Recombinant novel coronavirus as well as preparation method and application thereof

A technology of recombinant virus, influenza virus, applied in biological field

Active Publication Date: 2020-08-21
THE FIFTH MEDICAL CENT OF CHINESE PLA GENERAL HOSPITAL +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

So far, it has not been reported that influenza virus is used as a vector to express key proteins of SARS-CoV-2 virus for the development of recombinant SARS-CoV-2 virus strains

Method used

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  • Recombinant novel coronavirus as well as preparation method and application thereof
  • Recombinant novel coronavirus as well as preparation method and application thereof
  • Recombinant novel coronavirus as well as preparation method and application thereof

Examples

Experimental program
Comparison scheme
Effect test

experiment example 1

[0068] Experimental example 1 Preparation of recombinant novel coronavirus (SARS-CoV-2) vaccine of chimeric S-RBD+E+M-NS epitope

[0069] This embodiment provides recombinant novel coronavirus (SARS-CoV-2), and this recombinant SARS-CoV-2 expresses the protein named S-RBD+E+M, and described S-RBD+E+M is SARS-CoV -2 S protein receptor binding domain (receptorbinding domain, RBD) and E protein and M protein connected, its amino acid sequence is shown in SEQ ID NO.3, by S-RBD+E+ shown in SEQ ID NO.4 The M gene codes.

[0070] 1. Construction of recombinant plasmids:

[0071] 1. Using the NS gene fragment of the cold-adapted and attenuated influenza virus strain A / AnnArbor / 6 / 60 as the target for inserting the S-RBD+E+M gene, construct a recombinant plasmid and name it pAD-S-RBD+E+ M-NS.

[0072] Synthesize the gene shown in SEQ ID NO.5 (structure such as figure 1 shown), wherein, the 1st to 11th positions from the 5' end of SEQ ID No.5 are protective bases + enzyme cutting sit...

Embodiment 2

[0122] Example 2 Preparation of recombinant novel coronavirus (SARS-CoV-2) vaccine of chimeric S-RBD+E+M-PR8-NS

[0123]This embodiment provides recombinant novel coronavirus (SARS-CoV-2), and this recombinant SARS-CoV-2 expresses the protein named S-RBD+E+M, and described S-RBD+E+M is SARS-CoV -2 S protein receptor binding domain (receptorbinding domain, RBD) and E protein and M protein connected, its amino acid sequence is shown in SEQ ID NO.3, by S-RBD+E+ shown in SEQ ID NO.4 The M gene codes.

[0124] 1. Construction of recombinant plasmids:

[0125] 1. Using the NS gene fragment (named PR8-NS) of influenza virus strain A / PR / 8 / 1934 as the target point for inserting the epitope gene, construct a recombinant plasmid named pHW-S-RBD+E+M- PR8-NS.

[0126] Synthesize the gene shown in SEQ ID NO.8 (structure as shown in figure 2 Shown), wherein, the 1st to 11th positions from the 5' end of SEQ ID No.8 are protective bases + enzyme cutting sites, and the 12th-41st positions ...

experiment example 3

[0152] Experimental example 3 Preparation of recombinant SARS-CoV-2 vaccine of chimeric S-RBD-NA epitope

[0153] This embodiment provides a recombinant new coronavirus (SARS-CoV-2), the recombinant S-RBD-NA expresses a protein named S-RBD, and the S-RBD is the receptor binding of the SARS-CoV-2 S protein Region (RBD), the amino acid sequence of which is SEQ ID NO.1, encoded by the S-RBD gene shown in SEQ ID NO.2.

[0154] 1. Construction of recombinant plasmids:

[0155] 1. Using the NA gene fragment of influenza virus H1N1 subtype (influenza virus strain A / Michigan / 45 / 2015) as the target for inserting the S-RBD gene, a recombinant plasmid was constructed and named pAD-S-RBD-NA.

[0156] Synthesize the gene shown in SEQ ID NO.6 (structure as shown in image 3 shown), wherein, the 1st to 11th positions of SEQ ID No.6 from the 5' end are protective bases + enzyme cutting sites, and the 12th to 34th positions of SEQ ID NO.6 are non-coding at the 5' end Region (23nt), the first ...

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Abstract

The invention discloses a recombinant novel coronavirus as well as a preparation method and application thereof. The invention firstly discloses a recombinant virus. Any one of an NS gene of an influenza virus, an HA gene of the influenza virus and an NA gene of the influenza virus of the recombinant virus is replaced. The invention further discloses application of the recombinant virus in preparation of a product for preventing and/or treating diseases caused by influenza virus and/or SARS-CoV-2. According to the invention, the SARS-CoV-2 antigen epitope and the influenza virus genome are operated from the gene level; the recombinant SARS-CoV-2 vaccine strain taking the influenza virus as the carrier is prepared on the basis of an RG technology, so that the problem of SARS-CoV-2 vaccine shortage is solved. The recombinant novel coronavirus will be a new milestone in the field of coronavirus vaccines, and the SARS-CoV-2 chimeric vaccine can protect more people from being harmed by theinfluenza virus and SARS-CoV-2.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to a recombinant novel coronavirus and its preparation method and application. Background technique [0002] Coronavirus is a type of RNA virus with an envelope and a linear single-stranded positive strand. The particles are round or oval, with a diameter of about 60-140nm. There are three kinds of glycoproteins on the membrane surface: spike glycoprotein (SpikeProtein, S): a ball-and-stick-shaped protrusion on the surface of the virus particle, which belongs to type I transmembrane protein. It can be decomposed by S enzyme into two functional units, S1 and S2. S1 is used to promote the binding of the virus to the host cell receptor, and the host receptor binding region (RBD region) to bind to the receptor. This region can bind to the ACE2 protein. If the binding is completed, the new coronavirus will infect human respiratory epithelial cells , leading to pneumonia; membrane glycoprot...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N7/01C12N15/50C07K14/165A61K39/295A61K39/215A61K39/145A61P31/14A61P31/16C12R1/93
CPCC07K14/005A61K39/12A61P31/14A61P31/16C12N2760/16122C12N2760/16134C12N2770/20034A61K2039/70A61K2039/53
Inventor 杨鹏辉王希良程晋霞王欢孙芳
Owner THE FIFTH MEDICAL CENT OF CHINESE PLA GENERAL HOSPITAL
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