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Gene chip for inspecting and grouping SARS coronavirus, its method and reagent kit

A coronavirus and gene chip technology, applied in the field of detection, can solve problems such as inability to type and predict epidemics, and achieve high accuracy, strong repeatability, and easy operation

Inactive Publication Date: 2007-06-20
TIANJIN BIOCHIP TECH CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0007] The technical problem to be solved in the present invention is to disclose a gene chip for detection and typing of SARS coronavirus, so as to complement existing detection technology that can detect SARS coronavirus but cannot carry out typing and epidemic prediction on it.

Method used

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  • Gene chip for inspecting and grouping SARS coronavirus, its method and reagent kit
  • Gene chip for inspecting and grouping SARS coronavirus, its method and reagent kit
  • Gene chip for inspecting and grouping SARS coronavirus, its method and reagent kit

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0032] Example one Preparation of oligonucleotide probes

[0033] 1. Sequence alignment: Use the multiple sequence alignment software clustalx to compare the sequences of the s genes of 38 different SARS coronaviruses, and obtain 27 mutation sites in the s genes. According to the number of mutation sites, they can be divided into 4 types: prototype (no mutation), low pathogenic type (mutation at 2-7 sites), high pathogenic type (occurring at 17-25 sites) Mutation) and epidemic type (27 sites have mutations).

[0034]2. Design probe: Use Primer5 biology software to design 106 oligonuclei for typing based on 25 mutation sites (the other two sites did not cause amino acid changes after mutations, so they are not tested) In addition, two probes were designed to detect SARS coronavirus in the conserved region of the s gene. The 108 oligonucleotide probes mentioned above are shown in Table 1.

[0035] Table 1. Oligonucleotide probes for SARS coronavirus typing and detection

[0036] ...

Embodiment 2

[0041] Example two Preparation of primers for amplification of s gene

[0042] 1. Primer design: Use Primer 5.0 and other bioinformatics software to analyze the genome of SARS coronavirus, and design 4 pairs of primers to amplify the s gene in the conserved region of the s gene. The primer sequence is shown in Table 2.

[0043] Table 2. Primers used to amplify the s gene

[0044] SEQ ID

[0045] 2. Primer synthesis: send the selected primer sequence to the primer synthesis company for entrusted synthesis (synthesized by Aoke Company) for use.

Embodiment 3

[0046] Example three Chip preparation

[0047] 1. Preparation of probe solution: Dissolve the probe prepared in Example 2 in a 50% DMOS solution so that the final concentration of the probe reaches 1 μg / μl.

[0048] 2. Chip spotting: Take 57.5mm×25.5mm×1mm (length×width×height) aldehyde-based glass slide as the solid phase carrier, on which there are three spotting areas I, II, and III (as shown in the figure) 1). Use the automatic biochip spotting instrument (Spotarray 72) to spot samples in the three spotting areas with the 9 (row) × 15 (column) probe arrangements shown in Figures 2a, 2b, and 2c to make a gene chip. As shown in Figures 2a, 2b and 2c, Cy represents oligonucleotides labeled with fluorescein Cy3. These points are used to determine the start and end positions of the array during the scanning process; P1-P2 represent the positive control used to detect SARS coronavirus Probe; neg stands for negative control probe; S001-S106 stands for specific probes used to detect...

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Abstract

Gene chip for inspecting and parting SARS coronavirus, its method and reagent kit are disclosed. The gene chip consists of solid-phase carrier and oligonucleotide probe fixed on the solid-phase carrier, which comprises DNA fragment containing S gene mutation site selected from S gene sequence with one or multiple SARS coronavirus and DNA fragment selected from S gene conservative sequence of SARS coronavirus. The process is carried out by PCR amplifying for sample cDNA to be inspected, hybridizing with the gene chip, determining nucleotide kind by different fluorescent value and parting for SARS sample to be inspected. It's simple, accurate and repeated.

Description

Technical field [0001] The present invention belongs to the field of detection technology, and relates to a gene chip (or DNA microarray) and its use method and a kit, in particular to a gene chip for SARS coronavirus detection and typing and its detection typing Method and test typing kit. Background technique [0002] The severe acute respiratory syndrome (SARS) caused by atypical pneumonia is a serious disease that can lead to death of patients. This disease was first discovered in China’s Guangdong Province in November 2002. As of September 2003, 8,436 people worldwide had been infected with this disease, of which 812 had died, with a mortality rate of nearly 10%. On April 16, 2003, the World Health Organization (WHO) announced that the source of SARS was a member of a family of coronaviruses that had never been found in humans before. At present, the sequence of the SARS virus has been deciphered, proving that it is a brand new coronavirus. Although it can cause serious dise...

Claims

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Application Information

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IPC IPC(8): C12Q1/68
CPCY02A50/30
Inventor 王磊徐建国郭玺
Owner TIANJIN BIOCHIP TECH CO LTD
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