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Construction of Goat Pox Virus Recombination System and Dual-expression pprv H/F Protein Vaccine Without Plaque Cloning and Screening Tags

A goat pox virus and screening tag technology, applied in the field of medicine, can solve the problems of increasing the immunogen and the length of the inserted sequence, and achieve the effects of improving the construction efficiency, increasing the expression burden and avoiding the purification work.

Active Publication Date: 2020-10-16
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

At the same time, for ease of purification, green fluorescent protein (GFP) was used as a tracer tag and gpt was used as a drug screening gene, which made the insertion sequence longer and added unnecessary immunogens, which would hinder the future clinical application of recombinant virus vaccines. potential adverse effects

Method used

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  • Construction of Goat Pox Virus Recombination System and Dual-expression pprv H/F Protein Vaccine Without Plaque Cloning and Screening Tags
  • Construction of Goat Pox Virus Recombination System and Dual-expression pprv H/F Protein Vaccine Without Plaque Cloning and Screening Tags
  • Construction of Goat Pox Virus Recombination System and Dual-expression pprv H/F Protein Vaccine Without Plaque Cloning and Screening Tags

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Embodiment 1

[0050] Embodiment 1 Double expression PPRV H and the construction of F protein recombinant goat pox virus vaccine

[0051] 1 Materials and methods

[0052] 1.1 Virus strains and cells

[0053] The PPRV attenuated vaccine strain (Nigeria / 75 / 1) and the GPV temperature-sensitive attenuated vaccine strain (CVCC AV41) were provided by the China Veterinary Drug Administration; the primary lamb testis cells (Lamb testis, LT) were subjected to routine The method was prepared by culturing in MEM containing 10% fetal bovine serum; Vero cells were cultured in DMEM containing 10% fetal bovine serum. Amplification and titration of GPV or recombinant GPV were performed on LT cells in MEM containing 2% fetal bovine serum. The amplification and titration of PPRV attenuated vaccine strain (Nigeria / 75 / 1) was performed on Vero cells using DMEM containing 2% fetal bovine serum.

[0054] 1.2 Main reagents and instruments

[0055]Various restriction enzymes, Phanta DNA polymerase, and homologou...

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Abstract

The invention discloses a goat pox virus recombination system free of bacteriophage plaque cloning and screening labels and dual-expression PPRV (peste des petits ruminants virus) H / F protein vaccine construction. The recombination system is not only capable of constructing single-expression and dual-expression recombinant viruses or recombinant vaccines and even capable of constructing recombinant vaccines or recombinant viruses which expressing more than three proteins, the recombinant viruses obtained by the recombination system are free of any screening labels, and accordingly potential adverse effects caused by the screening labels in vaccine use can be avoided. In addition, the invention discloses a peste des petits ruminants virus H and F protein dual-expressed screening-label-free recombinant goat pox virus prepared by the recombination system, wherein the recombinant goat pox virus. According to researches, the recombinant is capable of inducing cell and humoral immunity more comprehensive than single antigen expression and simultaneously preventing two important diseases including PPR and goat pox, and an induced high-enough neutralizing antibody is beneficial to immunoserology monitoring, so that the system has a promising application prospect in prevention and treatment of peste des petits ruminants and goat pox.

Description

technical field [0001] The present invention relates to a goat pox virus recombination system that does not require plaque cloning and screening tags, and also relates to a dual expression of Peste des petits ruminants virus (Peste des petits ruminants virus, PPRV) H and F proteins without screening tags constructed by the system The recombinant goat pox virus vaccine and its construction method and application, the invention belongs to the technical field of medicine. Background technique [0002] Peste des petits of ruminants (PPR) and goat pox (GP) are caused by Peste des petits of ruminants virus (Peste des petits of ruminants), PPRV) and goat pox virus (GPV) of the Orthopoxvirus family (Orthopoxvirus) goat pox virus (Capripoxvirus), two major diseases that harm small ruminants such as sheep and goats. Among them, PPR has been prevalent in Africa, the Middle East, Central Asia, South Asia and Southeast Asia for a long time. As a transnational animal disease with great h...

Claims

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Application Information

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IPC IPC(8): C12N15/85C12N7/01A61K39/275A61K39/295A61K39/155A61P31/20A61P31/14
CPCA61K39/12A61K2039/5256A61K2039/552A61K2039/70C12N7/00C12N15/85C12N2710/24221C12N2710/24234C12N2710/24252C12N2760/18422C12N2760/18434C12N2800/107
Inventor 步志高陈伟业胡森
Owner HARBIN VETERINARY RES INST CHINESE ACADEMY OF AGRI SCI
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