Artificial recombinant H7N9 influenza virus and preparation method and application thereof

A technology of influenza virus and avian influenza virus, which is applied in the field of reverse genetics technology and animal infectious diseases, can solve the problems of isolating influenza virus strains, difficulty, etc., and achieve the goals of reducing production costs, high biological safety, and improving vaccine quality Effect

Active Publication Date: 2022-01-11
HARBIN VETERINARY RES INST CHINESE ACADEMY OF AGRI SCI
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, it is difficult to isolate influenza strain...

Method used

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  • Artificial recombinant H7N9 influenza virus and preparation method and application thereof
  • Artificial recombinant H7N9 influenza virus and preparation method and application thereof
  • Artificial recombinant H7N9 influenza virus and preparation method and application thereof

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Embodiment 1

[0039] Example 1: Preparation and Identification of Recombinant H7N9 Influenza Virus H7-Re4

[0040] 1. Construction and identification of 2 surface genes (HA and NA) of the recombinant virus:

[0041] Using the viral RNA extraction kit, extract the RNA of the H7N9 subtype highly pathogenic avian influenza virus isolate CK / YN / SD024 / 21 (H7N9) in China as a template, and use HA1 fragment-specific primers (HA1-U: atg aac act caa atc ctgata ttc g; HA1-L-: gca cca aat agg cct cgt ccc ttt gga acc tca gga aca ttc tt, where HA1-L is a primer that simultaneously mutates the HA cleavage site as a molecular feature of low pathogenicity) and HA2 fragment Specific primers (HA2-U: gga cga ggc cta ttt ggt gct ata gcg g; HA2-L: tta tat aca aat agt gcaccg cat g) were used to amplify two fragments of HA, HA1 and HA2, respectively, by RT-PCR ; Then use HA1 and HA2 as templates, HA1-U / HA2-L as primers, use phusion high-fidelity polymerase to perform overlap extension reaction (SOE-PCR), and ampl...

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Abstract

The invention discloses an artificial recombinant H7N9 influenza virus and a preparation method and application thereof. The artificial recombinant H7N9 influenza virus has a highly pathogenic avian influenza virus A/chicken/Yunnan/SD024/2021(H7N9) surface antigen Hemagglutinin (HA) gene and a Neuraminidase (NA) gene, but the amino acid sequence of the HA cleavage site is -PKGR-, the virus has six internal genes of PB2, PB1, PA, NP, M and NS of influenza virus A/PR/8/34(H1N1), the strain number is H7-Re4, and the preservation number is CCTCC NO: V202161. The strain has good chick embryo adaptability, and the virus growth titer can be increased by 9 times or above compared with that of a wild virus; the strain is used as a vaccine strain to produce vaccines, the production cost of the vaccines can be greatly reduced, and the quality of the vaccines is improved.

Description

technical field [0001] The invention relates to the field of reverse genetics technology and animal infectious disease technology, in particular to an artificially recombined and attenuated H7N9 subtype recombinant influenza virus and its preparation method and application. Background technique [0002] Avian influenza virus (AIV) is a segmented negative-strand RNA virus that can widely infect birds and mammals. According to the surface proteins of the virus, AIV is divided into 16 hemagglutinin (HA) and 9 neuraminidase (NA) subtypes. Among them, H5 and H7 subtypes of AIV can cause epidemics in poultry or wild birds, resulting in significant economic losses, and can also infect humans, causing public health hazards. China has developed a series of whole-virus inactivated vaccines, which have successfully controlled the H5 subtype since 2004 and the H7N9 subtype avian influenza since 2017. [0003] In response to poultry epidemics and public health hazards that may be cause...

Claims

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Application Information

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IPC IPC(8): C12N7/01C12N15/44C12N15/85A61K39/145A61P31/16C12R1/93
CPCC12N7/00C07K14/005C12N15/85A61K39/12A61P31/16C12N2760/16121C12N2760/16122C12N2760/16151C12N2760/16134C12N2800/107
Inventor 陈化兰曾显营施建忠邓国华田国彬李呈军
Owner HARBIN VETERINARY RES INST CHINESE ACADEMY OF AGRI SCI
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