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Antigen epitope I of neutrality B cell of chicken IBDV VP2 protein

An antigenic epitope, B cell technology, applied in the direction of viral peptides, peptide sources, peptides, etc., can solve the problem that the shortest amino acid sequence of the antigenic epitope cannot be finally determined, and the antigenic epitope cannot be distinguished.

Inactive Publication Date: 2005-10-26
河南省动物免疫学重点实验室
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The method of expressing antigenic protein by site-directed mutagenesis is one of the most commonly used techniques. This method is convenient, simple, and low in cost. However, this method can only roughly identify the key amino acid sites that exert antigenicity in protein antigens, and cannot finally determine the antigen. The shortest amino acid sequence of an epitope, and it is impossible to distinguish whether the antigenic epitope is a linear epitope or a conformational epitope

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0030] Example 1: Gene Cloning of Neutralizing B Cell Antigen Epitope Fragment of Chicken Infectious Bursal Disease Virus IBDV VP2 Protein:

[0031] SPF eggs were incubated until 11 days old, chicken embryos were taken to prepare chicken embryo fibroblasts, inoculated with IBDV-H4 strain virus, cultured at 37°C for 96 hours, and the virus was collected. The virus was separated by ultracentrifugation and 40-60% sucrose density gradient centrifugation to obtain virus particles. The virus total RNA was extracted by TRIZOL method, and the genomic RNA was treated with proteinase K to degrade the double-stranded RNA molecule, which was used as a template to amplify the IBDV A fragment gene by RT-PCR. The purified PCR product was recovered and connected to the pGEM-T vector, transformed into Escherichia coli, and the A segment gene clone was constructed.

[0032]A series of primers for overlapping expression of viral proteins were designed according to the structural characteristics...

Embodiment 2

[0034] Example 2: Fmoc solid-phase peptide synthesis method overlapping synthesis of a series of VP2 antigen peptide fragments and Dot-ELISA screening:

[0035] According to the indirect ELISA monoclonal antibody reaction spectrum of prokaryotic expression of VP2 and overlapping expression of VP2 antigen fragments, and the sequence of VP2 neutralizing antibody site screened by phage peptide library technology, the preliminary sequence of the overlapping synthetic polypeptide was determined. And apply the Chou and Fasman algorithm to analyze the secondary structure characteristics, hydrophilicity, antigenicity, surface accessibility and other characteristics of these polypeptide fragments by computer, and design the sequence of the synthetic polypeptide. And use the peptide program to analyze the difficulty of synthesis, design the peptide synthesis program, and use the peptide synthesizer to synthesize the peptide sequence by combining automatic synthesis and manual synthesis. ...

Embodiment 3

[0037] Example 3: Synthesis of Neutralizing B Cell Antigen Epitope of Chicken Infectious Bursal Disease Virus IBDV VP2 Protein:

[0038] According to the amino acid sequence of the neutralizing B cell epitope of VP2 protein, the solid-phase peptide synthesis technology is applied, and the automatic peptide synthesizer or artificial peptide synthesis method is adopted. The solid-phase resin adopts Fmoc-protected amino acid Wang resin or other resins. After the resin is swollen with DMF and piperidine is de-Fmoc-protected, Fmoc-protected amino acids are added according to the amino acid sequence order, and the acylation reaction is carried out in the presence of HBTU. After the acylation reaction is completed, after washing, the second Fmoc-amino acid is added for acylation reaction and washed. Such a cycle starts from the C-terminus of the polypeptide sequence to the N-terminus, and a complete polypeptide chain is synthesized in sequence. After the synthesis is completed, sele...

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PUM

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Abstract

The present invention belongs to the field of molecular immunology, mainly relates to series polypeptide fragment of chicken infective Fabricius bursa disease virus IBDV VP2 protein, in particular, it relates to polypeptide fragment series formed into B cell antigen apitope in VP2 protein. After the animal body is immunized by immunogen or vaccine designed by utilizing the invented chicken infective Fabricius bursa disease virus IBDV VP2 protein neutrality B cell antigen epitope, the neutrality antibody for IBDV can be produced, and can neutralize IBDV in vivo or in vitro, and can prevent the virus from infecting animal body.

Description

[0001] One. Technical field: the present invention relates to the immunology field of molecule, particularly relate to a series of neutralizing B cell epitopes of chicken infectious bursal disease virus IBDV VP2 protein. 2. Background technology: [0002] Infectious bursal disease (IBD) is an acute, highly contagious, lymphocytic infectious disease of chickens caused by infectious bursal disease virus (IBDV). It was established in the United States in 1957. Delaware State's Gumboro town broiler flocks first occurred, also known as Gambro's disease, in 1962 Winterfield isolated and identified IBDV. The disease occurs and is prevalent in the United States and other countries in the world. It broke out in a large area in my country from 1988 to 1992, causing huge economic losses. Studies have found that IBD is an immune injury disease mainly based on humoral immune response. IBDV belongs to the Birnaviridae family (Birnaviridae), and the avian birnavirus (avianbirnavirus) has two...

Claims

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Application Information

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IPC IPC(8): C07K7/06
Inventor 张改平王选年赵东乔宏兴江涛席俊杨艳艳李青梅柴书军邢广旭杨继飞李学伍肖志军邓瑞广
Owner 河南省动物免疫学重点实验室
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