RBD in a buffer without dextran, but lost its binding to the RBD in a buffer containing dextran. These results indicate that the binding nucleic acid molecules 1, 2, 5-9, and 11-13 bind to the RBD with greater specificity than the control nucleic acid molecule (SEQ ID NO: 14).

[0117] FIG. 6 is a graph showing the binding of the binding nucleic acid molecules 3, 4, and 10 to the trimer, where (A) to (D) show the results without dextran, and (E) and (F) show the results with dextran. In each graph in FIG. 6, the horizontal axis shows the time (seconds) elapsed after the start of injection of the sample, and the vertical axis shows the signal intensity (RU). As shown in FIG. 6(A) to (C), (E), and (F), the binding nucleic acid molecules 3, 4, and 10 had a constant signal intensity (RU) after the start of injection under conditions without dextran and with dextran. In contrast, as shown in FIG. 6(D), the signal intensity (RU) of the control nucleic acid molecule (SEQ ID NO: 14) decreased with the passage of time after the start of injection. These results demonstrated that the binding nucleic acid molecules 3, 4, and 10 had a slower off rate and higher binding affinity to the trimer than the control nucleic acid molecule (SEQ ID NO: 14).

[0118] Furthermore, kinetic parameters were calculated from the results of the SPR analysis in Figures 2 to 6. The results are shown in Tables 2 and 3. Tables 2 and 3 show the dissociation constants (KD) of the binding nucleic acid molecules 1 to 13 for the RBD and the trimer, respectively. In Table 2, "*" indicates that the dissociation constant (KD) was below the detection limit that can be measured by SPR. This is considered to be because the dissociation of the binding nucleic acid molecule was slow and the off rate was a very small value. As shown in Tables 2 and 3, the binding nucleic acid molecules 1 to 13 had very low dissociation constants (KD) for the RBD and the trimer compared to the control nucleic acid molecule (SEQ ID NO: 14), indicating excellent binding properties. [Table 2] [Table 3]

[0119] As described above, the binding ability of the nucleic acid molecule of the present invention to the SARS-CoV-2 spike glycoprotein was confirmed by SPR under solution conditions in which dextran, which has the effect of preventing non-specific binding of nucleic acids, was added. In addition, the kinetic parameters were able to be obtained.

[0120] [Example 3] The binding ability of the SARS-CoV-2 spike glycoprotein-binding nucleic acid molecule of the present invention to the SARS-CoV-2 spike glycoprotein was evaluated by ELAA (Enzyme Linked Aptamer Assay).

[0121] (1) Aptamers Polynucleotides consisting of the base sequences of SEQ ID NOs: 1 and 8 (binding nucleic acid molecule 1 and binding nucleic acid molecule 8) were synthesized in the same manner as in Example 1.

[0122] (2) Sample As a sample, the RBD shown in Table 1 of Example 1 was used.

[0123] (3) Analysis of binding ability by ELAA Analysis of binding ability by ELAA was performed in the same manner as described in Non-Patent Document 2 (I. Shiratori et al., Biochem Biophys Res Commun 443 (2014) 37-41.). Specifically, the RBD was diluted with 50 mmol / l carbonate buffer (pH 9.6) to 1 μg / 100 μl. After the dilution, the RBD was added to wells of a Maxisorp plate (Thermo Scientific / Nunc, Waltham, MA) to 1 μg / well, and immobilized at 4° C. overnight. After the solid phase, the wells were washed once with 200 μl of SB1T buffer (40 mmol / l HEPES (pH 7.4), 125 mmol / l NaCl, 1 mmol / l MgCl2, 5 mmol / l KCl, 0.01% Tween20), and after the washing, blocking was performed with 200 μl of TBS blocking buffer (Cat No.: 37570, Pierce Biotechnology) at room temperature (about 25° C., the same applies below) for 1 hour. Next, the 5′ end of the aptamer was biotinylated in the same manner as in Example 1 (3). After the biotinylation, the aptamer was adjusted to 1 μmol / l with SB1T buffer and thermally denatured at 95° C. for 5 minutes. After the denaturation, the wells were rapidly cooled to 4° C. and folded. The aptamer was added to the wells with RBD immobilized at 50 μl / well, and incubated at 25° C. for 1 hour. After the incubation, the wells were washed three times with 200 μl of SB1T buffer. After the washing, 100 μl of streptavidin-horseradish peroxidase (SA-HRP) (manufactured by Citiva) diluted 1000-fold with SB1T buffer was added, and incubated at 25° C. for 30 minutes. After the incubation, the wells were washed three times with SB1T buffer. After the washing, 100 μl of TMB solution (manufactured by MOSS) was added, and incubated at room temperature for 10 minutes. After the incubation, 0.5 N sulfuric acid was added to stop the reaction.Thereafter, the absorbance of the wells was measured at 490 nm and 620 nm (background) using infinite M1000Pro (manufactured by Tecan). Then, the difference in absorbance between 490 nm and 620 nm (ABS (490 nm-620 nm)) was calculated for each bound nucleic acid molecule. As a control, the difference in absorbance was calculated in the same manner except that the bound nucleic acid molecules 1 and 8 were not added.

[0124] The results are shown in Figure 8. Figure 8 is a graph showing the binding ability of the binding nucleic acid molecules 1 and 8 to the RBD. In Figure 8, the vertical axis shows the average of the difference in absorbance for three wells of the same sample, the error bars show the standard deviation, and the horizontal axis shows the type of binding nucleic acid molecule and the presence or absence of addition of RBD. In the absence of an aptamer, the difference in absorbance did not change regardless of the presence or absence of addition of RBD. On the other hand, in the binding nucleic acid molecules 1 and 8, the difference in absorbance increased in the presence of RBD.

[0125] Based on the above, the binding ability of the binding nucleic acid molecule of the present invention to the SARS-CoV-2 spike glycoprotein was confirmed by ELAA.

[0126] Although the present invention has been described above with reference to the embodiments and examples, the present invention is not limited to the above-mentioned embodiments and examples. Various modifications that can be understood by those skilled in the art can be made to the configuration and details of the present invention within the scope of the present invention.

[0127] This application claims priority based on Japanese Patent Application No. 2020-198556, filed on November 30, 2020, the disclosure of which is incorporated herein in its entirety.

[0128] <Additional Notes> Some or all of the above-described embodiments and examples may be described as follows, but are not limited to the following: (Appendix 1) A SARS-CoV-2 spike glycoprotein-binding nucleic acid molecule comprising any one of the following polynucleotides (a), (b), (c), and (d): (a) a polynucleotide consisting of the nucleotide sequence of any one of SEQ ID NOs: 1 to 7 or a partial sequence of the nucleotide sequence of any one of SEQ ID NOs: 1 to 7; (b) a polynucleotide having a nucleotide sequence having 80% or more identity to the nucleotide sequence of (a), and binding to the SARS-CoV-2 spike glycoprotein; (c) a polynucleotide that has a nucleotide sequence complementary to a polynucleotide that hybridizes under stringent conditions to the polynucleotide having the nucleotide sequence of (a), and that binds to the SARS-CoV-2 spike glycoprotein; (d) a polynucleotide that is composed of the base sequence of (a) in which one or several bases are deleted, substituted, inserted and / or added, and that binds to the SARS-CoV-2 spike glycoprotein; SEQ ID NO: 1: GGTATGTCTCCGCCACTGAAATCCG T GCC T AA T C T CACCCCACGGAA TT CA T GGCAAAGCCGAGG T G T C TT G T A TT C; SEQ ID NO: 2: GGTATGTCTCCGCCACTGAAATC T AA T C T CACA TT G T AAGCAAAGGAGAA T AAGCAAGCCGAGG T G T C TT G T A TT C; SEQ ID NO: 3: GGTATGTCTCCGCCACTGAAATCCC T GACCGC T GACCAAA T CT CAG T GCAGA T GCAAAGCCGAGG T G T C TT G T A TT C; Phone 4:GGTATGTCTCCGCCACTGAAATC TT G T CCCC T SECOND T AGG TT CCGAC T GACAAG T GCAAAGCCGAGG T G T C TT G T A TT C; Photo 5:GGTTTAGCCCTCGACTCCAAATG T GCAC T C T CCCGG T A T CCC T SECOND T C T CACCCGA T ACCG TT THOUGH T G T GC T GCCAA T AC; Page 6:GGTTTAGCCCTCGACTCCAAATGACA T GAGCCAGG T GCA T C TT GAACG T CA T AGA T ACCG TT THOUGH T G T GC T GCCAA T AC; Photo 7:GGTTTAGCCCTCGACTCCAAATG T GC T THOUGH T AC T CGG T A TCCC T AA T C T CACCCGA T ACCG TT GA T G T GC T GCCAA T AC. (Appendix 2) The partial sequence of the base sequence of SEQ ID NO: 1 is the base sequence of SEQ ID NO: 8. The partial sequence of the base sequence of SEQ ID NO: 2 is the base sequence of SEQ ID NO: 9. The partial sequence of the base sequence of SEQ ID NO: 3 is the base sequence of SEQ ID NO: 10. The partial sequence of the base sequence of SEQ ID NO: 5 is the base sequence of SEQ ID NO: 11. A partial sequence of the base sequence of SEQ ID NO: 6 is the base sequence of SEQ ID NO: 12, or The nucleic acid molecule according to Appendix 1, wherein the partial sequence of the nucleotide sequence of SEQ ID NO: 7 is the nucleotide sequence of SEQ ID NO: 13: SEQ ID NO: 8: CCACTGAAATCCG T GCC T AA T C T CACCCCACGGAA TT CA T GG; SEQ ID NO: 9: TCCGCCACTGAAATC T AA T C T CACA TT G T AAGCAAAGGAGAA T A.A.; SEQ ID NO: 10: CCACTGAAATCCC T GACCGC T GACCAAA T C T CAG T GCAGA T; SEQ ID NO: 11: TGTGCACTCTCCCGG T A T CCC T AA T C T CACCCGA T ACC; SEQ ID NO: 12: TGACATGAGCCAGG T GCA T C TT GAACG T CA T AGA T ACCG TT GA T G T GC T G; SEQ ID NO: 13: GCTGATACTCGG T A T CCC T AA T C T CACCCGA T A.C.C.G. (Appendix 3) The nucleic acid molecule according to Appendix 1 or 2, wherein the polynucleotide (b) is a polynucleotide (b1) below: (b1) A polynucleotide consisting of a base sequence having 80% or more identity to the base sequence of (a), comprising any of the base sequences of SEQ ID NOs: 8 to 13, and binding to the SARS-CoV-2 spike glycoprotein. (Appendix 4) The nucleic acid molecule according to Appendix 1 or 2, wherein the polynucleotide (d) is a polynucleotide (d1) below: (d1) A polynucleotide consisting of a base sequence in which one or several bases are deleted, substituted, inserted and / or added in the base sequence of (a), which contains any of the base sequences of SEQ ID NOs: 8 to 13, and which binds to the SARS-CoV-2 spike glycoprotein. (Appendix 5) 5. The SARS-CoV-2 spike glycoprotein-binding nucleic acid molecule of any one of claims 1 to 4, wherein the binding nucleic acid molecule comprises a modified base, the base being modified with a modifying group. (Appendix 6) 6. The SARS-CoV-2 spike glycoprotein-binding nucleic acid molecule of claim 5, wherein the modified base is a modified thymine base, wherein the thymine base is modified with a modifying group. (Appendix 7) The SARS-CoV-2 spike glycoprotein-binding nucleic acid molecule according to appendix 6, wherein the modified thymine base is a modified thymine base represented by the following formula (1): [ka] (Appendix 8) The SARS-CoV-2 spike glycoprotein-binding nucleic acid molecule according to any one of Appendices 5 to 7, wherein in the polynucleotide, the underlined thymine base in the base sequence of SEQ ID NOs: 1 to 13 is a modified base. (Appendix 9) 9. The SARS-CoV-2 spike glycoprotein-binding nucleic acid molecule of any one of claims 1 to 8, wherein the polynucleotide is DNA. (Appendix 10) A sensor for detecting SARS-CoV-2, comprising a SARS-CoV-2 spike glycoprotein-binding nucleic acid molecule described in any one of appendix 1 to 9. (Appendix 11) A reagent for detecting SARS-CoV-2, comprising a SARS-CoV-2 spike glycoprotein-binding nucleic acid molecule according to any one of claims 1 to 9. (Appendix 12) contacting a sample with a nucleic acid molecule and detecting SARS-CoV-2 spike glycoprotein in the sample; the nucleic acid molecule is a SARS-CoV-2 spike glycoprotein-binding nucleic acid molecule according to any one of claims 1 to 9, A method for detecting SARS-CoV-2, characterized in that in the detection step, the SARS-CoV-2 spike glycoprotein in the sample is bound to the nucleic acid molecule, and the SARS-CoV-2 spike glycoprotein in the sample is detected through the binding. (Appendix 13) The method for detecting SARS-CoV-2 described in Appendix 12, wherein the sample is at least one selected from the group consisting of aerosol, saliva, urine, plasma, and serum. (Appendix 14) An agent for inactivating or neutralizing the SARS-CoV-2 virus, characterized in that it comprises a SARS-CoV-2 spike glycoprotein-binding nucleic acid molecule described in any one of appendices 1 to 9. [Industrial Applicability]

[0129] As described above, the SARS-CoV-2 spike glycoprotein-binding nucleic acid molecule of the present invention can specifically bind to the SARS-CoV-2 spike glycoprotein, has high binding strength, and is slow to dissociate. Therefore, the SARS-CoV-2 spike glycoprotein-binding nucleic acid molecule of the present invention can detect the SARS-CoV-2 spike glycoprotein with high accuracy, for example, based on the presence or absence of binding to the SARS-CoV-2 spike glycoprotein in a sample. Therefore, the SARS-CoV-2 spike glycoprotein-binding nucleic acid molecule of the present invention can be said to be an extremely useful tool for detecting SARS-CoV-2 in the fields of preventive medicine, health management, and diagnosis of infectious diseases, for example. The SARS-CoV-2 spike glycoprotein-binding nucleic acid molecule of the present invention can be used, for example, for detecting viruses in biological samples and aerosols, and for inactivating viruses.

Claims

1. The present invention comprises any one of the following polynucleotides (a), (b1), (c), and (d): A partial sequence of the base sequence of SEQ ID NO: 1 is the base sequence of SEQ ID NO:

8. A partial sequence of the base sequence of SEQ ID NO: 2 is the base sequence of SEQ ID NO:

9. A partial sequence of the base sequence of SEQ ID NO: 3 is the base sequence of SEQ ID NO:

10. A partial sequence of the base sequence of SEQ ID NO:5 is the base sequence of SEQ ID NO:

11. A partial sequence of the base sequence of SEQ ID NO:6 is the base sequence of SEQ ID NO:12, or A partial sequence of the base sequence of SEQ ID NO: 7 is the base sequence of SEQ ID NO: 13, A SARS-CoV-2 spike glycoprotein-binding nucleic acid molecule, characterized in that the underlined T in the following SEQ ID NOs: 1 to 13 is a thymine base or a modified thymine base partially or entirely modified with a modifying group: (a) a polynucleotide consisting of the nucleotide sequence of any one of SEQ ID NOs: 1 to 7 or a partial sequence of any one of SEQ ID NOs: 1 to 3 and 5 to 7; (b1) a polynucleotide consisting of a base sequence having 80% or more identity to the base sequence of (a), comprising any of the base sequences of SEQ ID NOs: 8 to 13, and binding to the SARS-CoV-2 spike glycoprotein; (c) a polynucleotide that has a base sequence complementary to a polynucleotide that hybridizes under stringent conditions to the polynucleotide having the base sequence of (a) and binds to the SARS-CoV-2 spike glycoprotein; (d) A polynucleotide comprising the base sequence of (a) in which one or several bases have been deleted, substituted, inserted and / or added, and which binds to the SARS-CoV-2 spike glycoprotein.

2. The SARS-CoV-2 spike glycoprotein-binding nucleic acid molecule of claim 1, wherein the binding nucleic acid molecule comprises a modified base, the base being modified with a modifying group.

3. The SARS-CoV-2 spike glycoprotein-binding nucleic acid molecule according to claim 2, wherein the modified base is a modified thymine base represented by the following formula (1): 【Chemistry 1】

4. The SARS-CoV-2 spike glycoprotein-binding nucleic acid molecule according to claim 2 or 3, wherein in the polynucleotide, the underlined thymine bases in the base sequences of SEQ ID NOs: 1 to 13 are modified bases.

5. A sensor for detecting SARS-CoV-2, comprising the SARS-CoV-2 spike glycoprotein-binding nucleic acid molecule according to any one of claims 1 to 4.

6. A reagent for detecting SARS-CoV-2, comprising the SARS-CoV-2 spike glycoprotein-binding nucleic acid molecule according to any one of claims 1 to 4.

7. contacting a sample with a nucleic acid molecule and detecting SARS-CoV-2 spike glycoprotein in said sample; The nucleic acid molecule is a SARS-CoV-2 spike glycoprotein-binding nucleic acid molecule according to any one of claims 1 to 4, A method for detecting SARS-CoV-2, characterized in that in the detection step, the SARS-CoV-2 spike glycoprotein in the sample is bound to the nucleic acid molecule, and the SARS-CoV-2 spike glycoprotein in the sample is detected through the binding.

8. An agent for inactivating SARS-CoV-2 virus, comprising the SARS-CoV-2 spike glycoprotein-binding nucleic acid molecule according to any one of claims 1 to 4.

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