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Substrate, gene chip and preparation method thereof and target detection method

A gene chip, target technology, applied in biochemical equipment and methods, biological testing, material inspection products, etc., can solve problems such as poor detection sensitivity and recognition

Active Publication Date: 2010-09-08
CHANGCHUN INST OF APPLIED CHEMISTRY - CHINESE ACAD OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] The technical problem to be solved by the present invention is to provide a gene chip with higher sensitivity and recognition and a method for detecting targets for the defects of existing biochips with poor detection sensitivity and recognition

Method used

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  • Substrate, gene chip and preparation method thereof and target detection method
  • Substrate, gene chip and preparation method thereof and target detection method
  • Substrate, gene chip and preparation method thereof and target detection method

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preparation example Construction

[0036] The present invention also provides a method for preparing a gene chip, comprising:

[0037] Step 1: Soaking the surface aldehydelated solid support in the polyamide-amine dendrimer solution for 10h to 14h to obtain a solid support covalently bonded to the polyamide-amine dendrimer;

[0038] Step 2: activating the solid support covalently bound to the polyamide-amine type dendrimer with a glutaraldehyde solution with a pH value of 7-7.4;

[0039] Step 3: Take the target probe and apply it on the solid support obtained in Step 2 to obtain a gene chip.

[0040] According to the present invention, put the aldehyde-modified glass slide into 25mL-35mL of the fourth-generation PAMAM methanol solution, the mass percentage of the fourth-generation PAMAM is preferably 0.05%-0.15%, and under slight stirring, the amino terminal on the PAMAM It will react with the aldehyde group on the glass slide. PAMAM is covalently bound to the aldehyde group-modified glass slide. After reactin...

Embodiment 1

[0052] Follow the steps below to prepare detection probe-coupled gold nanoparticles:

[0053] Gold nanoparticles with a diameter of 13nm were prepared by the Turkevich-Frens method. Mix 25 μL of SP aqueous solution with a concentration of 40 μmol / L and 500 μL of 6 nmol / L GNPs solution, let it stand overnight, and then add 500 μL of phosphate buffer solution to obtain the first mixed solution. The pH of the phosphate buffer solution is 7.4, and the phosphate buffer solution is a PBS solution containing 10mmol / L phosphate and 0.2mol / L sodium chloride. After the first mixed solution is left to stand for 2 hours, the solution is preferably concentrated to 150 μL using a vacuum centrifugal concentrator to obtain a concentrated solution. Then centrifuge the concentrated solution 3 times for each 15min at a speed of 9000rad / min, and dissolve the gold nanoparticles coupled with the detection probe obtained after centrifugal concentration in 60mmol / L trisodium citrate, 0.6mol / L sodium...

Embodiment 2

[0055] Follow the steps below to create a 3D base:

[0056] Put the aldehyde-modified glass slide into 30 mL of the fourth-generation PAMAM methanol solution with a mass fraction of 0.1 wt%, and react at room temperature for 12 h under stirring, then wash with 30 mL of methanol for 3 min each time, and then wash with N 2 blow dry. With 30mL, 10mmol / L sodium borohydride (NaBH 4 ) solution to reduce the Schiff base generated in the above reaction. Then, put the substrate modified with dendrimers into 30 mL of glutaraldehyde solution to react for 4 hours. The glutaraldehyde solution preferably includes 5 wt% glutaraldehyde and 95% phosphate buffer solution, and the phosphate The buffer solution is a PBS solution containing 50 mmol / L phosphate and 0.15 mol / L sodium chloride, and the pH value is 7.4. Wash 3 times with 30 mL of the above phosphate buffer solution, 3 min each time, wash 3 times with Milli-Q ultrapure water, 3 min each time, centrifuge and dry for 1 min to obtain a...

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Abstract

The invention relates to the technical field of biology and provides a substrate for a biological chip. The substrate is formed by jointing a polyamide-amine dendritic macromolecule on a solid support through a covalent bond; the polyamide-amine dendritic macromolecule is a dendritic macromolecule which radiates 64 amino terminals with ethidene diamine as a center, and a molecular weight is 14,214 to 14,220; and the polyamide-amine dendritic macromolecule is activated by an aldehyde group. The substrate for the biological chip adds reaction points through the multiple amino terminals of the dendritic macromolecule, so that the sensitivity of the biological chip is enhanced. The invention also provides a gene chip prepared from the substrate, a method for preparing the gene chip and a method for detecting a target by using the gene chip.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to a substrate, a gene chip, a preparation method thereof, and a method for detecting a target. Background technique [0002] Gene chip technology is a high-throughput analysis method widely used in the study of gene expression, protein-gene sequence interaction and single nucleotide polymorphism. In the application process of gene chips, the sensitivity and selectivity of detection are two very important investigation parameters. Therefore, it is necessary to make an ideal one that has high immobilization efficiency and does not interact with other substances in the reaction system non-specifically. Reactive chip substrates become the key to gene chip technology. [0003] A gene chip consists of a substrate and target probes. Substrates can be roughly divided into two categories: 1. Two-dimensional (2D) flat substrates with functional groups such as aldehyde groups, epoxy groups, or ...

Claims

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Application Information

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IPC IPC(8): C12Q1/68C12M1/34G01N33/68
Inventor 王振新李小梅刘殿俊
Owner CHANGCHUN INST OF APPLIED CHEMISTRY - CHINESE ACAD OF SCI
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