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Method of detecting inorganic phosphoric acid, pyrophosphate and nucleic acid, and method of typing snp sequence of DNA

A technology of inorganic phosphoric acid and detection method, which is used in biochemical equipment and methods, determination/inspection of microorganisms, material analysis by electromagnetic means, etc., and can solve problems such as inability to detect target nucleic acid molecules.

Inactive Publication Date: 2004-11-10
PANASONIC CORP
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  • Abstract
  • Description
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  • Application Information

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Problems solved by technology

However, since the target sample for detection here is L-phenylalanine, this method cannot be used to detect the presence of target nucleic acid molecules

Method used

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  • Method of detecting inorganic phosphoric acid, pyrophosphate and nucleic acid, and method of typing snp sequence of DNA
  • Method of detecting inorganic phosphoric acid, pyrophosphate and nucleic acid, and method of typing snp sequence of DNA
  • Method of detecting inorganic phosphoric acid, pyrophosphate and nucleic acid, and method of typing snp sequence of DNA

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Embodiment approach 1

[0037] In Embodiment 1 of the present invention, simple quantitative detection of inorganic phosphate is performed using an enzyme reaction. Embodiment 1 of the present invention relates to a method for detecting inorganic phosphoric acid, the method comprising: supplying a sample to a sample containing glyceraldehyde-3-phosphate, oxidized nicotinamide adenine dinucleotide (NAD + ) or oxidized nicotinamide phosphate adenine dinucleotide (NADP + ), a step in the measurement system of glyceraldehyde phosphate dehydrogenase and an electron transport mediator; and, a step in which the current value is measured in the system. In the present specification, "measurement system" refers to the whole of a series of reactions of the mechanism involved in the detection of inorganic phosphate and the place where such a series of reactions are carried out. Components necessary for carrying out a series of reactions exist in the "measurement system". The "measurement system" is usually obt...

Embodiment approach 2

[0062] In Embodiment 2 of the present invention, pyrophosphate is simply and quantitatively detected using an enzymatic reaction.

[0063] Embodiment 2 of the present invention will be described using the reaction formula (V).

[0064] First, as shown in the reaction formula (V), pyrophosphoric acid (PPi in the reaction formula (V)) is hydrolyzed to become inorganic phosphoric acid (Pi in the reaction formula (V)).

[0065] Reaction formula (V)

[0066]

[0067] This hydrolysis reaction may be carried out at a high temperature, or may be carried out using pyrophosphatase, which is an enzyme that catalyzes the reaction.

[0068] Next, pyrophosphoric acid can be quantitatively detected by detecting the inorganic phosphoric acid produced by the reaction of the reaction formula (V) in the same manner as in the first embodiment.

[0069] In this embodiment, the reaction of the reaction formula (V) does not need to be performed separately in the reaction in the detection of...

Embodiment approach 3

[0071] In Embodiment 3 of the present invention, a method for detecting DNA having a specific sequence with high sensitivity is implemented.

[0072] In the present embodiment, first, a sample is supplied to a reaction system containing a DNA probe having a sequence complementary to a target DNA sequence, a DNA polymerase, and a deoxynucleotide. The "reaction system" as used herein refers to a series of nucleic acid extension reactions as described below and a place where such reactions are carried out. In a "reaction system", there are components necessary to carry out a series of reactions. A "reaction system" typically dissolves the above components in a suitable solvent (eg, Tris-HCl buffer, any buffer commonly used in nucleic acid elongation reactions or nucleic acid propagation reactions (including those in commercially available kits). buffer)), provided as a solution. The DNA polymerase may be any DNA polymerase that is commercially available or prepared by those ski...

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Abstract

According to the present invention, an inorganic phosphoric acid is detected by a method which includes: subjecting a sample to a measurement system containing glyceraldehyde-3-phosphate, oxidized nicotinamide adenine dinucleotide or oxidized nicotinamide adenine dinucleotide phosphate, glyceraldehyde phosphate dehydrogenase, and an electron mediator; and measuring a current value in the measurement system. In the method, a pyrophosphate is quantitatively measured with high sensitivity and at a high speed through converting the pyrophosphate in a sample into an inorganic phosphoric acid. Such a measurement of a pyrophosphate allows for quantitative determination of the pyrophosphate which is produced concurrent with the extension of a DNA, thereby enabling the detection of the presence of a targeted nucleic acid, and typing of a base in a SNP site of a targeted DNA.

Description

technical field [0001] The present invention relates to a method for detecting inorganic phosphoric acid or pyrophosphoric acid, and also relates to a method for detecting a target nucleic acid in a sample with a simple and high sensitivity using the above-mentioned detection, in particular, a method for discriminating SNP. Background technique [0002] In recent years, the development of technologies to study genetic information has been at its peak. In the medical field, it is possible to treat diseases at the molecular level by analyzing the genes associated with diseases. With genetic diagnosis, specific medical treatment corresponding to each patient can also be performed. In the pharmaceutical field, protein molecules such as antibodies and hormones are identified using genetic information and used as pharmaceuticals. In the fields of agriculture and food, products utilizing various genetic information have also been produced. [0003] Among these genetic informatio...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/68G01N33/84
CPCC12Q1/6827C12Q1/6858G01N33/84C12Q2535/125C12Q2565/301
Inventor 行政哲男夜久英信深日希美
Owner PANASONIC CORP
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