Method for preparing tetramer from Asia I 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 Asia I foot and mouth disease virus polypeptide
  • Method for preparing tetramer from Asia I foot and mouth disease virus polypeptide
  • Method for preparing tetramer from Asia I 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 Amethyst; 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 ...

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Abstract

The invention belongs to the technical field of biology and particularly relates to a method for preparing tetramer from Asia I foot and mouth disease virus polypeptide. According to the designed foot and mouth disease virus polypeptide, the foot and mouth disease virus polypeptide is screened and identified by the ELISPOT method and the tetramer construction method, pouch pig SLA-2 heavy-chain tetramer is constructed and expressed in a prokaryotic expression system pET-21a(+) / BL21, and an inclusion body is extracted. Polypeptide capable of causing specificity CTL reaction is screened from the foot and mouth disease virus polypeptide through the ELISPOT method, the polypeptide and SLA-2 heavy-chain and known light-chain beta2m are subjected to renaturation and biotinylation, the polypeptide and FITC labeled-streptavidin react to generate SLA-2-BSP / tetramer, and then the function of tetramer is detected through flow cytometry. A specificity peptide epitope of a pig Asia I foot and mouth disease virus VP1 is obtained through identification through the ELISPOT method, the tetramer is successfully constructed, the specificity CTL of the tetramer is detected, a pig tetramer technical platform is primarily built, and a foundation is laid for researching pig specificity CTL immune response 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 Asian type I 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 ...

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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