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Recombinant turkey herpesvirus as well as preparation method and application thereof

A technology for turkey herpes virus and chicken Newcastle disease virus, which is applied in the field of genetic engineering vaccines, can solve the problems of time-consuming recombinant vaccines and low efficiency of recombinant viruses, and achieves the effect of simple knock-in method and efficient insertion.

Pending Publication Date: 2022-03-04
YANGZHOU UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, these recombinant HVT vaccines are mainly produced by conventional homologous recombination in virus-infected cells, or by recombination in bacterial artificial chromosomes (BACs) from which full-length genomes are cloned. Methods to generate recombinant vaccines are time consuming

Method used

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  • Recombinant turkey herpesvirus as well as preparation method and application thereof
  • Recombinant turkey herpesvirus as well as preparation method and application thereof
  • Recombinant turkey herpesvirus as well as preparation method and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment

[0073] 1. Experimental materials

[0074] 1.1 Virus strains, plasmids and antibodies

[0075] HVT FC-126 vaccine strain, NDV gene type VII JS strain and AIV H9N2 (2019) strain were identified and preserved by the Key Laboratory of Poultry Preventive Medicine, Ministry of Education, Yangzhou University.

[0076] Plasmids: pX330, pX459, pcDNA3.1 and pcDNA3.1-Cre plasmids were purchased from the website of Hunan Keai Medical Instrument Co., Ltd. (Youbao Biology). -T Easy Vector Systems were purchased from Promega Corporation. Plasmid pCMV-N-GFP was purchased from Biyuntian Company. Plasmid pX459-sgA was constructed and preserved in the Key Laboratory of Poultry Preventive Medicine, Ministry of Education, Yangzhou University. https: / / doi.org / 10.1093 / nar / gkw064) into the pX459 plasmid through the BbsI restriction site.

[0077] Antibody: anti-HA (H9) monoclonal antibody 2G4 (reference "Analysis of amino acid variation of key epitope epitope of H9N2 influenza virus envelope gly...

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Abstract

The invention provides a recombinant turkey herpesvirus as well as a preparation method and application thereof, and particularly provides a recombinant turkey herpesvirus, an exogenous gene is inserted into a spacer region between an HVT005 region and an HVT006 region of a turkey herpesvirus genome, and the exogenous gene is selected from genes from a Newcastle disease virus, an avian influenza virus or an avian infectious bursal disease virus; the spacer region between the HVT005 region and the HVT006 region of the turkey herpesvirus genome is between 8867nt and 9319nt of the turkey herpesvirus genome, and the nucleotide sequence of the spacer region is SEQ ID NO: 1. The invention discovers that the HVT005-HVT006 spacer region can be used as a site for inserting an HVT exogenous gene for the first time. An exogenous gene is inserted into a spacer region HVT005-HVT006 of an HVT genome by utilizing a CRISPR / Cas9 technology, and the vaccine strain can be used as a vaccine candidate strain for preventing the Newcastle disease and the avian influenza.

Description

technical field [0001] The invention belongs to the technical field of genetic engineering vaccines, and in particular relates to constructing a recombinant turkey herpes virus strain using CRISPR-Cas9 technology. The recombinant virus can provide good protection as a vaccine. Background technique [0002] As a recently emerging gene editing technology, the CRISPR-Cas9 system has achieved great success in efficiently producing genetically engineered cells and animal models, and has also been used to edit the genomes of a variety of large DNA viruses, including herpes simplex virus, adenovirus , pseudorabies virus, poxvirus, guinea pig cytomegalovirus and duck enteritis virus. [0003] Turkey herpesvirus (HVT) has been widely used as a vaccine vector expressing heterologous antigens for various poultry diseases, and these vaccines provided good and long-lasting immunity against Marek's disease virus (MDV) and the virus to which the gene inserted in the vector belongs. Howeve...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N7/01C12N15/45C12N15/44C12N15/40C12N15/869A61K39/12A61K39/145A61K39/17A61P31/14A61P31/16
CPCC12N7/00C07K14/005C12N15/86A61K39/12A61P31/16A61P31/14C12N2710/16021C12N2710/16043C12N2710/16052C12N2760/18122C12N2760/16122C12N2720/00022C12N2760/18134C12N2760/16134C12N2720/00034A61K2039/5256A61K2039/53A61K2039/552C12N9/22C12N2310/20C12N2710/16343C12N2710/16351C12N2720/10022C12N2720/10034C12N2710/16034A61K2039/54A61K2039/545A61K39/145A61K39/17C12N5/0603C12N15/11C12N2710/16022C12N2720/10021C12N2720/10071C12N2760/16121C12N2760/16171C12N2760/18121C12N2760/18133C12N2760/18171C12N2800/107C12N2800/80
Inventor 秦爱建宰栩生石彬钱琨邵红霞叶建强
Owner YANGZHOU UNIV
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