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Process of detecting gene mutation in cell with one-dimensional microflow controlled biochip

A biochip and microfluidic technology, which is applied in the field of biological application of the micro-total analysis system, can solve the problems such as the difficult coexistence of micro-quantity detection, high sensitivity and high-throughput analysis

Inactive Publication Date: 2007-10-03
HUNAN UNIV
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0003] The purpose of the present invention is to provide a novel detection method applicable to the identification and analysis of cell gene mutations, so as to solve the defects that the characteristics of micro detection, high sensitivity and high-throughput analysis are difficult to coexist in the current gene mutation analysis methods, and Promote further development in this technical field

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  • Process of detecting gene mutation in cell with one-dimensional microflow controlled biochip
  • Process of detecting gene mutation in cell with one-dimensional microflow controlled biochip
  • Process of detecting gene mutation in cell with one-dimensional microflow controlled biochip

Examples

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Embodiment 1

[0016] Example 1, one-dimensional microfluidic mutation detection chip detects p53 gene mutation with DNA sequence as object:

[0017] 1. Preparation of silica microbeads for mutation detection: Take about 10-20 mg of silica microbeads with a diameter of 40 μm and place them in a 1.5ml Ep tube, add 500 μl of 0.01M NaOH to activate the surface for 20 minutes, wash with TE 3 times, remove Supernatant: add 80μl 0.1~1.0mg / ml biotinylated bovine serum albumin (Biotin-BSA), place on a low-temperature shaker at 4°C, shake at 350rpm for 12~48h, then wash 3 times with TE, Biotin- BSA is bound to the particle surface through physical adsorption; add 80 μl of 0.1-1.0 mg / ml streptavidin (Streptavidin), place it on a low-temperature shaker at 4°C, shake at 350 rpm for 4-8 hours, and the streptavidin can bind to the particle The biotin on the surface is strongly bound and fixed on the particle surface, washed with TE for 3 times; add 50 μl of 0.1-1.0 μM biotin-modified mutation detection ca...

Embodiment 2

[0021] Example 2, one-dimensional microfluidic mutation detection chip detects p53 gene mutation with mRNA sequence as object:

[0022]1. Preparation of silica microbeads for mutation detection: Take about 10-20 mg of silica microbeads with a diameter of 40 μm and place them in a 1.5ml Ep tube, add 500 μl of 0.01M NaOH to activate the surface for 20 minutes, wash with TE 3 times, remove Supernatant: add 80μl 0.1~1.0mg / ml biotinylated bovine serum albumin (Biotin-BSA), place on a low-temperature shaker at 4°C, shake at 350rpm for 12~48h, then wash 3 times with TE, Biotin- BSA is bound to the particle surface through physical adsorption; add 80 μl of 0.1-1.0 mg / ml streptavidin (Streptavidin), place it on a low-temperature shaker at 4°C, shake at 350 rpm for 4-8 hours, and the streptavidin can bind to the particle The biotin on the surface is strongly bound and fixed on the surface of the particles, washed with TE three times; 50 μl of 0.1-1.0 μM biotin-modified mutation detectio...

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Abstract

The process of detecting gene mutation in cell with one-dimensional microflow controlled biochip includes constructing mutation detecting chip based on one-dimensional microflow controlled micro bead array, diluting hybridization buffering solution with total RNA or target DNA sample and fluorescent probe, pressurizing the hybridization buffering solution through the micro channel to the micro bead array, hybridizing and eluting high precision eluent containing formamide, observing in a turned fluorescent microscope and taking micro bead surface fluorescence image, calculating the surface fluorescence intensity, and judging the mutation type of the detected site based on the fluorescence intensity. The present invention is hopeful in realizing the gene mutation analysis in single cell level.

Description

technical field [0001] The invention relates to the biological application field of a micro-total analysis system, in particular to the application and development of a one-dimensional microfluidic biochip in cell gene mutation analysis. Background technique [0002] Seven countries including the United States, Britain, and China announced in February 2001 that they had completed the first draft of the analysis of the human genome sequence, which transformed the field of human biomedical research from the genome era to the post-genome era, that is, the functional genome era. Geneticists generally believe that an important direction of efforts in the post-genome era is to analyze the variation of human gene sequences, because the study of mutations in human DNA sequences will help to better understand the occurrence, development, and response to drug treatments of human diseases. Therefore, the analysis and detection method of the mutation is very important. Before 1985, mut...

Claims

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Application Information

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IPC IPC(8): G01N21/64G01N33/48C12Q1/68
Inventor 王柯敏张何羊小海文建辉谭蔚泓
Owner HUNAN UNIV
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