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CTL epitope polypeptide of porcine reproductive and respiratory syndrome virus (PRRSV) and application thereof

A technology of recombining viral vectors and expression cassettes, applied in the fields of application, antiviral agents, viral antigen components, etc., can solve problems such as ambiguity of the method and not involving the restriction of the SLA-I allele of the pig species

Inactive Publication Date: 2013-08-14
CHINA AGRI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, these methods are very vague and do not address the restriction of the pig species SLA-I allele

Method used

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  • CTL epitope polypeptide of porcine reproductive and respiratory syndrome virus (PRRSV) and application thereof
  • CTL epitope polypeptide of porcine reproductive and respiratory syndrome virus (PRRSV) and application thereof
  • CTL epitope polypeptide of porcine reproductive and respiratory syndrome virus (PRRSV) and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0077] Example 1, Selection and Synthesis of CTL Epitope Polypeptides of Porcine Reproductive and Respiratory Syndrome Virus

[0078] According to the NetMHCpan 2.0server (http: / / www.cbs.dtu.dk / services / NetMHCpan) website, the PRRSV-derived CTL epitope polypeptide that SLA-I is restricted to bind is predicted, and an amino acid sequence is selected on the NSP9 protein of PRRSV It is the CTL cell epitope polypeptide of sequence 1 in the sequence listing (its nucleotide sequence is sequence 2 in the sequence listing, named NSP9 -TMP ), polypeptide NSP9 -TMP It has a higher affinity prediction score (0.540) for SLA-I. Peptide NSP9 -TMP Send to Beijing Zhongke Yaguang Biotechnology Co., Ltd. for synthesis, and purify by HPLC (High Performance Liquid Chromatography), with a purity of ≥90%. And identified by mass spectrometry (MS).

Embodiment 2

[0079] Embodiment 2, the binding ability of the CTL epitope polypeptide of porcine reproductive and respiratory syndrome virus and SAL-I molecule is detected by protein folding renaturation experiment

[0080] 1. Preparation of porcine SAL-I molecular light chain β2m inclusion body protein

[0081] The construction process of the β2m inclusion body protein recombinant prokaryotic expression vector is as follows: DNA (nucleoside The acid sequence (as shown in Sequence 4 in the sequence table) was inserted between the restriction sites EcoR I and HindIII of the prokaryotic expression plasmid pET-21a(+), to obtain a recombinant expression plasmid named pET-21a(+) / sβ2m.

[0082]The preparation process of the light chain β2m inclusion body protein is as follows: (1) Transform the pET-21a(+) / sβ2m plasmid into BL21(DE3) competent cells, and culture at 37°C overnight. (2) Pick a monoclonal colony and inoculate it into 100 mL of LB liquid medium containing Amp at 37° C. and 200 rpm fo...

Embodiment 3

[0093] Example 3. Detection of CTL Epitope Immunological Activity of Porcine Reproductive and Respiratory Syndrome Virus by Tetramer Technology

[0094] 1. Preparation of porcine SAL-I molecular heavy chain inclusion body protein with BirA substrate peptide (BSP) sequence added to the carboxy-terminus

[0095] The extracellular domain amino acid (amino acid sequence is as the sequence in the sequence listing) of porcine SAL-I class molecular heavy chain SLA-I (SLA-1*1502, GenBank No.HQ909439) encoding the sequence of adding BirA substrate peptide (BSP) at the carboxy-terminus 7) DNA molecule (nucleotide sequence as shown in sequence 8 in the sequence table) is inserted between the restriction sites EcoR I and Hind III of the prokaryotic expression plasmid pET-21a (+), to obtain the recombinant expression plasmid, Designated as pET-21a(+) / SLA-1*1502-BSP. The specific preparation process of the porcine SAL-I molecular heavy chain inclusion body protein (named SLA-1*1502-BSP) wi...

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PUM

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Abstract

The present invention discloses a CTL epitope polypeptide of porcine reproductive and respiratory syndrome virus (PRRSV) and application thereof. The amino acid sequence of the CTL epitope polypeptide of PRRSV provided by the present invention is as shown in sequence 1 in a sequence table. The invention also protects a nucleic acid molecule encoding the polypeptide. The CTL epitope polypeptide of PRRSV provided by the present invention can combine with a swine SLA-I molecule, can be used for preparing a vaccine for PRRSV and preparing a chicken CD8 + T lymphocyte proliferation agent.

Description

technical field [0001] The invention relates to a CTL epitope polypeptide of porcine reproductive and respiratory syndrome virus and its application. Background technique [0002] Porcine Reproductive and Respiratory Syndrome (PRRS) is a highly contagious disease of pigs caused by Porcine Reproductive and Respiratory Syndrome Virus (PRRSV). Pigs of different breeds, ages and sexes can be infected, with pregnant sows and piglets within 1 month of age being the most susceptible. PRRS is mainly characterized by abortion, stillbirth, weak fetus, mummified fetus, dyspnea, sepsis and high mortality in piglets. The disease was first reported in the United States in 1987, and it quickly spread to countries and regions with developed pig farming in the world in just a few years. Initially, because people could not determine its cause, it was called "swine mystery disease" (swine Mystery disease, SMD), "swine infertility and respiratory syndrome" (Swine infertility and respiratory s...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K7/06C12N15/40C12N15/63C12N5/10C12N1/15C12N1/19C12N1/21C12N15/86C12N5/0783A61K48/00A61K39/12A61P31/14
Inventor 夏春潘孝成
Owner CHINA AGRI UNIV
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