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Self-assembled bovine parainfluenza virus type 3 nanoparticle-like antigen and application thereof

A bovine parainfluenza virus and nanoparticle technology, applied in the direction of virus antigen components, viruses, virus peptides, etc., can solve the problems of strong virulence, insufficient immune response, and dispersal of viruses

Active Publication Date: 2020-09-01
NORTHEAST AGRICULTURAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Although there have been research reports on inactivated vaccines and attenuated vaccines, there are safety risks such as loose virus and strong virulence; there are also various forms of new vaccines, and the induced immune response is often not comprehensive enough, resulting in the body's inability to completely resist BPIV3 virus insufficient infection

Method used

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  • Self-assembled bovine parainfluenza virus type 3 nanoparticle-like antigen and application thereof
  • Self-assembled bovine parainfluenza virus type 3 nanoparticle-like antigen and application thereof
  • Self-assembled bovine parainfluenza virus type 3 nanoparticle-like antigen and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0062] Example 1 Preparation of self-assembled BPIV3 nanoparticle-like antigen

[0063] method:

[0064] 1. Cloning of bovine BPIV3 HN extramembrane region (HNex) gene

[0065] (1) BPIV3 (5555-9258) gene fragment cloning

[0066]①Primer design, according to the homology comparison of the BPIV3 genome sequence in Genbank, with the gene sequence of BPIV3 NM09 strain as a reference, use the biological software Primer 5 to design the cloning primers at the conserved regions at both ends of the HN gene, namely BPIV3-5555 (5'-TCTGTAGGTAATCTAATTGTTGC -3') and BPIV3-9158 (5'-GGTTATACCATTTGTCTGATTGA-3'), synthesized by Xinhai Gene Biotechnology Co., Ltd.

[0067] ② BPIV3 genomic RNA extraction and reverse transcription, according to Reagent instruction manual Extract BPIV3 genomic RNA and perform reverse transcription, BPIV3-9158 is the reverse transcription primer, and the cDNA of the reverse transcription product is named "BPIV3-cDNA9158".

[0068] ③PCR amplification of BPIV3 (5...

Embodiment 2

[0142] Example 2 The immune effect experiment of BPIV3 HNex-RFNp

[0143] method:

[0144] 1. Animal grouping and immunization

[0145] C57 BL / 6 mice, 6-8 weeks old, were randomly divided into 5 groups. They were HNex-RFNp (100μg / head), HNex-RFNp (50μg / head), HNex (50μg / head), inactivated BPIV3 (50μg / head) and PBS (200μL) groups respectively. At 0w and 1w, each mouse was inoculated with 200 μL of the corresponding antigen. Before immunization and after immunization (6w), blood was collected by tail docking every week, serum was separated and stored at -70°C.

[0146] 2. Humoral immunity test

[0147] (1) ELISA antibody against BPIV3 and HNex

[0148] BPIV3 and HNex were used as antigens, and ELISA plates were coated at 50 ng / well. The I-ELISA method was used to detect the contents of BPIV3 ELISA antibodies and HNex ELISA antibodies in the collected serum, to clarify the rules of antibody growth and decline, and to compare and analyze the antibody levels among the groups. ...

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Abstract

The invention discloses a self-assembled bovine parainfluenza virus type 3 nanoparticle-like antigen and application thereof. The self-assembled BPIV3 nanoparticle-like antigen is a self-assembled BPIV3 nanoparticle-like antigen (named as HNex-RFNp) which is obtained by expression of a Bac-to-Bac baculovirus expression system and taking ferritin protein of Helicobacter pylori as a carrier to carryBPIV3 HN protein. The HN protein is fused at the N-terminal of ferritin, and the HN protein is a fragment located at the amino terminal 128-572aa of the HN protein of the BPIV3-vaccine strain. According to the invention, the immune effect of HNex-RFNp is evaluated from three aspects of humoral immunity, cellular immunity and animal protective tests, and results show that the immune effect of HNex-RFNp is significantly better than that of inactivated BPIV3 protein and HNex protein. The invention provides a theoretical basis for developing a novel, safe and efficient vaccine capable of being used for preventing BPIV3 infection, and provides a technical platform.

Description

technical field [0001] The present invention relates to a self-assembled BPIV3 particle-like antigen and its preparation method and application, in particular to a self-assembled BPIV3 nanoparticle-like antigen carrying BPIV3 HNex protein with ferritin of Helicobacter pylori as a carrier and its preparation method and application. The invention belongs to the field of biotechnology. Background technique [0002] Bovine parainfluenza (Bovine parainfluenza) is an acute, contagious disease caused by Bovine parainfluenza virus type 3 (BPIV3). The incidence of the disease often increases significantly under stress conditions such as transportation, climate change, physical decline, and malnutrition. It manifests as severe dyspnea, loss of appetite, and depression, accompanied by clinical symptoms such as runny nose, tearing, and coughing. In production, the disease mostly occurs in cattle that have been transported long distances, so it is also called "transportation fever". BP...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K39/155A61P31/16C07K19/00
CPCA61K39/12C07K14/005C07K14/195A61P31/16C12N2760/18622C12N2760/18634A61K2039/552A61K2039/575A61K2039/572A61K2039/57A61K2039/5258
Inventor 王君伟曲哲会高明春曹翀李明珠安冉于越洋李臣锋
Owner NORTHEAST AGRICULTURAL UNIVERSITY