Method for preparing tetramer from O type foot and mouth disease virus polypeptide

A foot-and-mouth disease virus, tetramer technology, applied in viral peptides, viruses/phages, biochemical equipment and methods, etc.

Active Publication Date: 2016-08-24
DALIAN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] At present, the Tetramer technology has not been applied to the identification of the function of the foot-and-mouth disease virus polypeptide bound by the porcine SLA class I molecule in China.

Method used

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  • Method for preparing tetramer from O type foot and mouth disease virus polypeptide
  • Method for preparing tetramer from O type foot and mouth disease virus polypeptide
  • Method for preparing tetramer from O type foot and mouth disease virus polypeptide

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Embodiment Construction

[0062] Now further illustrate the present invention in conjunction with embodiment.

[0063] First, the reagents involved in the present invention are described;

[0064] IPTG, DNA Marker and Protein Molecular Weight Marker are all products of TaKaRa Company; Propionamide, PBS solution, NH 4 Cl, DMSO, Tris-base, L-arginine, EDTA, reduced glutathione (GSH), oxidized glutathione (GSSG), PMSF, ammonium persulfate, TEMED, DTT, Coomassie brilliant blue R250 Purchased from Sangon Bioengineering Co., Ltd.; BCA protein concentration assay kit, western blot developer and fixer were purchased from Beyontian; protein concentration ultrafiltration tubes were purchased from MilliPore; Anti-His Tag Mouse Monoclonal Antibody and HRPConjugated Goat anti -MouseIgG was purchased from Ametek; ELISpot PLUSfor Porcine IFN-γ Kit was purchased from Mabtech Company; Porcine Peripheral Blood Lymphocyte Separation Kit was purchased from TBD Company; Con-A (concanavalin A) was purchased from SIGMA Co...

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Abstract

The invention belongs to the technical field of biology, and particularly relates to a method for preparing tetramer from O type foot and mouth disease virus polypeptide. Aiming at designed foot and mouth disease virus polypeptide, two methods of ELISPOT and tetramer construction are adopted for screening and identifying the foot and mouth disease virus polypeptide, a Hebao pig SLA-2 heavy chain tetramer is constructed, expression is carried out on a prokaryotic expression system pET-21a(+)/BL21, and an inclusion body is extracted; in the foot and mouth disease virus polypeptide, polypeptide causing a specific CTL reaction is screened out through an ELISPOT method, then the polypeptide, the SLA-2 heavy chain, and existing light chains beta 2m are subjected to renaturation, biotinylation and FITC label streptavidin reactions to generate the SLA-2-BSP/peptide tetramer, and then the function of the tetramer is detected through flow cytometry. Through an ELISPOT method, a specificity polypeptide epitope of O type foot and mouth disease viruses VP1 is obtained through identification, the peptide tetramer is successfully constructed, a pig tetramer technology platform is preliminarily built, and a foundation is laid for study of pig specificity CTL immune responses and T epitope screening.

Description

technical field [0001] The invention belongs to the field of biotechnology, and in particular relates to a method for preparing a tetramer of an O-type foot-and-mouth disease virus polypeptide. Background technique [0002] Foot-and-mouth disease is an acute thermal and highly contagious infectious disease caused by Foot and Mouth Disease Virus (FMDV). It mainly affects artiodactyls. Pigs are a relatively susceptible group. Blisters and festers are the main features, and are the most contagious among livestock diseases. Every outbreak of foot-and-mouth disease will bring great losses to the agricultural economy. Therefore, the International Office of Epizootics (OIE) has always listed it as the top of the list of Class A animal diseases that must be reported. Therefore, the prevention and diagnosis of FMD in pigs is particularly important. At present, the prevention and treatment of FMD virus mainly relies on traditional inactivated vaccines. Inactivated vaccines have the...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K14/09
CPCC07K14/005C12N2770/32122
Inventor 高凤山
Owner DALIAN UNIV
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