Looking for breakthrough ideas for innovation challenges? Try Patsnap Eureka!

Primers for determining full-gene sequence of influenza A H1N1 viruses and determination method

An influenza virus and whole gene technology, applied in the field of primers and kits for determining the whole gene sequence of influenza A (H1N1) virus, can solve the problems of poor primer specificity and sensitivity that affect the monitoring of the whole gene sequence of influenza A (H1N1) virus Low-level problems, to achieve the effect of stable results, simple sampling and high sensitivity

Active Publication Date: 2013-03-06
SUN YAT SEN UNIV
View PDF2 Cites 2 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, we also found that according to the 46 pairs of whole gene sequencing primers recommended by WHO (http: / / www.who.int / csr / disease / swineflu / en / index.html), more than one-third of the primers have poor specificity and sensitivity The low phenomenon seriously affects the progress of monitoring the whole gene sequence determination of influenza A (H1N1) virus

Method used

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
View more

Image

Smart Image Click on the blue labels to locate them in the text.
Viewing Examples
Smart Image
  • Primers for determining full-gene sequence of influenza A H1N1 viruses and determination method
  • Primers for determining full-gene sequence of influenza A H1N1 viruses and determination method
  • Primers for determining full-gene sequence of influenza A H1N1 viruses and determination method

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0030] Example 1: Design of primers and probes

[0031] By comparing and analyzing the gene sequences of 100 known A / H1N1 virus strains in all GenBank databases, a highly conserved segment was selected, and 48 pairs of primers and probes were designed, as described in SEQ ID NO: 1-96. The principles of primer design are as follows:

[0032] (1) The length of the primer is about 20-23nt, and the GC content is between 45%-55%.

[0033] (2) The length of PCR amplification is generally between 400-600 bp.

[0034] (3) For the same template fragment, design primers so that each site is covered by 2-3 PCR amplified fragments, so as to avoid point mutations in the PCR reaction from affecting the interpretation of the results.

[0035] Analyze the information of the above-mentioned virus-specific genes, and use the sequence analysis software DNASTAR to exclude inter-primer / inner dimers, and verify the specificity of the primers and the homology of similar pathogens by BLAST, and d...

Embodiment 2

[0036] Embodiment 2: Construction and optimization of PCR reaction system

[0037] 1. Sample Collection

[0038] Our laboratory collected samples from three suspected cases on May 17 and May 28, 2009, including the first suspected imported H1N1 case in Guangdong Province and the first and second second-generation suspected cases of H1N1 in my country Nasal / throat swab specimens of Dai and Xue.

[0039] 2. Extraction of RNA from nasal / throat swab specimens

[0040] Use the QIAmp MiniElute Virus Spin Kit (Qiagen, Chatsworth, CA) to extract RNA from nasal / pharyngeal swab specimens, and follow the instructions:

[0041] (1) When using this kit for the first time, add 100% ethanol to the AW1 and AW2 buffers according to the volume indicated on the reagent bottle, add 25 ml absolute ethanol to 19 ml AW1, add 30 ml absolute ethanol to 13 ml AW1 Ethanol; add 28 μg / ml carrier RNA to AL.

[0042] (2) Put 25 μl Qiagen Protease into a 1.5 ml centrifuge tube.

[0043] (3) Add 200 μl ...

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

PUM

No PUM Login to View More

Abstract

The invention discloses full-gene sequence primers for determining influenza A H1N1 viruses and a determination method. The invention relates to 48 pairs of primers with strong specificity, high sensitivity and good stability, which are used for determining 8 fragments of gene sequences of the influenza A H1N1 viruses, wherein the 8 fragments comprise HA, NA, NP, M1, M2, PA, PB1 and PB2. The method for detection by utilizing the 48 pairs of primers has the advantages of simple operation, short time consumption, strong specificity and high sensitivity and is suitable for identification of the influenza A H1N1 viruses, epidemiological survey and research and the like.

Description

technical field [0001] The invention relates to a detection technology of type A (H1N1) influenza virus, in particular to a primer and a kit for measuring the whole gene sequence of type A (H1N1) influenza virus. Background technique [0002] Influenza (flu) is a respiratory infection caused by the influenza virus (flu virus). Influenza viruses belong to the Orthomyxoviridae family and are negative single-stranded RNA viruses. Influenza viruses are divided into three types: A, B, and C according to the antigenicity of nucleoprotein (NP) and matrix protein (MP), all of which can infect humans. Among them, influenza A virus has the strongest ability to mutate and can cause influenza seasonal epidemics or influenza pandemics. Influenza B virus has a weak ability to mutate, and generally causes seasonal influenza epidemics. The antigenicity of influenza C virus is relatively stable, and it is less likely to cause epidemics. [0003] Influenza A virus spread from birds to hum...

Claims

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

Application Information

Patent Timeline
no application Login to View More
Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/70C12Q1/68C12N15/11
Inventor 黎孟枫曹开源吴珏珩朱勋何振健袁洁李隽张定梅徐霖杨纨李蔓英
Owner SUN YAT SEN UNIV
Who we serve
  • R&D Engineer
  • R&D Manager
  • IP Professional
Why Patsnap Eureka
  • Industry Leading Data Capabilities
  • Powerful AI technology
  • Patent DNA Extraction
Social media
Patsnap Eureka Blog
Learn More
PatSnap group products

Browse by: Latest US Patents, China's latest patents, Technical Efficacy Thesaurus, Application Domain, Technology Topic, Popular Technical Reports.

© 2024 PatSnap. All rights reserved.Legal|Privacy policy|Modern Slavery Act Transparency Statement|Sitemap|About US| Contact US: help@patsnap.com