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Primer group and method for rapidly identifying influenza A virus subtypes through combination of DNA barcodes and second-generation high-throughput sequencing

An influenza virus, high-throughput technology, applied in biochemical equipment and methods, DNA/RNA fragments, recombinant DNA technology, etc., can solve the problems that the development of barcode technology has not yet been reported, and achieve the goal of rapid influenza typing diagnostic procedures Effect

Inactive Publication Date: 2016-01-20
INSPECTION & QUARANTINE TECH CENT SHANDONG ENTRY EXIT INSPECTION & QUARANTINE BUREAU
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, there is no corresponding report on the development of barcode technology for the virus subtype itself

Method used

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  • Primer group and method for rapidly identifying influenza A virus subtypes through combination of DNA barcodes and second-generation high-throughput sequencing
  • Primer group and method for rapidly identifying influenza A virus subtypes through combination of DNA barcodes and second-generation high-throughput sequencing
  • Primer group and method for rapidly identifying influenza A virus subtypes through combination of DNA barcodes and second-generation high-throughput sequencing

Examples

Experimental program
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Effect test

Embodiment Construction

[0049] This example provides a method for rapidly identifying influenza A virus subtypes by using DNA barcodes combined with next-generation high-throughput sequencing.

[0050] 1 Materials and methods

[0051] 1.1 Strain materials (09pdm swine flu H1N1, equine flu H3N8, bird flu H2N9, H4N1, H5N1, H7N2, H9N2, H13N6, H16N3) inactivated samples are kept by our laboratory, H6N5, H8N4, H10N8, H11N6, H12N5, H14N5, The H15N8 nucleic acid sample was donated by the Jilin Entry-Exit Quarantine Bureau (see the table below for details)

[0052] No.

Isolate

Acc. no.

1

A / swine / Shandong / 01 / 2009(H1N1)

KM368313

2

A / duck / Nanchang / 4-190 / 2000(H2N9)

CY117211

3

A / equine / Haerbin / 01 / 2010(H3N8)

KM892526

4

A / duck / Hong Kong / 951 / 1980(H4N1)

AB292404

5

A / chicken / Shandong / K01 / 2004(H5N1)

DQ767725

6

A / duck / Guangxi / 1455 / 2004(H6N5)

HM144533

7

A / duck / Hong Kong / 293 / 78(H7N2)

CY006029

8 ...

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Abstract

The invention discloses a primer group and a method for rapidly identifying influenza A virus subtypes through combination of DNA barcodes and second-generation high-throughput sequencing. The method comprises steps as follows: (1), genome extraction and inverse transcription; (2), primer design and PCR (polymerase chain reaction) amplification; (3), Ion Torrent sequencing; (4), bioinformatic analysis; and (5), phylogenetic analysis and genetic distance calculation. According to the primer group and the method, sequences of influenza A virus materials of 16 kinds of different areas and subtypes are analyzed, and the DNA barcodes which can be used for distinguishing the subtypes are found out; with the adoption of the technical idea of the DNA barcodes and a traditional RT-PCR (reverse transcription-polymerase chain reaction) detection method, whether the influenza virus exists can be rapidly judged with preference of a touch-down program, the subtype and a pathotype of the influenza A virus can be measured simultaneously, and one simpler and more rapid influenza typical diagnosis program is provided.

Description

Technical field: [0001] The invention relates to a primer set and a method for a method for quickly identifying type A influenza virus subtypes by combining DNA barcodes with second-generation high-throughput sequencing, and belongs to the field of biological genetic engineering. Background technique: [0002] According to the antigenic differences of influenza virus surface glycoproteins hemagglutinin (HA) and neuraminidase (NA), influenza A viruses can be divided into 18 HA subtypes and 11 NA subtypes. However, due to the lack of proofreading function of RNA polymerase, which relies on RNA, the virus has a high mutation rate in the genome during replication. Statistics show that three new subtypes of avian influenza viruses including H10N8, H5N2, and H7N9 have been newly discovered in my country. The number of annual outbreaks and seasonal differences also indicate that influenza cross-transmission among different hosts is becoming more frequent . However, traditional ide...

Claims

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Application Information

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IPC IPC(8): C12N15/11C12Q1/70C12Q1/68C12R1/93
Inventor 孙涛尹伟力房保海岳志芹梁成珠
Owner INSPECTION & QUARANTINE TECH CENT SHANDONG ENTRY EXIT INSPECTION & QUARANTINE BUREAU
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