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Compound having structure derived from mononucleoside or mononucleotide, nucleic acid, labeling substance, and method and kit for detection of nucleic acid

A single nucleotide and compound technology, applied in biological testing, biochemical equipment and methods, sugar derivatives, etc., can solve problems such as increased fluorescence intensity

Active Publication Date: 2010-01-20
DNAFORM
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Although an example of making a fluorescent probe by covalently bonding thiazole orange to DNA has been attempted, it still emits strong fluorescence through the interaction with single-stranded DNA containing purine bases (Non-Patent Document 5), so in The increase in fluorescence intensity when forming a double helix is ​​small and cannot be said to be successful (Non-Patent Documents 6 and 7)

Method used

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  • Compound having structure derived from mononucleoside or mononucleotide, nucleic acid, labeling substance, and method and kit for detection of nucleic acid
  • Compound having structure derived from mononucleoside or mononucleotide, nucleic acid, labeling substance, and method and kit for detection of nucleic acid
  • Compound having structure derived from mononucleoside or mononucleotide, nucleic acid, labeling substance, and method and kit for detection of nucleic acid

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preparation example Construction

[0261] The method for producing the compound and nucleic acid of the present invention is not particularly limited, and known synthesis methods (production methods) can be suitably used. For example, in the case of the compound represented by the formula (21), it can be produced by a production method comprising the following steps: after activating the carboxyl group of the compound represented by the following formula (26), it is combined with tris(2-aminoethyl group) amine reaction; protection of amino group; and the reaction of protecting the hydroxyl group present in the compound obtained above with a protecting group, and the reaction of adding a phosphoric acid or phosphoramidite group to the hydroxyl group present in the obtained compound.

[0262] [Chemical 52]

[0263]

[0264] In formula (26), A represents a hydrogen atom or a hydroxyl group. B represents a residue of adenine, guanine, cytosine, thymine or uracil.

[0265] The production method (synthesis metho...

Embodiment 1-3

[0320] According to the following scheme 1, compounds 102 and 103 in which two active amino groups were each protected with a trifluoroacetyl group were synthesized (produced), and phosphoramidite 104 was further synthesized.

[0321] [Chemical 53]

[0322]

[0323] plan 1

[0324] Reagents and reaction conditions (a) (i) N-hydroxysuccinimide, EDC / DMF, (ii) tris(2-aminoethyl)-amine / CH 3 CN, (iii) CF 3 COO Et, Et 3 N; (b) DMTrCl / pyridine; (c) 2-cyanoethyl-N,N,N',N'-tetraisopropylphosphoramidite, 1H-tetrazole / CH 3 CN.

[0325] Regarding Scheme 1 above, it is described in more detail below.

Embodiment 1

[0326] Example 1: 2-[2-[N,N-bis(2-trifluoroacetylaminoethyl)]-aminoethyl]carbamoyl-(E)-vinyl]-2'-deoxyuridine

[0327] Synthesis of (2-[2-[N,N-bis(2-trifluoroacetamidoethyl)]-aminoethyl]carbamoyl-(E)-vinyl)-2′-deoxyuridine, compound 102)

[0328]Starting material (E)-5-(2-carboxyvinyl)-2'-deoxyuridine ((E)-5-(2-carboxyvinyl)-2'-deoxyuridine, compound 101) according to Tetrahedron1987, 43, 20 , 4601-4607 Synthesis. That is, first, 71 mL of 1,4-dioxane was added to 430 mg of palladium(II) acetate (FW224.51) and 1.05 g of triphenylphosphine (FW262.29), and then 7.1 mL of triethylamine (FW 101.19, d=0.726), heated and stirred at 70°C. When the reaction solution changed from reddish brown to dark brown, add 14.2g 2'-deoxy-5-iodouridine (FW354.10) and 7.0mL methyl acrylate (FW 86.09, d=0.956) in 1,4-diox The suspension in alkanes was heated to reflux at 125°C for 1 hour. Afterwards, it was filtered while still hot, the residue was washed with methanol, and the filtrate was recov...

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Abstract

Disclosed is a labeling substance which enables to detect a double-stranded structure in a nucleic acid effectively. Specifically disclosed is: a compound having a structure derived from a mononucleoside or mononucleotide, wherein the structure is represented by the formula (1), (1b) or (1c); a tautomer or stereoisomer of the compound; or a salt of the compound, the tautomer or the stereoisomer. Wherein B represents an atomic group having a nucleobase skeleton; E represents an atomic group having a deoxyribose skeleton, a ribose skeleton or a structure derived from the deoxyribose skeleton or the ribose skeleton, or an atomic group having a peptide structure or a peptoid structure; and Z<11> and Z<12> independently represent a hydrogen atom, a protecting group or a fluorescent atomic group, and may be the same as or different from each other.

Description

technical field [0001] The present invention relates to a compound, a nucleic acid, a marker, a nucleic acid detection method and a kit having a structure derived from a single nucleoside or a single nucleotide. Background technique [0002] In genetic diagnosis of diseases and gene expression analysis, etc., it is necessary to detect nucleic acids having a specific sequence. Methods using fluorescence are often used for this purpose, for example fluorescent probes obtained by covalently binding a fluorescent dye to DNA are commonly used as labels. [0003] A problem with such labels (fluorescent probes) is that they fluoresce, for example without forming a double helix with a complementary nucleic acid. The method of using FRET to eliminate the fluorescence of the probe itself is effective (Non-Patent Documents 1-4, etc.), but there are problems such as the cost of introducing two kinds of fluorescent dyes. [0004] In addition, thiazole orange is known as one of cyanine ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07H19/073C07H21/04G01N21/78G01N33/53G01N33/58C12N15/09C12Q1/68C12N15/11
CPCC09B23/04
Inventor 冈本晃充池田修司久保田健
Owner DNAFORM