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HCV genotyping DNA microarray chip

A microarray chip, genotyping technology, applied in DNA/RNA fragmentation, recombinant DNA technology, microorganism determination/inspection, etc., can solve the problems of cumbersome operation, low throughput, low sensitivity, etc.

Inactive Publication Date: 2010-03-03
DAAN GENE CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] Currently, HCV genotyping methods mainly include: restriction length polymorphism (RFLP), type-specific primer PCR, type-specific probe nucleic acid hybridization analysis, RDB, direct sequencing and other methods, but most of these methods are cumbersome to operate, Long detection time, low throughput, low sensitivity, low specificity, etc.

Method used

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  • HCV genotyping DNA microarray chip
  • HCV genotyping DNA microarray chip
  • HCV genotyping DNA microarray chip

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0027] Example 1: Design of PCR primers and probes for HCV genotyping DNA microarray chip

[0028] Design RT and PCR primers in the 5'UTR region of the HCV genome, and use an asymmetric RT-PCR system to amplify the target fragments required for typing; within the range of target fragments, design 3 type 1 probes and type 2 probes 4, 3 type 3 probes, 3 type 6 probes and 1 universal probe.

Embodiment 2

[0029] Example 2: Preparation of HCV genotyping DNA microarray chip

[0030] Using a gene chip automatic spotting instrument, 14 typing probes and 2 control probes are spotted on a specific area of ​​the glass slide to make a DNA microarray.

[0031] (1) Spot probe solution: Dilute the probe into 5×SSC, 0.05% SDS solution, the final concentration is 30μM;

[0032] (2) Spot matrix: The probes are dotted on the glass slide, and each probe is horizontally repeated 2 dots in the hybridization area, 8 dots per row, 4 rows, 32 dots (see attached figure 1 ); hybridization is divided into 10 areas, 8 copies can be detected at the same time

[0033] Sample (9, 10 hybridization area is the control area).

Embodiment 3

[0034] Example 3: Using the product of the present invention to test 8 clinical samples

[0035] Take 8 clinical samples and one negative and positive control, extract RNA, and perform PCR amplification according to the operating instructions of RT-PCR reagents to obtain DNA hybridization template.

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Abstract

The invention relates to a DNA microarray chip, in particular to a hepatitis C virus (HCV) genotyping DNA microarray chip. A solid-phase carrier substrate is used, and an oligonucleotide probe is designed in view of different genotypes of HCV so as to prepare the DNA microarray chip; and when matched with a PCR primer and other components, the DNA microarray chip can quickly and accurately classify the types of the hepatitis C virus in a blood sample.

Description

Technical field [0001] The invention relates to a DNA microarray chip, in particular to a hepatitis C virus (HCV) genotyping DNA microarray chip. The invention adopts a solid-phase carrier substrate, designs oligonucleotide probes according to different genotypes of HCV, and prepares a DNA microarray chip for rapid and accurate typing detection of hepatitis C virus in blood samples. Background technique [0002] Hepatitis C is an infectious disease that seriously harms human health and has a wide range of epidemics. Its pathogen is hepatitis C virus (HCV). At present, there are more than 170 million people infected with HCV in the world, and more than 100,000 HCV patients develop liver cancer each year, and then have gastrointestinal bleeding and ascites. According to the WHO 2002 annual report, in 2001, chronic liver disease caused 14 million deaths, including 796,000 cases caused by cirrhosis and 616,000 cases caused by primary liver cancer. [0003] HCV is a positive single-st...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/70C12Q1/68C12N15/11
Inventor 彭春梅张帆胡守旺李明何蕴韶
Owner DAAN GENE CO LTD
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