Y-probe and variation thereof, and DNA microarray, kit, and gene analysis method using the Y-probe and the variation thereof

A gene analysis and micro-display technology, applied in the field of gene analysis and DNA micro-display, to achieve the effects of reducing space obstruction, accurate statistical analysis, and improving sensitivity and specificity

Active Publication Date: 2013-05-08
GOODGENE
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0019] The object of the present invention is to provide a novel Y-type probe and its modified form in which the target gene to be detected and the gene of the reference standard substance are used as probes together at each point, thereby improving conventional oligonucleotide probes. , and solve the problems of existing oligonucleotide microdisplays, and actually apply to clinical diagnosis

Method used

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  • Y-probe and variation thereof, and DNA microarray, kit, and gene analysis method using the Y-probe and the variation thereof
  • Y-probe and variation thereof, and DNA microarray, kit, and gene analysis method using the Y-probe and the variation thereof
  • Y-probe and variation thereof, and DNA microarray, kit, and gene analysis method using the Y-probe and the variation thereof

Examples

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Embodiment 1

[0125] Embodiment 1: Design of Y-type probe

[0126] The process that plays the most important role in the development of the DNA chip is to develop the structure of the double oligonucleotide probe in the Y shape of the present invention. Among them, the following steps are included: a step of pasting the connecting peptide for placing the probe on a solid support (solid support, features); a step of pasting a spacer and pasting a labeling dye for signal observation.

[0127] The Y-type probe of the present invention is a continuous oligonucleotide, which is in the form of a Y-shaped branch, and has a tree-like structure in which two different oligonucleotide probes are placed on the stem region. Like the root of the tree planted on the ground, the part where the above-mentioned probe is placed on a fixed carrier such as a glass slide is called a connecting peptide or a spacer. The sample falls on the tree like snow. For the sample, if the DNA or RNA with the sequence comple...

Embodiment 2

[0167] Embodiment 2: the synthesis of Y type probe

[0168] The Y-type probe designed as in Example 1 can be synthesized through the following process. The synthesis process of Y-type oligonucleotide probes is divided into 1) removal of trityl (detritylation, DMT removal), 2) coupling, 3) capping (capping) and 4) oxidation (oxidation), in a A nucleotide is bound during the cycle. Therefore, an oligomer can be synthesized by allowing dA, dG, dT, and dC to participate in each reaction in the order of the base sequence to be synthesized. When the synthesis is completed, ammonium hydroxide is added to deprotect the oligomer from the carrier. The oligonucleotide firmly binds the nucleotide at the 3' end to the solid phase support so that the oligonucleotide does not move the nucleotide at the 3' end, and then reacts on the chromatography column to achieve synthesis. Therefore, synthesis is carried out from the 3' to 5' direction. Controlled pore glass (CPG, controlled pore glas...

Embodiment 3-8

[0209] Example 3-8: Development of DNA Microarray for HPV Diagnosis

[0210] The present invention relates to a novel method for diagnosing human papillomavirus (human papillomavirus, HPV) infection by using DNA micro-array of spotted Y-type probes. The present embodiment 3 to embodiment 8 include the following steps: taking HPV diagnosis as an example, the step of preparing a Y-type probe (embodiment 3); ); the step of labeling after separating DNA from the specimen (Example 5); the step of hybridization reaction (Example 6); after the reaction, the step of analyzing its signal (Example 7); Steps in clinical diagnosis (Example 8). The present Examples 3 to 8 show an example related to the utilization method of the Y-type probe, and show that the DNA microarray using the Y-type probe is useful for the diagnosis of important diseases.

[0211] The genome of HPV is composed of double-helix DNA, and there are early protein genes from E1 to E7 and late protein genes from L1 and ...

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Abstract

The present invention relates to a Y-type nucleotide probe having two probe portions on a single body such that it can be widely used for genotype testing and analysis with improved sensitivity, specificity, and accuracy, as well as to a DNA microarray, kit, and gene analysis method using the y-probe. The Y-probe of the invention is configured with five portions, comprising: a left probe portion, a left stem portion, a linker, a right stem portion, and a right probe portion. The DNA microarray of the present invention may have improved testing accuracy as it can perform a simultaneously overlapping test on the same gene or a simultaneous test on two different target genes. In particular, by simultaneously testing a target gene and a control gene in one spot, analysis errors can be reduced, quantitative analysis is made possible, and standardization is facilitated. The Y-probe of the present invention can be used for genotype analysis and gene expression analysis, as well as for mutation and SNP analysis, and can thus be broadly used for the diagnosis and prognosis of diseases and for genetic diagnosis for determining a course of treatment, etc.

Description

technical field [0001] The present invention relates to a Y-type nucleotide probe (probe) having two probe sites in one body, which can be widely used in diagnosis by improving sensitivity, specificity and accuracy in genotype detection and analysis, and Its deformed type (d-type or b-type probe), and a DNA microarray, kit and gene analysis method using the Y-type nucleotide probe. Background technique [0002] DNA microdisplay or DNA chip is to spot dozens to hundreds of millions of gene probes on a solid support such as a glass slide in the form of spots. DNA, RNA, cDNA, cRNA, microRNA, polymerase chain reaction (polymerase chain reaction) labeled with fluorescent substances (fluorescent dye) after extraction from tissues, cells, body fluids and other specimens are placed on the DNA microarray. reaction; PCR) products and other nucleic acids, and perform hybridization reactions or sequencing (sequencing) reactions, the signals of the marker substances that appear in the r...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/11C07H21/00C12Q1/68G01N33/52
CPCC12Q2600/156C12Q1/6816C12Q1/708C12Q1/6888C12Q1/6886C12Q1/701C12Q1/689C12Q1/6837C12Q2565/543C12Q2525/107
Inventor 文宇哲吴明烈
Owner GOODGENE
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