Dry composition for nucleic acid amplification and nucleic acid amplification method using same

Abstract

Provided is a dry composition for nucleic acid amplification, which contains a polymer that contains a structural unit derived from a monomer represented by formula (1) (wherein the symbols are as defined in the description). # imgabs0 #

Description

Technical Field

[0001] The present invention relates to a drying composition for nucleic acid amplification and a nucleic acid amplification method using the same. Background Art

[0002] As a reaction for newly synthesizing a nucleic acid complementary to a nucleic acid using the nucleic acid as a template, a nucleic acid amplification method is known. To perform a nucleic acid amplification reaction, in addition to the nucleic acid as a template, various nucleic acid amplification reagents such as oligonucleotides called primers and enzymes are required. It is known that these nucleic acid amplification reagents deteriorate significantly due to a decrease in enzyme activity at room temperature (e.g., 15°C to 25°C) or under refrigerated conditions. As a method for preventing this deterioration, drying of the nucleic acid amplification reagents is known.

[0003] As a drying method, the most commonly used is freeze-drying (e.g., Patent Documents 1 to 3). As other drying methods, for example, a method of drying at room temperature and atmospheric pressure (air drying method) and a method of drying at a temperature above room temperature and a pressure of 90% or less of atmospheric pressure (evaporative drying method) are described in Patent Document 4. Prior Art Documents Patent Documents

[0004] Patent Document 1: Japanese Patent Publication No. 2000-513940 Patent Document 2: Japanese Patent Publication No. 10-503383 Patent Document 3: Japanese Patent Application Laid-Open No. 02-265984 Patent Document 4: Japanese Patent Application Laid-Open No. 2015-092870 Summary of the Invention Problems to be Solved by the Invention

[0005] When a composition for nucleic acid amplification containing nucleic acid amplification reagents is dried using the drying methods described in Patent Documents 1 to 4, the stability of the resulting dried composition for nucleic acid amplification is improved compared to the solution-like composition before drying. However, when the dried composition for nucleic acid amplification is stored for a long time, problems such as aggregation of nucleic acid amplification reagents, structural changes, and adsorption on the container surface occur, resulting in a decrease in their activity. That is, even when using the drying methods described in Patent Documents 1 to 4, the storage stability of the resulting dried composition for nucleic acid amplification is not sufficient.

[0006] The present invention has been made in view of the above problems, and an object thereof is to provide a dried composition for nucleic acid amplification having excellent storage stability. Means for Solving the Problems

[0007] The present inventors have conducted in-depth research and found that if a polymer containing a structural unit derived from the monomer represented by the following formula (1) is used, the storage stability of the dry composition for nucleic acid amplification can be improved. The present invention based on this finding is as follows.

[0008] [1], A dry composition for nucleic acid amplification, comprising a polymer containing a structural unit derived from the monomer represented by formula (1):

[0009] [Chemical Formula 1]

[0010] (In the formula, X 1 represents (meth)acryloyloxy or (meth)acrylamino, L 1 represents an alkylene group having 2 to 4 carbon atoms which may have 1 hydroxyl group, or an alkyleneoxyalkylene group having 2 to 4 carbon atoms, and R 1 ~R 3 each independently represents an alkyl group having 1 to 3 carbon atoms.) [2] The dry composition for nucleic acid amplification according to the above [1], wherein the polymer is a homopolymer composed of one structural unit derived from the monomer represented by formula (1). [3] The dry composition for nucleic acid amplification according to the above [1], wherein the polymer is a copolymer further containing a structural unit derived from the monomer represented by formula (2),

[0011] [Chemical Formula 2]

[0012] (In the formula, R 4 represents a hydrogen atom or a methyl group, and R 5 represents a hydrogen atom or an alkyl group having 1 to 20 carbon atoms.) [4] The dry composition for nucleic acid amplification according to the above [1] or [3], wherein the polymer is a copolymer further containing a structural unit derived from the monomer represented by formula (3),

[0013] [Chemical Formula 3]

[0014] (In the formula, R 6 represents a hydrogen atom or a methyl group, and R 7 represents an alkyl group having 3 to 6 carbon atoms having 2 or more hydroxyl groups.) [5] The dry composition for nucleic acid amplification according to [1], [3], or [4] above, wherein the polymer is a copolymer further containing a structural unit derived from the monomer represented by formula (4),

[0015] [Chemical formula 4]

[0016] (In the formula, R 8 represents a hydrogen atom or a methyl group, n represents a number from 1 to 10, and R 9 represents a hydrogen atom, a methyl group, or an ethyl group.) [6] The dry composition for nucleic acid amplification according to any one of [1] to [5] above, which further contains a polymerase. [7] The dry composition for nucleic acid amplification according to [6] above, which is used for the quantitative polymerase chain reaction method. [8] The dry composition for nucleic acid amplification according to any one of [1] to [7] above, which further contains primers. [9] A nucleic acid amplification method, which includes the steps of preparing a reaction solution by mixing the dry composition for nucleic acid amplification according to any one of [1] to [8] above, the nucleic acid to be amplified, and water, and performing a nucleic acid amplification reaction using the obtained reaction solution. Effects of the Invention

[0017] According to the present invention, a dry composition for nucleic acid amplification with excellent storage stability can be obtained. Brief Description of the Drawings

[0018] Figure 1 is a photograph showing the results of electrophoresis performed in Examples 1 to 4 and Comparative Example 1. Detailed Description of the Invention

[0019] The present invention will be described in detail below. In addition, in this specification, when a stepwise numerical range is described, the lower limit value and the upper limit value of each numerical range can be combined. For example, when it is described that "preferably 10 to 100, more preferably 20 to 90", the "preferred lower limit value: 10" can be combined with the "more preferred upper limit value: 90" (that is, the numerical range of "10 to 90" is also within the scope of this specification).

[0020] [The Dry Composition of the Present Invention] ​The present invention provides a dry composition for nucleic acid amplification (which may be referred to as "the dry composition of the present invention" in this specification) containing a polymer having a structural unit derived from a monomer represented by the formula (1) (which may be referred to as "the polymer of the present invention" in this specification).

[0021] In this specification, the "dry composition" refers to a solid composition obtained by drying a solution or a dispersion liquid.

[0022] In this specification, the "dry composition for nucleic acid amplification" refers to a dry composition for a nucleic acid amplification method. Specifically, the "dry composition for nucleic acid amplification" refers to a dry composition used for preparing a reaction solution by mixing the dry composition, the nucleic acid to be amplified, and water, and performing a nucleic acid amplification reaction using the obtained reaction solution.

[0023] Examples of the nucleic acid amplification method include: Polymerase Chain Reaction (PCR) method, Loop mediated isothermal Amplification (LAMP) method, Transcription Mediated Amplification (TMA) method, Isothermal and Chimeric primer-initiated Amplification of Nucleic acids (IICAN) method, Strand Displacement Amplification (SDA) method, Ligase Chain Reaction (LCR) method, Nucleic Acid Seqence-Based Amplification (NASBA) method, and the like.

[0024] The nucleic acid amplification method is preferably the polymerase chain reaction (PCR method), more preferably the quantitative polymerase chain reaction. That is, the dry composition of the present invention is more preferably used for the quantitative polymerase chain reaction. In addition, in this specification, in the "polymerase chain reaction (PCR method)", it includes not only the typical PCR method using DNA as the target nucleic acid, but also the reverse transcription polymerase chain reaction (RT-PCR method) using RNA as the target nucleic acid. Furthermore, in this specification, in the "quantitative polymerase chain reaction", it includes not only the typical quantitative polymerase chain reaction using DNA as the target nucleic acid, but also the quantitative reverse transcription polymerase chain reaction using RNA as the target nucleic acid.

[0025] [The polymer of the present invention] The polymer of the present invention is a polymer containing a structural unit derived from a monomer represented by the formula (1) (hereinafter sometimes simply referred to as "monomer (1)").

[0026] [Chemical Formula 5]

[0027] (In the formula, X 1 represents (meth)acryloyloxy or (meth)acrylamino, L 1 represents an alkylene group having 2 to 4 carbon atoms which may have 1 hydroxyl group, or an alkyleneoxyalkylene group having 2 to 4 carbon atoms, and R 1 ~R 3 each independently represents an alkyl group having 1 to 3 carbon atoms.) The polymer of the present invention can be used alone, or two or more kinds can be used in combination.

[0028] The structural unit derived from monomer (1) (hereinafter sometimes simply referred to as "structural unit (1)") means a structural unit having a structure formed by the reaction of the carbon-carbon double bond of the (meth)acryloyl group contained in monomer (1). The structural units derived from other monomers have the same meaning as the structural units derived from monomer (1).

[0029] Monomer (1) can be used alone, or two or more kinds can be used in combination. That is, the polymer of the present invention can be a homopolymer composed of one kind of structural unit (1), or a copolymer containing two or more kinds of structural units (1). The copolymer can be a random copolymer, a block copolymer, or a copolymer containing both a random part and a block part.

[0030] Hereinafter, the groups in the formula (1) will be described in turn. X in the formula (1) 1 represents (meth)acryloyloxy (i.e., CH2=CR-CO-O-, R: hydrogen atom or methyl) or (meth)acrylamino (i.e., CH2=CR-CO-NH-, R: hydrogen atom or methyl). From the viewpoint of starting material chirality, X 1 is preferably (meth)acryloyloxy, more preferably methacryloyloxy.

[0031] L in the formula (1) 1represents an alkylene group having 2 to 4 carbon atoms and 1 hydroxyl group, or an alkyleneoxyalkylene group having 2 to 4 carbon atoms. The above alkylene group can be linear or branched. As the alkylene group having 2 to 4 carbon atoms and 1 hydroxyl group, for example, -C2H4- can be cited. As the alkyleneoxyalkylene group having 2 to 4 carbon atoms, for example, -C2H4-O-C2H4- can be cited. From the perspective of starting material chirality, L 1 is preferably -C2H4- or -C2H4-O-C2H4-, more preferably -C2H4- (i.e., ethylene).

[0032] R in formula (1) 1 ~R 3 each independently represents an alkyl group having 1 to 3 carbon atoms. The above alkyl group can be linear or branched. As the alkyl group having 1 to 3 carbon atoms, for example, methyl, ethyl, propyl, etc. can be cited. From the perspective of starting material chirality, R 1 ~R 3 are preferably all methyl groups.

[0033] The preferred monomer (1) is: X 1 is (meth)acryloyloxy, L 1 is -C2H4- or -C2H4-O-C2H4-, and R 1 ~R 3 are methyl groups. A more preferred monomer (1) is: X 1 is (meth)acryloyloxy, L 1 is ethylene, and R 1 ~R 3 are methyl groups (i.e., 2-(meth)acryloyloxyethylphosphorylcholine). A further preferred monomer (1) is: 2-methacryloyloxyethylphosphorylcholine. Monomer (1) can be a commercially available product.

[0034] In this specification, "2-(meth)acryloyloxyethylphosphorylcholine" basically refers to "2-acryloyloxyethylphosphorylcholine or 2-methacryloyloxyethylphosphorylcholine". When there are multiple 2-(meth)acryloyloxyethylphosphorylcholines, "2-(meth)acryloyloxyethylphosphorylcholine" refers to "2-acryloyloxyethylphosphorylcholine and / or 2-methacryloyloxyethylphosphorylcholine". Other terms similar to "2-(meth)acryloyloxyethylphosphorylcholine" also have the same meaning as "2-(meth)acryloyloxyethylphosphorylcholine".

[0035] As an embodiment of the polymer of the present invention, a polymer composed of structural unit (1) can be cited. In the present specification, the "polymer composed of structural unit (1)" means a polymer in which all the structural units (repeating units) in the polymer chain are composed of structural unit (1). The same expression as the "polymer composed of structural unit (1)" also has the same meaning as the "polymer composed of structural unit (1)". The polymer composed of structural unit (1) can be a homopolymer composed of one kind of structural unit (1) (hereinafter sometimes simply referred to as "homopolymer (1)"), or a copolymer composed of two or more kinds of structural units (1), but a homopolymer (1) is preferred.

[0036] The weight-average molecular weight of the homopolymer (1) is not particularly limited, and is preferably 20,000 to 2,000,000, more preferably 100,000 to 1,500,000, and still more preferably 500,000 to 1,500,000. The weight-average molecular weight can be determined, for example, by gel permeation chromatography using an EcoSEC system (manufactured by Tosoh Corporation) or the like, and converted based on polyethylene glycol.

[0037] In addition to structural unit (1), the polymer of the present invention may further contain a structural unit (hereinafter sometimes simply referred to as "structural unit (2)") derived from the monomer represented by formula (2) (hereinafter sometimes simply referred to as "monomer (2)").

[0038] [Chemical formula 6]

[0039] (In the formula, R 4 represents a hydrogen atom or a methyl group, and R 5 represents a hydrogen atom or an alkyl group having 1 to 20 carbon atoms.)

[0040] Only one kind of monomer (2) can be used, or two or more kinds can be used in combination. That is, the polymer of the present invention can also be a copolymer containing one or more structural units (1) and one or more structural units (2). The copolymer can be a random copolymer, a block copolymer, or a copolymer containing both a random part and a block part.

[0041] Hereinafter, the groups in formula (2) will be described in turn. R 4 in formula (2) represents a hydrogen atom or a methyl group. From the viewpoint of the storage stability of the polymer, R 4 is preferably a methyl group.

[0042] R 5represents a hydrogen atom or an alkyl group having 1 to 20 carbon atoms. The above alkyl group may be linear or branched, preferably a hydrogen atom or an alkyl group having 2 to 20 carbon atoms, more preferably a hydrogen atom or an alkyl group having 3 to 19 carbon atoms, and further preferably a linear alkyl group having 4 to 18 carbon atoms. In addition, when the polymer of the present invention further contains a structural unit derived from a monomer represented by the following formula (3) in addition to the structural units (1) and (2), R 5 is preferably a linear alkyl group having 3 to 6 carbon atoms.

[0043] Specific examples of the monomer (2) include: (meth)acrylic acid, methyl (meth)acrylate, ethyl (meth)acrylate, propyl (meth)acrylate, butyl (meth)acrylate, pentyl (meth)acrylate, hexyl (meth)acrylate, heptyl (meth)acrylate, octyl (meth)acrylate, nonyl (meth)acrylate, decyl (meth)acrylate, undecyl (meth)acrylate, dodecyl (meth)acrylate, tridecyl (meth)acrylate, tetradecyl (meth)acrylate, pentadecyl (meth)acrylate, hexadecyl (meth)acrylate, heptadecyl (meth)acrylate, stearyl (meth)acrylate, etc. The monomer (2) can be a commercially available product.

[0044] Among the above specific examples of the monomer (2), (i) are preferably (meth)acrylic acid, methyl (meth)acrylate, ethyl (meth)acrylate, propyl (meth)acrylate, butyl (meth)acrylate, pentyl (meth)acrylate, hexyl (meth)acrylate, heptyl (meth)acrylate, octyl (meth)acrylate, nonyl (meth)acrylate, decyl (meth)acrylate, undecyl (meth)acrylate, dodecyl (meth)acrylate, tridecyl (meth)acrylate, tetradecyl (meth)acrylate, pentadecyl (meth)acrylate, hexadecyl (meth)acrylate, heptadecyl (meth)acrylate, and stearyl (meth)acrylate; (ii) are more preferably (meth)acrylic acid, methyl (meth)acrylate, ethyl (meth)acrylate, propyl (meth)acrylate, butyl (meth)acrylate, dodecyl (meth)acrylate, tridecyl (meth)acrylate, tetradecyl (meth)acrylate, pentadecyl (meth)acrylate, hexadecyl (meth)acrylate, heptadecyl (meth)acrylate, and stearyl (meth)acrylate; (iii) are further preferably (meth)acrylic acid, butyl (meth)acrylate, and stearyl (meth)acrylate; (iv) are particularly preferably methacrylic acid, butyl methacrylate, and stearyl methacrylate.

[0045] As an embodiment of the polymer of the present invention, there may be mentioned: a copolymer composed of structural unit (1) and structural unit (2) (hereinafter, there may be a case where it is simply referred to as "copolymer (1-2)"). In copolymer (1-2), with respect to a total of 100 moles of structural unit (1) and structural unit (2) (i.e., a total of 100 moles of monomer (1) and monomer (2) used in the polymerization), the molar ratio of structural unit (1) (i.e., the molar ratio of monomer (1) used in the polymerization) is preferably 5 to 95 moles, more preferably 10 to 95 moles, further preferably 15 to 95 moles, and particularly preferably 20 to 90 moles; the molar ratio of structural unit (2) (i.e., the molar ratio of monomer (2) used in the polymerization) is preferably 5 to 95 moles, more preferably 5 to 90 moles, further preferably 5 to 85 moles, and particularly preferably 10 to 80 moles.

[0046] The weight-average molecular weight of copolymer (1-2) is not particularly limited, and is preferably 10,000 to 2,000,000, more preferably 20,000 to 1,500,000, and further preferably 30,000 to 1,000,000.

[0047] In addition to structural unit (1), the polymer of the present invention may further contain a structural unit (hereinafter, there may be a case where it is simply referred to as "structural unit (3)") derived from the monomer represented by formula (3) (hereinafter, there may be a case where it is simply referred to as "monomer (3)")

[0048] [Chemical formula 7]

[0049] (In the formula, R 6 represents a hydrogen atom or a methyl group, and R 7 represents an alkyl group having 3 to 6 carbon atoms and having two or more hydroxyl groups.)

[0050] Only one kind of monomer (3) may be used, or two or more kinds may be used in combination. That is, the polymer of the present invention may be a copolymer containing one or more structural units (1) and one or more structural units (3), and may also be a copolymer containing one or more structural units (1), one or more structural units (2) and one or more structural units (3). The copolymer may be a random copolymer, a block copolymer, or a copolymer containing both a random part and a block part.

[0051] Hereinafter, the groups in formula (3) will be described in turn. R 6 represents a hydrogen atom or a methyl group. From the viewpoint of the storage stability of the polymer, R 6 is preferably a methyl group.

[0052] R in formula (3) 7 represents an alkyl group having 3 to 6 carbon atoms and having 2 or more hydroxyl groups. The number of hydroxyl groups in R 7 is preferably 2 to 5. The above alkyl group may be linear or branched. Examples of the alkyl group having 3 to 6 carbon atoms include propyl, butyl, pentyl, hexyl, etc.

[0053] Specific examples of the monomer (3) include glycerol mono(meth)acrylate, threitol mono(meth)acrylate, erythritol mono(meth)acrylate, xylitol mono(meth)acrylate, arabitol mono(meth)acrylate, mannitol mono(meth)acrylate, galactitol mono(meth)acrylate, sorbitol mono(meth)acrylate, etc. Among these, glycerol mono(meth)acrylate and xylitol mono(meth)acrylate are preferred, glycerol mono(meth)acrylate is more preferred, and glycerol monomethacrylate is further preferred.

[0054] The monomer (3) can be a commercially available product or can be produced according to a known method. For example, the monomer (3) can be produced by an esterification reaction of (meth)acrylic acid or its derivative (e.g., acyl chloride) with a polyol having 3 or more hydroxyl groups. The esterification reaction is well-known, and those skilled in the art can appropriately set the conditions for carrying out the reaction.

[0055] As an embodiment of the polymer of the present invention, a copolymer composed of the structural unit (1) and the structural unit (3) can be cited (hereinafter, there is a case where it is simply referred to as "copolymer (1-3)"). In the copolymer (1-3), with respect to a total of 100 moles of the structural unit (1) and the structural unit (3) (i.e., a total of 100 moles of the monomer (1) and the monomer (3) used in the polymerization), the molar ratio of the structural unit (1) (i.e., the molar ratio of the monomer (1) used in the polymerization) is preferably 25 to 95 moles, more preferably 30 to 90 moles, and further preferably 35 to 85 moles; the molar ratio of the structural unit (3) (i.e., the molar ratio of the monomer (3) used in the polymerization) is preferably 5 to 75 moles, more preferably 10 to 70 moles, and further preferably 15 to 65 moles.

[0056] The weight average molecular weight of the copolymer (1-3) is not particularly limited, and is preferably 100,000 to 1,000,000, more preferably 100,000 to 800,000, and further preferably 100,000 to 500,000.

[0057] As an embodiment of the polymer of the present invention, a copolymer composed of structural unit (1), structural unit (2), and structural unit (3) (hereinafter, there may be a case where it is simply referred to as "copolymer (1-2-3)") can be cited. In copolymer (1-2-3), with respect to a total of 100 moles of structural unit (1), structural unit (2), and structural unit (3) (i.e., a total of 100 moles of monomer (1), monomer (2), and monomer (3) used in the polymerization), the molar ratio of structural unit (1) (i.e., the molar ratio of monomer (1) used in the polymerization) is preferably 30 to 80 moles, more preferably 30 to 70 moles, and still more preferably 30 to 60 moles; the molar ratio of structural unit (2) (i.e., the molar ratio of monomer (2) used in the polymerization) is preferably 10 to 60 moles, more preferably 20 to 60 moles, and still more preferably 30 to 60 moles; the molar ratio of structural unit (3) (i.e., the molar ratio of monomer (3) used in the polymerization) is preferably 10 to 60 moles, more preferably 10 to 50 moles, and still more preferably 10 to 40 moles.

[0058] The weight average molecular weight of copolymer (1-2-3) is not particularly limited, and is preferably 10,000 to 500,000, more preferably 10,000 to 100,000, and still more preferably 10,000 to 50,000.

[0059] In addition to structural unit (1), the polymer of the present invention may further contain a structural unit (hereinafter, there may be a case where it is simply referred to as "structural unit (4)") derived from the monomer represented by formula (4) (hereinafter, there may be a case where it is simply referred to as "monomer (4)")

[0060] [Chemical formula 8]

[0061] (In the formula, R 8 represents a hydrogen atom or a methyl group, n represents a number from 1 to 10, and R 9 represents a hydrogen atom, a methyl group, or an ethyl group.)

[0062] Only one kind of monomer (4) can be used, or two or more kinds can be used in combination. That is, the polymer of the present invention can be a copolymer containing one or more structural units (1) and one or more structural units (4), and can also be a copolymer containing one or more structural units (1), one or more structural units (4), and one or more other structural units (for example, one or more structural units (2) and / or one or more structural units (3)). The copolymer can be a random copolymer, a block copolymer, or a copolymer containing both a random part and a block part.

[0063] Hereinafter, the groups in formula (4) will be described. R 8 represents a hydrogen atom or a methyl group. From the viewpoint of the storage stability of the polymer, R 8 is preferably a methyl group.

[0064] In formula (4), n represents a number from 1 to 10, preferably a number from 8 to 10. In addition, n represents the number of OCH2CH2 units in the polyethylene glycol chain, and n of the actual monomer (4) is an average value. Therefore, n can also be a decimal. In formula (4), R 9 represents a hydrogen atom, a methyl group or an ethyl group, preferably a methyl group or an ethyl group, more preferably a methyl group.

[0065] As specific examples of the monomer (4), polyethylene glycol mono(meth)acrylate, methoxypolyethylene glycol (meth)acrylate, ethoxypolyethylene glycol (meth)acrylate, etc. can be cited. From the viewpoint of the storage stability of the polymer, methoxypolyethylene glycol (meth)acrylate and ethoxypolyethylene glycol (meth)acrylate are preferred, and methoxypolyethylene glycol methyl acrylate is more preferred.

[0066] As an embodiment of the polymer of the present invention, a copolymer composed of structural unit (1) and structural unit (4) (hereinafter sometimes simply referred to as "copolymer (1-4)") may be included. In the copolymer (1-4), with respect to a total of 100 moles of structural unit (1) and structural unit (4) (i.e., a total of 100 moles of monomer (1) and monomer (4) used in the polymerization), the molar ratio of structural unit (1) (i.e., the molar ratio of monomer (1) used in the polymerization) is preferably 40 to 95 moles, more preferably 60 to 95 moles, and further preferably 80 to 95 moles; the molar ratio of structural unit (4) (i.e., the molar ratio of monomer (4) used in the polymerization) is preferably 5 to 60 moles, more preferably 5 to 40 moles, and further preferably 5 to 20 moles.

[0067] The weight average molecular weight of the copolymer (1-4) is not particularly limited, and is preferably 100,000 to 1,000,000, more preferably 200,000 to 1,000,000, and further preferably 300,000 to 800,000.

[0068] Within the scope not impairing the effects of the present invention, the polymer of the present invention may also contain other structural units derived from other monomers different from the above monomers (1) to (4). Only one kind of other monomer may be used, or two or more kinds may be used in combination. There is no particular limitation on the other monomer, and examples thereof include benzyl (meth)acrylate, isobornyl (meth)acrylate, etc. The amount of the other structural unit in the polymer of the present invention is preferably 20 mol% or less relative to the total structural units. The polymer of the present invention more preferably does not contain other structural units.

[0069] In one embodiment of the present invention, the polymer of the present invention is preferably at least one selected from the group consisting of homopolymer (1), copolymer (1-2), copolymer (1-3), copolymer (1-4), and copolymer (1-2-3), more preferably at least one selected from the group consisting of homopolymer (1), copolymer (1-2), copolymer (1-4), and copolymer (1-2-3), even more preferably homopolymer (1), copolymer (1-2), copolymer (1-3), or copolymer (1-2-3). As the monomers (1) to (4) for forming the structural units (1) to (4) in this embodiment, the above-mentioned monomers can be cited.

[0070] The polymer of the present invention can be produced according to a known method (for example, the method described in International Publication No. 2018 / 216628, etc.).

[0071] From the viewpoint of the stabilization effect, the content of the polymer of the present invention in the whole dry composition for nucleic acid amplification of the present invention is preferably 0.01% by mass to 10% by mass, more preferably 0.05% by mass to 5% by mass, and even more preferably 0.1% by mass to 1.0% by mass. In addition, when two or more kinds of the polymers of the present invention are used, the content means the total of the contents of two or more kinds of the polymers of the present invention. The same applies to the content or amount described for the following other components. When two or more kinds of this component are used, the content or amount means the total of the contents or amounts of two or more kinds of this component.

[0072] [Other components] The dry composition of the present invention contains components other than the polymer of the present invention (in this specification, there are cases where it is described as "other components"). As other components, known components for nucleic acid amplification methods represented by the PCR method can be used. As other components, for example, polymerase, primer, matrix, fluorescent DNA staining reagent, fluorescent probe, passive reference, salt, surfactant, protein, nucleic acid, etc. can be cited. Each of the other components can be used alone or two or more kinds can be used in combination.

[0073] As the polymerase, known DNA polymerases can be used. From the viewpoint of heat resistance, enzymes derived from thermophilic bacteria, thermophilic archaea, hyperthermophilic bacteria, hyperthermophilic archaea and their mutant enzymes are preferred. The DNA polymerase can be appropriately selected from DNA-dependent DNA polymerases, RNA-dependent DNA polymerases or enzymes having both functions according to the purpose of nucleic acid amplification. In addition, either a DNA polymerase having nuclease activity or a DNA polymerase not having nuclease activity can be appropriately selected and used.

[0074] The dry composition of the present invention preferably contains a polymerase. When the dry composition of the present invention contains a polymerase, its content (i.e., the amount of polymerase (U) per 1 mg of dry composition) is preferably 0.01 to 3.0 U / mg, more preferably 0.05 to 2.0 U / mg, and further preferably 0.1 to 1.0 U / mg.

[0075] The primer is not particularly limited. For example, oligonucleotides of about 15 to 30 bases designed / formulated according to known methods can be cited. The primer can be modified with an appropriate fluorescent dye such as fluorescein (FAM). One kind of primer can be used alone, or two kinds of primers can be used as a pair. In order to amplify multiple regions simultaneously, multiple primers can also be used.

[0076] The dry composition of the present invention preferably contains a primer. When the dry composition of the present invention contains a primer, its content (i.e., the amount of primer (pmol) per 1 mg of dry composition) is preferably 1.0 to 10.0 pmol / mg, more preferably 2.0 to 8.0 pmol / mg, and further preferably 3.0 to 6.0 pmol / mg.

[0077] The matrix is not particularly limited. For example, a mixture of deoxyadenosine triphosphate (dATP), deoxythymidine triphosphate (dTTP), deoxyguanosine triphosphate (dGTP), and deoxycytidine triphosphate (dCTP) (dNTPs) can be cited. Among them, part and / or all of dTTP can be replaced with deoxyuridine triphosphate (dUTP).

[0078] As the fluorescent DNA staining reagent, there is no particular limitation. For example, SYBR TM Green I etc. can be cited. As the fluorescent probe, there is no particular limitation. For example, TaqMan TM probe can be cited. The passive reference can be appropriately selected according to the purpose of nucleic acid amplification. As the passive reference, for example, ROX TM Dye etc. can be cited.

[0079] As the salt, there is no particular limitation. For example, salts of organic bases such as tris(hydroxymethyl)aminomethane, tris(hydroxymethyl)methylglycine (tricine), N,N-bis(2-hydroxyethyl)glycine (Bicine) and acids such as sulfuric acid, hydrochloric acid, acetic acid, phosphoric acid, etc. can be cited. In addition, as the salt, magnesium salts (such as magnesium chloride), manganese salts (such as manganese acetate), potassium chloride, ammonium sulfate, etc. can also be used.

[0080] As the surfactant, there is no particular limitation. For example, polyoxyethylene sorbitan fatty acid ester, polyoxyethylene alkyl phenyl ether can be cited. As the protein, there is no particular limitation. For example, bovine serum albumin can be cited.

[0081] As the nucleic acid, in addition to the above primers and fluorescent probes, for example, as an exogenous control gene, any DNA and / or RNA can also be used. Among them, the nucleic acid can be a nucleic acid synthesized in vitro (in vitro), or a nucleic acid prepared from cells, microorganisms, viruses, etc. by a known method. Among them, the cells, microorganisms, viruses, etc. can be substances collected from nature, the environment, humans or animals and plants, and can also be substances that have been isolated / cultured.

[0082] The dry composition of the present invention may further contain oils such as mineral oil; stationary phase carriers (carriers) such as glass beads and magnetic beads.

[0083] In addition, products of the kit type that select and combine multiple of the above components, products of the master mix (Master Mix) type that further mix these components in advance (there are also cases where it is called primer mixture, premix, etc.) can also be used.

[0084] [Manufacturing method of the dry composition of the present invention] The dry composition of the present invention can be manufactured, for example, by drying a solution containing the polymer of the present invention, other components (especially polymerase and / or primer) and water. As the drying method for this, for example, a method of drying at room temperature and atmospheric pressure (air drying method), a method of drying at a temperature above room temperature and a pressure of 90% or less of atmospheric pressure (evaporation drying method), a method of freezing the solution and drying at low temperature and high vacuum (freeze drying method), etc. can be cited. The drying method is preferably the freeze drying method.

[0085] The solution for producing the dry composition of the present invention may also contain a buffer. There is no particular limitation on the buffer. For example, a buffer having a pH of 6 to 9, more preferably about 7 to 8, prepared by mixing an organic base such as tris(hydroxymethyl)aminomethane, tris(hydroxymethyl)methylglycine, N,N-bis(2-hydroxyethyl)glycine with an acid such as sulfuric acid, hydrochloric acid, acetic acid, phosphoric acid, etc. can be cited. In addition, the buffer is desirably appropriately contained with a magnesium salt (e.g., magnesium chloride) and / or a manganese salt (e.g., manganese acetate). In addition, the buffer may further contain salts such as potassium chloride and ammonium sulfate. In addition, the buffer may further contain water-soluble organic solvents such as dimethyl sulfoxide, dimethylformamide, formamide, and glycerol. In addition, the buffer may further contain surfactants such as polyoxyethylene sorbitan fatty acid esters and polyoxyethylene alkyl phenyl ethers. In addition, the buffer may further contain proteins such as bovine serum albumin.

[0086] [Nucleic Acid Amplification Method of the Present Invention] The present invention also provides a nucleic acid amplification method, which includes mixing the dry composition of the present invention, the nucleic acid to be amplified, and water to prepare a reaction solution, and performing a nucleic acid amplification reaction using the obtained reaction solution. The reaction solution can be produced, for example, by mixing a sample containing the nucleic acid to be amplified and water (i.e., nucleic acid aqueous solution) with the dry composition of the present invention.

[0087] The nucleic acid to be amplified can be any of DNA or RNA. The description of the nucleic acid amplification method is as described above. The nucleic acid amplification method of the present invention is preferably polymerase chain reaction (PCR method), and more preferably quantitative polymerase chain reaction. The nucleic acid amplification method (especially the PCR method) is well-known to those skilled in the art, and those skilled in the art can appropriately perform it. Examples

[0088] Hereinafter, the present invention will be specifically described by way of examples, etc., but the present invention is not limited by these.

[0089] [Synthesis Example 1] Weigh 40.0 g of 2-methacryloyloxyethyl phosphorylcholine (hereinafter referred to as "MPC") as monomer (1) in a glass flask for polymerization. Add 60.0 g of purified water to dissolve monomer (1), and add 0.31 g of PEROYL SA (manufactured by NOF Corporation, hereinafter referred to as "PRSA") as a polymerization initiator to the resulting solution. After thoroughly purging the inside of the reaction vessel with nitrogen, polymerization is carried out by heating at 70 °C for 6 hours with stirring. The resulting reaction solution is cooled, and the polymer is precipitated by dropping it into acetone. The precipitate is filtered, washed with acetone, and then dried under vacuum to obtain a white powder homopolymer (hereinafter referred to as "polymer 1"). As determined by gel permeation chromatography (hereinafter referred to as "GPC") under the conditions described below, the weight-average molecular weight of polymer 1 in terms of polyethylene glycol is 1,030,000.

[0090] [Synthesis Example 2] Weigh 8.4 g of MPC as monomer (1), 2.1 g of butyl methacrylate (hereinafter referred to as "BMA") as monomer (2), and 4.5 g of glycerol monomethacrylate (hereinafter referred to as "GLM") as monomer (3) (monomer (1) / monomer (2) / monomer (3) = 40 / 40 / 20 (molar ratio)) in a glass flask for polymerization. Add 42.5 g of purified water and 42.5 g of ethanol to dissolve monomers (1) to (3), and add 0.15 g of PRSA to the resulting solution. Thereafter, a copolymer (hereinafter referred to as "polymer 2") is prepared in the same manner as in Synthesis Example 1. As determined by GPC under the conditions described below, the weight-average molecular weight of polymer 2 in terms of polyethylene glycol is 22,000.

[0091] [Synthesis Example 3] Weigh 6.0 g of MPC as monomer (1) and 4.0 g of methacrylic acid (hereinafter referred to as "MAc") as monomer (2) (monomer (1) / monomer (2) = 30 / 70 (molar ratio)) in a glass flask for polymerization. Add 90.0 g of purified water to dissolve monomers (1) and (2), and add 0.78 g of PRSA to the resulting solution. Thereafter, a copolymer (hereinafter referred to as "polymer 3") is prepared in the same manner as in Synthesis Example 1. As determined by GPC under the conditions described below, the weight-average molecular weight of polymer 3 in terms of polyethylene glycol is 680,000.

[0092] [Synthesis Example 4] Weigh 25.3 g of MPC as monomer (1) and 4.7 g of methoxypolyethylene glycol methacrylate (BLEMMER PME, manufactured by NOF Corporation) (number average molecular weight: approximately 500, n in formula (4): approximately 9, hereinafter referred to as "PEGMA") as monomer (4) in a glass flask for polymerization (monomer (1) / monomer (4) = 90 / 10 (molar ratio)). Add 70.0 g of purified water, dissolve monomers (1) and (4), and add 0.23 g of PRSA to the resulting solution. Thereafter, a copolymer (hereinafter referred to as "Polymer 4") was prepared in the same manner as in Synthesis Example 1. By GPC measurement under the conditions described below, the weight average molecular weight of Polymer 4 was 501,000 in terms of polyethylene glycol conversion.

[0093] [Synthesis Example 5] Weigh 6.0 g of MPC as monomer (1) and 3.3 g of stearyl methacrylate (hereinafter referred to as "SMA") as monomer (2) in a glass flask for polymerization (monomer (1) / monomer (2) = 80 / 20 (molar ratio)). Add 85.0 g of ethanol, dissolve monomers (1) and (2), and add 0.06 g of azobisisobutyronitrile as a polymerization initiator to the resulting solution. Thereafter, a copolymer (hereinafter referred to as "Polymer 5") was prepared in the same manner as in Synthesis Example 1. By GPC measurement under the conditions described below, the weight average molecular weight of Polymer 5 was 43,000 in terms of polyethylene glycol conversion.

[0094] [GPC Measurement] The GPC measurements of Polymers 1 to 5 prepared in Synthesis Examples 1 to 5 were carried out under the following conditions. GPC system: EcoSEC system (manufactured by Tosoh Corporation) Chromatographic column: Shodex OHpak SB-802.5HQ (manufactured by Showa Denko K.K.) and SB-806HQ (manufactured by Showa Denko K.K.) connected in series Developing solvent: 20 mM sodium phosphate buffer (pH 7.4) Detector: Differential refractive index detector Molecular weight standard: EasiVial PEG / PEO (manufactured by Agilent Technologies) Flow rate: 0.5 mL / minute Column temperature: 40 °C Sample: Dilute the resulting polymer with the developing solvent to a final concentration of 0.1 wt% Injection volume: 100 μL

[0095] The monomers used in Synthesis Examples 1 to 5, their molar ratios, and the weight average molecular weights of the resulting polymers are summarized in Table 1.

[0096] [Table 1] Molar ratio of monomers = monomer (1) / monomer (2) / monomer (3) / monomer (4) MPC: 2-methacryloyloxyethyl phosphorylcholine BMA: Butyl methacrylate GLM: Glycerol monomethacrylate MAc: Methacrylic acid PEGMA: Methoxypolyethylene glycol methacrylate SMA: Stearyl methacrylate

[0097] [Examples 1 - 4] Examples showing the case where the nucleic acid to be amplified is double-stranded DNA and the nucleic acid amplification method is the PCR method are presented.

[0098] <Preparation of Solution A> Mix the components shown in Table 2 to prepare Solution A. Additionally, the preparation of this Solution A is carried out while cooling with ice.

[0099] [Table 2] (1) Addition amount: The addition amount per well (2) 1x The concentration obtained by diluting the colorless G0Taq Flexi buffer 5-fold

[0100] <Preparation of the composition for nucleic acid amplification before drying> Add the polymers obtained in Synthesis Examples 1 - 4 to the resulting Solution A to prepare the composition for nucleic acid amplification. Specifically, in Examples 1, 2, or 4, add 2 μL of an aqueous solution of Polymer 1, 2, or 4 with a concentration of 0.5 w / v% to Solution A per well to prepare a 15 μL composition for nucleic acid amplification (the concentration of the polymer in each composition: 0.067 w / v%). In addition, in Example 3, add 2 μL of an aqueous solution of Polymer 3 with a concentration of 0.05 w / v% to Solution A per well to prepare a 15 μL composition for nucleic acid amplification (the concentration of the polymer in the composition: 0.0067 w / v%). Additionally, the preparation of these compositions is carried out while cooling with ice.

[0101] <Preparation of the dried composition for nucleic acid amplification> The obtained composition for nucleic acid amplification was placed in a PCR tube (manufactured by Eppendorf), and the tube was placed in an ultra-low temperature freezer (a freezing ultra-low temperature bath manufactured by Nihon Freezer). The composition was frozen by storing it at -80°C for 30 minutes. Subsequently, the PCR tube containing the frozen composition was placed in a freeze dryer ("FDU-2100" manufactured by Tokyo Rika Kikai Co., Ltd.), and the composition was freeze-dried (sublimation drying) to prepare a dried composition for nucleic acid amplification. Freeze-drying was performed at a reduced pressure of 15 kPa to 20 kPa and a temperature of 22°C for 24 hours. The content of each polymer in the obtained dried composition was 0.4% by mass, the content of the polymerase was 0.2 U / mg, and the content of the primers (i.e., the total content of the first and second primers) was 4.8 pmol / mg.

[0102] Using the dried composition immediately after freeze-drying or the dried composition stored for 3 months, the following operations were performed to measure the band intensity of the nucleic acid amplification product.

[0103] <Storage of the dried composition> The storage of the dried composition was carried out as follows. Specifically, after freeze-drying, the PCR tube was taken out from the freeze dryer, and the PCR tube was directly sealed with Parafilm to prevent the dried composition from absorbing moisture. The sealed PCR tube containing the dried composition was stored at a temperature of 37°C for 3 months.

[0104] <Preparation of a composition for nucleic acid amplification from the dried composition> To the dried composition immediately after freeze-drying or the dried composition stored for 3 months, 15 μL of nuclease-free distilled water (manufactured by NIPPON GENE) was added per well to dissolve the dried composition and prepare a composition for nucleic acid amplification. In addition, the preparation of these compositions was carried out while cooling with ice.

[0105] <Preparation of the PCR reaction solution> An aqueous solution of the target nucleic acid with a nucleic acid concentration of 10 ng / μL was added to the composition for nucleic acid amplification in an amount of 5 μL per well to prepare a PCR reaction solution (the concentration of the polymer in each PCR reaction solution: 0.05 w / v% in Examples 1, 2, and 4, and 0.005 w / v% in Example 3). In addition, the preparation of the PCR reaction solution was carried out while cooling with ice.

[0106] <pcr> PCR was performed using a qPCR device (the "StepOnePlus Real-Time PCR System" manufactured by Applied Biosystems) with the PCR reaction solution prepared as described above. The reaction conditions (temperature program) are as described in Table 3 below. TM The reaction conditions (temperature program) are as described in Table 3 below.

[0107] [Table 3]

[0108] <Electrophoresis> The obtained PCR products were electrophoresed as follows to observe the amount of DNA (band concentration). First, 4 μL of 6x loading buffer (manufactured by NIPPON GENE) was added to 20 μL of the obtained PCR products to prepare an electrophoresis sample. In addition, an electrophoresis gel was prepared using Agarose S (manufactured by NIPPON GENE). In addition, 1x TEA (manufactured by NIPPON GENE) was used as the electrophoresis buffer. The marker (Gene Ladder 100 (0.1 - 2 kbp), manufactured by NIPPON GENE) and the electrophoresis sample were added to the prepared electrophoresis gel at 4 μL / well, respectively, and the resulting mixture was electrophoresed at 100 V for 30 minutes. After that, the sample components were stained with Midori Green Advance (manufactured by Genetics), then decolorized with nuclease-free distilled water (manufactured by NIPPON GENE), and the band of the target nucleic acid amplification product (size: 1.3 kbp) was detected using a gel imaging device (Fas-Digi Compact, manufactured by Nippon Genetics). Using Image J (manufactured by the National Institutes of Health, USA), the band intensity obtained using the dried composition immediately after lyophilization or after storage for 3 months was calculated. The band intensity (relative value) obtained using the dried composition after storage for 3 months when the band intensity obtained using the dried composition immediately after lyophilization was 100% is shown in Table 4 below. In addition, Figure 1 the results of electrophoresis using the dried compositions stored for 3 months in Examples 1 - 4 are shown. In addition, Figure 1 the results of electrophoresis using the marker, electrophoresis without adding the target nucleic acid solution, and electrophoresis using the dried composition stored for 3 months in Comparative Example 1 described below are also shown.

[0109] [Comparative Example 1] In <Preparation of the dry composition for nucleic acid amplification>, except for using 2 μL of nuclease-free distilled water (manufactured by NIPPONGENE) instead of 2 μL of the aqueous solution of the polymer, the same operations as in Examples 1 to 4 were carried out in Comparative Example 1. The band intensities (relative values) of the dry composition obtained after 3 months of use and storage when the band intensity of the dry composition immediately after freeze-drying was 100% are shown in Table 4 below. In addition, Figure 1 The results of electrophoresis using the dry composition after 3 months of use and storage are also shown.

[0110] [Table 4] Polymer used Band intensity (relative value) Example 1 Polymer 1 36％ Example 2 Polymer 2 44％ Example 3 Polymer 3 31％ Example 4 Polymer 4 51％ Comparative Example 1 - 0 Band intensity (relative value) = Band intensity (relative value) of the dry composition obtained after 3 months of use and storage when the band intensity of the dry composition immediately after freeze-drying was 100%

[0111] As shown in Table 4 and Figure 1 shown, no bands were detected in the electrophoresis using the dry composition of Comparative Example 1 that does not contain Polymers 1 to 4 after 3 months of storage. In contrast, bands were detected in the electrophoresis using the dry composition of Examples 1 to 4 that contain any one of Polymers 1 to 4 after 3 months of storage. In particular, in Example 4, the band intensity (relative value) of the dry composition obtained after 3 months of use and storage was 51%. From these results, it can be seen that the storage stability of the dry composition for nucleic acid amplification of the present invention is excellent.

[0112] [Examples 5 to 8] Examples are shown when the nucleic acid to be amplified is single-stranded RNA and the nucleic acid amplification method is the RT-qPCR method.

[0113] <Preparation of Solution B> Mix the components shown in Table 5 to prepare Solution B. In addition, the preparation of this Solution B is carried out while cooling with ice.

[0114] [Table 5] (1) Added amount: Added amount per well (2) 1x: Concentration obtained by diluting the RapiDxFire Lyo-Flex 1-Step RT-qPCR 5X master mix 5-fold

[0115] <Preparation of the composition for nucleic acid amplification before drying> To the obtained Solution B, add the polymers prepared in Synthesis Example 1 or 3 to 5 to formulate a composition for nucleic acid amplification. Specifically, in Example 5, 7, or 8, to each well of Solution B, add 2 μL of an aqueous solution of Polymer 1, 4, or 5 with a concentration of 0.5 w / v% to formulate a 15-μL composition for nucleic acid amplification (the concentration of the polymer in each composition: 0.067 w / v%). In addition, in Example 6, to each well of Solution B, add 2 μL of an aqueous solution of Polymer 3 with a concentration of 0.05 w / v% to formulate a 15-μL composition for nucleic acid amplification (the concentration of the polymer in the composition: 0.0067 w / v%). Additionally, the formulation of these compositions is carried out while cooling with ice.

[0116] <Preparation of a dried composition for nucleic acid amplification> Put the obtained composition for nucleic acid amplification into a PCR tube (manufactured by Eppendorf), place this tube into an ultra-low temperature freezer (a freezing ultra-low temperature bath manufactured by Nihon Freezer), and store it at -80°C for 30 minutes to freeze the composition. Then, put the PCR tube containing the frozen composition into a freeze dryer ("FDU-2100" manufactured by Tokyo Rika Kikai Co., Ltd.) to perform freeze drying (sublimation drying) of the composition to prepare a dried composition for nucleic acid amplification. The freeze drying is carried out at a reduced pressure of 15 kPa to 20 kPa and a temperature of 22°C for 24 hours. The content of each polymer in the obtained dried composition is 0.4% by mass, and the content of the primers (i.e., the total content of the forward primer and the reverse primer) is 4.0 pmol / mg. Using the dried composition just after freeze drying or the dried composition stored for 12 days, perform the following operations to measure the fluorescence intensity of the nucleic acid amplification product.

[0117] <Storage of the dried composition> The storage of the dried composition is carried out as follows. Specifically, after freeze drying, take out the PCR tube from the freeze dryer and directly seal the PCR tube with Parafilm so that the dried composition does not absorb moisture. Store the sealed PCR tube containing the dried composition at a temperature of 60°C for 12 days.

[0118] <Formulation of a composition for nucleic acid amplification from the dried composition> To the dried composition just after freeze drying or the dried composition stored for 12 days, add 15 μL of nuclease-free distilled water (manufactured by NIPPON GENE) to each well to dissolve the dried composition and formulate a composition for nucleic acid amplification. Additionally, the formulation of these compositions is carried out while cooling with ice.

[0119] <Formulation of the RT-qPCR reaction solution> An aqueous solution of a target nucleic acid with a nucleic acid concentration of 100 copies / μL was added to the composition for nucleic acid amplification in an amount of 5 μL per well to prepare an RT-qPCR reaction solution (the concentration of the polymer in each RT-qPCR reaction solution: 0.05 w / v% in Examples 5, 7, and 8, and 0.005 w / v% in Example 6). In addition, the preparation of the PCR reaction solution was carried out while cooling with ice.

[0120] <rt-qpcr> Using the qPCR device (the "StepOnePlus Real-Time PCR System" manufactured by Applied Biosystems) with the RT-qPCR reaction solution prepared as above, RT-qPCR was performed to measure the fluorescence intensity at the end point of the RT-qPCR reaction solution. The reaction conditions (temperature program) are as described in Table 6 below. TM The fluorescence intensity (relative value) of the dried composition after 12 days of use and storage, when the fluorescence intensity obtained using the dried composition just after lyophilization is taken as 100%, is shown in Table 7 below.

[0121] [Table 6]

[0122] The fluorescence intensity (relative value) of the dried composition after 12 days of use and storage, when the fluorescence intensity obtained using the dried composition just after lyophilization is taken as 100%, is shown in Table 7 below.

[0123] [Comparative Example 2] In <Preparation of the Dried Composition for Nucleic Acid Amplification>, except that 2 μL of nuclease-free distilled water (manufactured by NIPPONGENE) was used instead of 2 μL of the aqueous solution of the polymer, the same operations as in Examples 5 to 8 were carried out in Comparative Example 2. The fluorescence intensity (relative value) of the dried composition after 12 days of use and storage, when the fluorescence intensity obtained using the dried composition just after lyophilization is taken as 100%, is shown in Table 7 below.

[0124] [Table 7] Polymer used Fluorescence intensity (relative value) Example 5 Polymer 1 54％ Example 6 Polymer 3 79％ Example 7 Polymer 4 82％ Example 8 Polymer 5 66％ Comparative Example 2 - 52％ Fluorescence intensity (relative value) = The fluorescence intensity (relative value) of the dried composition after 12 days of use and storage, when the fluorescence intensity obtained using the dried composition just after lyophilization is taken as 100%

[0125] As shown in Table 7, the dried compositions of Examples 5 to 8 containing Polymer 1 or 3 to 5 had higher fluorescence intensity (relative value) after 12 days of storage compared to the dried composition of Comparative Example 2 that did not contain these polymers. In particular, the fluorescence intensity (relative value) of the dried composition after 12 days of use and storage in Example 7 was 82%. From these results, it can be seen that the storage stability of the dried composition for nucleic acid amplification of the present invention is excellent. Industrial Applicability

[0126] The dried composition of the present invention is useful for nucleic acid amplification methods (especially quantitative polymerase chain reaction) used in gene testing, microbial testing, virus testing, etc.

[0127] This application is based on Japanese Patent Application No. 2022-185199, the content of which is incorporated herein in its entirety. < / pcr>

Claims

1. A dry composition for nucleic acid amplification, which comprises a polymer containing a structural unit derived from a monomer represented by formula (1), wherein, X 1 represents (meth)acryloyloxy or (meth)acrylamino, L 1 represents an alkylene group having 2 to 4 carbon atoms with or without 1 hydroxyl group, or an alkyleneoxyalkylene group having 2 to 4 carbon atoms, and R 1 ~R 3 Each independently represents an alkyl group having 1 to 3 carbon atoms.

2. The dry composition for nucleic acid amplification according to claim 1, wherein the polymer is a homopolymer composed of one structural unit derived from the monomer represented by formula (1).

3. The dry composition for nucleic acid amplification according to claim 1, wherein the polymer is a copolymer further containing a structural unit derived from the monomer represented by formula (2), wherein, R 4 represents a hydrogen atom or a methyl group, and R 5 represents a hydrogen atom or an alkyl group having 1 to 20 carbon atoms.

4. The dry composition for nucleic acid amplification according to claim 3, wherein the polymer is a copolymer further containing a structural unit derived from the monomer represented by formula (3), wherein, R 6 represents a hydrogen atom or a methyl group, and R 7 represents an alkyl group having 3 to 6 carbon atoms and having two or more hydroxyl groups.

5. The dry composition for nucleic acid amplification according to claim 1, wherein the polymer is a copolymer further containing a structural unit derived from the monomer represented by formula (4), wherein, R 8 represents a hydrogen atom or a methyl group, n represents a number from 1 to 10, and R 9 represents a hydrogen atom, a methyl group or an ethyl group.

6. The dry composition for nucleic acid amplification according to any one of claims 1 to 5, which further contains a polymerase.

7. The dry composition for nucleic acid amplification according to claim 6, which is used for quantitative polymerase chain reaction.

8. The dry composition for nucleic acid amplification according to claim 7, which further contains primers.

9. A nucleic acid amplification method, which comprises mixing the dry composition for nucleic acid amplification according to any one of claims 1 to 5, the nucleic acid to be amplified and water to prepare a reaction solution, and using the obtained reaction solution to carry out a nucleic acid amplification reaction.

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