Application of siniperca chuatsi ISKNV (Infectious Spleen and Kidney Necrosis Virus) ORF093 protein

A technology of spleen and kidney necrosis virus, ORF093, applied in the field of genetic engineering, can solve the problem that there is no research report on ISKNVORF093 gene antiserum preparations, and achieve good immune prevention effect

Active Publication Date: 2011-11-16
PEARL RIVER FISHERY RES INST CHINESE ACAD OF FISHERY SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0004] So far, there has been no research report on using the ISKNV ORF093 gene as a vaccine precursor and its antiserum preparation

Method used

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  • Application of siniperca chuatsi ISKNV (Infectious Spleen and Kidney Necrosis Virus) ORF093 protein
  • Application of siniperca chuatsi ISKNV (Infectious Spleen and Kidney Necrosis Virus) ORF093 protein
  • Application of siniperca chuatsi ISKNV (Infectious Spleen and Kidney Necrosis Virus) ORF093 protein

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Embodiment 1

[0018] 1 Prokaryotic expression of mandarin fish ISKNV ORF093 recombinant protein in Escherichia coli

[0019] 1.1 Primers were designed according to the Infectious Spleen and Kidney Necrosis Virus (ISKNV) [NC_003494] ORF093 sequence (SEQ ID NO: 1) in GenBank, and a restriction enzyme cleavage site was introduced (underlined). The primer sequences are as follows:

[0020] P1:【5`-CG GAATTC ATGAATAAAACGC -3'] (SEQ ID NO: 3), introducing EcoRI restriction site;

[0021] P2: [5'-CGG AAGCTT TTAAACATGGCGTC -3'] (SEQ ID NO: 4), introducing a HindⅢ restriction site.

[0022] 1.2 Use the ISKNV genome of mandarin fish self-preserved in our laboratory as a template for PCR amplification. The 25 μL PCR reaction system contains: 10×buffer (including Mg2+) 2.5 μL, 4×dNTP (10 mmol / L) 0.5 μL, upstream and downstream primers (20 μmol / L) 0.5 μL each, Taq DNA polymerase (5U / μL) 0.3 μL, DNA 1.0 μL, and sterile double distilled water to make up. PCR reaction conditions: pre-denaturation at...

Embodiment 2

[0065] 1. Construction of ISKNV ORF093 eukaryotic expression vector

[0066] 1.1 Primers were designed according to the Infectious Spleen and Kidney Necrosis Virus (ISKNV) [NC_003494] ORF093 sequence (SEQ ID NO: 1) in GenBank, and a restriction enzyme cleavage site was introduced (underlined). The primer sequences are as follows:

[0067] P3: [5'-AT GGTACC A TGGTTTCGAGTGCACTGCT-3'] (SEQ ID NO: 5), introduced Kpn Ⅰ Restriction site;

[0068] P4:【5'-AT GAATTC TTAGGCCATAATGCCGCG-3'] (SEQ ID NO: 6), introducing Eco RI restriction site.

[0069]1.2 Use the pET32a-ORF 093 plasmid as a template for PCR amplification. The 25 μL PCR reaction system contains: 10×buffer (including Mg2+) 2.5 μL, 4×dNTP (10 mmol / L) 0.5 μL, upstream and downstream primers (20 μmol / L) each 0.5 μL, Taq DNA polymerase (5U / μL) 0.3 μL, DNA 1.0 μL, supplemented with sterile double distilled water. PCR reaction conditions: pre-denaturation at 94°C for 4min; denaturation at 94°C for 45S, renaturation a...

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Abstract

The invention discloses application of a siniperca chuatsi ISKNV (Infectious Spleen and Kidney Necrosis Virus) ORF093 protein in preparing a vaccine or a medicament for preventing a siniperca chuatsi ISKNV disease. The siniperca chuatsi ISKNV ORF093 protein plays a better immunoprophylaxis role on a siniperca chuatsi iridescent virus disease, has protection ratio exceeding 45 percent and can be developed into a new vaccine or a new medicament for resisting the siniperca chuatsi ISKNV disease.

Description

technical field [0001] The invention belongs to the technical field of genetic engineering, and in particular relates to the application of ORF093 protein of mandarin fish infectious spleen and kidney necrosis virus (ISKNV). Background technique [0002] mandarin fish ( Siniperca chuatsi ) is one of the important freshwater fish characteristic breeding species in my country. The iridescent virus disease of mandarin fish caused by Infectious spleen and kidney necrosis virus (ISKNV) is the main disease of mandarin fish culture, and the mortality rate of mandarin fish is close to 100%, becoming the main bottleneck restricting the development of mandarin fish aquaculture. For a long time, there has been no effective drug for the prevention and treatment of mandarin fish virus disease, and there are many disadvantages in the prevention and treatment of diseases such as antibiotics and chemical drugs, such as pathogenic drug resistance, environmental pollution, food safety issues,...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K39/12A61P31/20
Inventor 付小哲吴淑勤李宁求石存斌潘厚军
Owner PEARL RIVER FISHERY RES INST CHINESE ACAD OF FISHERY SCI
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