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Method for detecting single nucleotide polymorphism (SNP) and point mutation in gene, detection apparatus and detection chip

A detection chip and detection device technology, which can be used in measurement devices, genetic engineering, plant genetic improvement, etc., and can solve problems such as impracticability

Inactive Publication Date: 2001-10-31
内田和彦 +2
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

It is almost impossible to collect and analyze all one-base substitution SNPs and point mutations with existing methods

Method used

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  • Method for detecting single nucleotide polymorphism (SNP) and point mutation in gene, detection apparatus and detection chip
  • Method for detecting single nucleotide polymorphism (SNP) and point mutation in gene, detection apparatus and detection chip
  • Method for detecting single nucleotide polymorphism (SNP) and point mutation in gene, detection apparatus and detection chip

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0044] An example of detection of a base substitution SNP in codon 72 of the gene p53 is shown. The oligonucleotides had base sequences corresponding to the following two polymorphisms (genetic polymorphisms), and the oligonucleotides were immobilized at the points of the respective gold electrodes.

[0045] p53Pro (the 72nd position codon is Pro)

[0046] p53Arg (the 72nd position codon is Arg)

[0047] The 72nd codon of p53 is Pro, the DNA taken from the peripheral blood of normal people and the amplified PCR product are subjected to a hybridization reaction after high-temperature denaturation, and the PCR product is a fragment amplified from this DNA, which includes Codon 72 in p53 exon 4. Electrolyte solution with 0.1MAcOH-AcOK (pH5.6), 0.1M KCl, 0.05mM NFc, at 20 degrees to measure the change of current value before and after hybridization at 470mV (Ag / AgCl reference electrode standard).

[0048] DNA collected from peripheral blood

[0049] Current change (%) of p53Pr...

Embodiment 2

[0063] In this example, the measured current value is different due to the difference in the number of base substitutions, and thus the amount of matching errors can be measured. Seven oligonucleotides of dT20, dT10dAdT9, dT8dA4dT8, dAdT19, dT3dT17, dT19dA, dT17dA3 were immobilized on each point of the gold electrode respectively. The dA20 was subjected to a hybridization reaction.

[0064] Electrolyte solution with 0.1M AcOH-AcOK (pH5.6), 0.1M KCl, 0.05mM NFc, at 20 degrees, with 470mV (Ag / AgCl reference electrode standard) to measure the change of current value before and after hybridization. The measurement results are shown in Table 1.

[0065] dT20

[0066] The current change in Table 1 is almost dependent on the change in the amount of base matching errors. Especially when there is a mismatch at the end, a change larger than the Tm value is seen. Such detection cannot be carried out in the previous SSCP, and this method is the first time to figure it out. ...

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Abstract

Space part S within a detecting chip 2 for single base substitution SNP and point mutation of genes where a plurality of gold electrodes 8 are formed in the base 7 of closed space part S, oligonucleotides 10 with different gene sequences are fixed to the gold electrodes 8, a common electrode 16 arranged not to contact the gold electrodes 8, are filled with DNA samples, voltage is applied between the common electrode 16 and the gold electrode 8, and current is measured to allow the double-stranded DNA to be detected and analyzed. It becomes possible to detect and analyze a large number of single base substitution SNP and point mutation for a plurality of DNA samples.

Description

technical field [0001] The invention relates to aspects such as gene expression and can detect and analyze a base substitution SNP (single nucleotide polymorphism: a variant in the human genetic code) and a point mutation in a DNA gene, that is, a base substitution SNP and a point mutation in a gene A detection method, a detection device and a detection chip device. Background technique [0002] The so-called one base substitution SNP means that there is a base change in the base sequence of human DNA between 1000bp and 2000bp. For humans, whether it is a normal person or a patient, it is generally believed that there are hundreds of thousands or even millions of bases. Therefore, it is expected that it can be used as a marker to uncover the etiology and effectively prevent diseases. [0003] The so-called point mutation is a change in the base sequence of a known gene, and the function of translation into protein is abnormal due to this base change, a...

Claims

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Application Information

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IPC IPC(8): C12N15/09B01L3/00B01L7/00C12M1/00C12Q1/68G01N27/327G01N27/416G01N33/483G01N33/53G01N37/00
CPCB01L2300/0636B01L7/52B01L2300/0809C12Q1/6827B01L2300/0645B01L2300/0877B01L2200/02B01L3/5027B01L3/502715C12Q2565/607C12Q2565/518C12Q1/68G01N33/53C12N15/00
Inventor 宫原孝俊内田和彦竹中繁织
Owner 内田和彦
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