Preparation method and application of bivalent DNA (Deoxyribose Nucleic Acid) vaccine for preventing prawn white spot syndrome virus

A DNA vaccine, a technology for white spot syndrome, which is applied in the field of preventing shrimp white spot syndrome virus bivalent DNA vaccine, and can solve problems such as difficulty in virus preservation and proliferation.

Active Publication Date: 2015-12-30
湛江出入境检验检疫局检验检疫技术中心
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, there is currently no cell line that can be used for the propagation of shrimp white spot syndrome virus, so it i

Method used

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  • Preparation method and application of bivalent DNA (Deoxyribose Nucleic Acid) vaccine for preventing prawn white spot syndrome virus
  • Preparation method and application of bivalent DNA (Deoxyribose Nucleic Acid) vaccine for preventing prawn white spot syndrome virus
  • Preparation method and application of bivalent DNA (Deoxyribose Nucleic Acid) vaccine for preventing prawn white spot syndrome virus

Examples

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Embodiment 1

[0073] The preparation method of preventing prawn white spot syndrome virus bivalent DNA vaccine, the steps are as follows:

[0074] Step 1: Design primers for the amplification of white spot virus VP19 and VP28 genes, as follows:

[0075] 1. According to the VP19 gene sequence, use PrimerPremier5 software to synthesize a pair of primers:

[0076] (1-1) Upstream primer: 5- GGATCC ATGGCCACCACGACTAACAC-3, where the underlined GGATCC for BamHI;

[0077] (1-2) Downstream primer: 5- GAATTC TTACTGCCTCCTCTTGGGGT-3, where the underlined GAATTC is EcoRI;

[0078] 2. According to the VP28 gene sequence, use PrimerPremier5 software to synthesize a pair of primers:

[0079] (2-1) Upstream primer: 5-AGA GAATTC ATGGATCTTTCTTTCAC-3, where the underlined GAATTC is EcoRI;

[0080] (2-2) Downstream primer: 5-CAC AAGCTT TTACTCGGTCTCAGTGC-3, where the underlined AAGCTT is XhoI;

[0081] Step 2: Extraction of total white spot virus DNA, as follows:

[0082] Use the DNA extractio...

Embodiment 2

[0124] The preparation method of preventing prawn white spot syndrome virus bivalent DNA vaccine, the steps are as follows:

[0125] Step 1: Design primers for the amplification of white spot virus VP19 and VP28 genes, as follows:

[0126] 1. According to the VP19 gene sequence, use PrimerPremier5 software to synthesize a pair of primers:

[0127] (1-1) Upstream primer: 5- GGATCC ATGGCCACCACGACTAACAC-3, where the underlined GGATCC for BamHI;

[0128] (1-2) Downstream primer: 5- GAATTC TTACTGCCTCCTCTTGGGGT-3, where the underlined GAATTC is EcoRI;

[0129] 2. According to the VP28 gene sequence, use PrimerPremier5 software to synthesize a pair of primers:

[0130] (2-1) Upstream primer: 5-AGA GAATTC ATGGATCTTTCTTTCAC-3, where the underlined GAATTC is EcoRI;

[0131] (2-2) Downstream primer: 5-CAC AAGCTT TTACTCGGTCTCAGTGC-3, where the underlined AAGCTT is XhoI;

[0132] Step 2: Extraction of total white spot virus DNA, as follows:

[0133] Use the DNA extractio...

Embodiment 3

[0175] The preparation method of preventing prawn white spot syndrome virus bivalent DNA vaccine, the steps are as follows:

[0176] Step 1: Design primers for the amplification of white spot virus VP19 and VP28 genes, as follows:

[0177] 1. According to the VP19 gene sequence, use PrimerPremier5 software to synthesize a pair of primers:

[0178] (1-1) Upstream primer: 5- GGATCC ATGGCCACCACGACTAACAC-3, where the underlined GGATCC for BamHI;

[0179] (1-2) Downstream primer: 5- GAATTC TTACTGCCTCCTCTTGGGGT-3, where the underlined GAATTC is EcoRI;

[0180] 2. According to the VP28 gene sequence, use PrimerPremier5 software to synthesize a pair of primers:

[0181] (2-1) Upstream primer: 5-AGA GAATTC ATGGATCTTTCTTTCAC-3, where the underlined GAATTC is EcoRI;

[0182] (2-2) Downstream primer: 5-CAC AAGCTT TTACTCGGTCTCAGTGC-3, where the underlined AAGCTT is XhoI;

[0183] Step 2: Extraction of total white spot virus DNA, as follows:

[0184] Use the DNA extractio...

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Abstract

The invention discloses a preparation method and application of a bivalent DNA (Deoxyribose Nucleic Acid) vaccine for preventing prawn white spot syndrome virus. When primers are designed, two restriction enzyme cutting sites are added at two ends of an upstream primer and a downstream primer, so that antigen genes of VP19 and VP28 obtained by cloning carry double restriction enzyme cutting sites; when double restriction enzyme cutting is performed, target genes are enabled to obtain cohensive ends; the antigen genes of VP19 and VP28 are serially connected and are cloned to a prokaryotic expression vector by using DNA ligase and are used for the prokaryotic expression vector; when the bivalent DNA vaccine is constructed, VP19 gene modification is needed, i.e., a termination codon of the VP19 gene is removed by primer design; when the primer design is performed, the restriction enzyme cutting site on the downstream of the VP19 gene is identical with that on the upstream of the VP28 gene, so that the VP19 gene and the VP28 gene are serially connected and are cloned by a restriction enzyme cutting technology.

Description

technical field [0001] The invention belongs to the technical field of DNA vaccines, in particular to a preparation method and application of a bivalent DNA vaccine for preventing prawn white spot syndrome virus, and specifically to the technical field of biological veterinary medicine. The vaccine contains white spot syndrome virus (whitespot syndrome virus, WSSV) Envelope protein VP19 antigen gene and envelope protein VP28 antigen gene. At the same time, the vaccine contains PVAX1.0 vector. Background technique [0002] Whitespot syndrome (Whitespotsyndrome) is a fulminant infectious disease of prawns caused by whitespot syndrome virus (WhitespotsyndromeVirus, WSSV). huge economic losses. WSSV is a kind of enveloped non-inclusion body baculovirus, which belongs to the Nimaviridea family. It is a circular double-stranded DNA virus with an envelope. The DNA of WSSV virus is 290KD, with 181 ORFs, including 39 structural proteins. and 9 membrane proteins, including VP15, VP...

Claims

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Application Information

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IPC IPC(8): A61K48/00A61K39/295A23K1/16A23K1/18A61P31/20
Inventor 张娜马新华龙阳谢艳辉袁俊杰刘骁杨劲李红权徐金龙孙良娟
Owner 湛江出入境检验检疫局检验检疫技术中心
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