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Antigen epitope III of neutrality B cell of chicken IBDV VP2 protein and application thereof

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: 2007-10-17
HENAN INST OF SCI & TECH
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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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  • Antigen epitope III of neutrality B cell of chicken IBDV VP2 protein and application thereof

Examples

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Embodiment 1

[0026] Example 1: Gene Cloning of Neutralizing B Cell Antigen Epitope Fragment of Chicken Infectious Bursal Disease Virus IBDV VP2 Protein:

[0027] 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. Recover and purify the PCR product, connect it with the pGEM-T vector, transform Escherichia coli, and construct the A segment gene clone.

[0028] A series of primers for overlapping expression of viral proteins were designed according to the structural characteristics and antigenicit...

Embodiment 2

[0030] Example 2: Fmoc solid-phase peptide synthesis method overlapping synthesis of a series of VP2 antigen peptide fragments and Dot-ELISA screening:

[0031] 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

[0033] Example 3: Synthesis of Neutralizing B Cell Antigen Epitope of Chicken Infectious Bursal Disease Virus IBDV VP2 Protein:

[0034] 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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Abstract

The present invention belongs to molecular immunology field, mainly relating to chicken infectious bursal disease virus IBDV VP2 polypeptide fragment, more specificly relating to B cell antigen epitope polypeptide fragment in VP2 protein. The present invention also relates to IBDV VP2 protein neutrality and uses of B cell antigen epitope in preparing immunogen, immune animal, immune protection and uses for detecting antibody of resisting chicken infectious bursal disease virus IBDV or polypeptide of chicken infectious bursal disease virus IBDV. Animal is immunized by immunogen or vaccine designed according to B cell antigen epitope which can neutralize chicken IBDV VP2 protein and can produce antibody for neutralizing IBDV in vitro or vivo, so the immunogen can prevent virus infection to animal. Chicken IBDV VP2 protein neutralized B cell antigen epitope vaccine can prevent infection of chicken IBD classic toxic strain, superstrong toxic strain or / and variant toxic strain.

Description

[0001] This application is a divisional application with the application number 200510017453.6. The applicant of the original application is Henan Institute of Science and Technology. The original application date is March 29, 2005. The original invention title is "Chicken IBDV VP2 protein neutralizing B cell antigen epitope II and its application". [0002] 1. Technical field: the present invention relates to the immunology field of molecule, particularly relate to a series of chicken infectious bursal disease virus IBDV VP2 protein neutralizing B cell epitope and application thereof. 2. Background technology: [0003] Infectious bursal disease (IBD) is an acute, highly contagious, lymphocytic infectious disease of chickens caused by infectious bursal disease virus (IBDV). In 1957, it occurred for the first time in broiler chickens in Gumboro Township, Delaware State, USA, also known as Gambro disease. IBDV was isolated and identified by Winterfield in 1962. The disease occ...

Claims

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Application Information

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IPC IPC(8): C07K7/08C07K14/18
Inventor 王选年张改平李培庆王三虎王自良保银梅
Owner HENAN INST OF SCI & TECH
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