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Subunit vaccine for grouper iridovirus and preparation method and application of vaccine

A subunit vaccine, iridescent virus technology, applied in the fields of molecular biology and immunology, to achieve high protection rate, improved immune protection effect, and low cost

Active Publication Date: 2018-05-29
SOUTH CHINA AGRI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

No recombinant subunit vaccines against Singaporean grouper iridescent virus SGIV have been reported

Method used

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  • Subunit vaccine for grouper iridovirus and preparation method and application of vaccine
  • Subunit vaccine for grouper iridovirus and preparation method and application of vaccine

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0031] The immunoprotective subunit vaccine is encoded by the base sequence of VP72 shown in SEQ ID NO.1 in the sequence table.

[0032] The preparation method of immunoprotective subunit vaccine:

[0033] 1) Construction of plasmid pET32a-VP72

[0034]Plasmid pET32 (Novagen, USA) was digested with BamHI / XhoI, and a 5.9 kb fragment was recovered. Using the independently isolated and preserved Singapore grouper iridescent virus SGIV DNA as a template, use primer VP72-F: 5'-TAAGGATCCATGCAAGAGTTTTCGGTCGG-3', primer VP72-R: 5'-AATCTCGAGTGGACATTGTTACAGTGATGCT AGC-3' to carry out PCR amplification according to the following conditions VP72 base sequence (shown in SEQ ID NO.1), specific PCR conditions are: 95°C 180s pre-denatured template DNA, then 94°C 30s, 55°C 60s, 72°C 60s, 35 cycles, and then extended at 72°C React for 10 minutes. After the PCR product was purified, it was digested with BamHII / XhoI, and the 1.1kb fragment was recovered, ligated with the above 5.9kb fragment, ...

Embodiment 2

[0038] Immune Application of Recombinant Subunit Vaccine

[0039] Step 1) Preparation of subunit vaccine protein and adjuvant mixture.

[0040] Subunit vaccine protein mixture: the subunit vaccine protein rVP72 (250mg / L, purity more than 90%) purified in Example 1 is adjusted to contain 200mg of bases in the sequence table SEQ ID No1 in each liter of buffer solution with PBS buffer Sequence-coded vaccine protein solution (i.e. containing 200 mg subunit vaccine protein rVP72 / L) to obtain a subunit vaccine protein mixture;

[0041] Subunit vaccine protein and adjuvant mixture: mix the subunit vaccine protein rVP72 (250mg / L, purity above 90%) purified in Example 1 with aluminum hydroxide solution, and adjust it to the concentration in each liter of buffer solution with PBS buffer solution. The vaccine protein (subunit vaccine protein rVP72) that contains the base sequence code in 200mg sequence table SEQ ID No.1 8 and a mixed solution of 0.1 mg / mL aluminum hydroxide (that is, c...

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Abstract

The invention discloses a subunit vaccine for grouper iridovirus and a preparation method and application of the vaccine. The vaccine for the grouper iridovirus is protein of a nucleotide sequence code as shown in SEQ ID No.1. Plasmid pET32a-VP72 is built, escherichia coli BL21(DE3) is converted, induction expression is conducted, cells are broken, then recombinant protein is recycled from supernatant, and the vaccine for the grouper iridovirus is obtained. Recombinant protein rVP72 can be mixed with adjuvant, and obtained vaccine mixed liquor has the immune protection effect on the Singaporegrouper iridovirus (SGIV). According to the vaccine for the grouper iridovirus, the immune effect is stable, the protection rate is high, the efficient protectiveness is achieved, and the vaccine canbe applied to the phenomena that in the grouper breeding process, fry and adult fishes are protected, and grouper iridovirus infection is resisted.

Description

Technical field: [0001] The invention belongs to the fields of molecular biology and immunology, and specifically relates to an immunoprotective Singapore grouper iridescent virus recombinant main capsid protein subunit vaccine and its preparation method and application. Background technique: [0002] Grouper is a marine fish with high economic value. In recent years, the scale of coastal grouper farming has been expanded, but the breeding environment is deteriorating day by day, and various diseases, especially viral diseases, frequently break out, causing significant economic losses. Iridescent virus is an important pathogenic agent of grouper, which can cause fish mortality from 30% (in adult fish) to 100% (juvenile stage). In Asia, especially when the water temperature is above 25°C, this virus is very likely to infect rare seawater fish cultured in East Asia and South Asia, such as grouper, barramundi, red sea bream, flounder and large yellow croaker, etc. , resulting...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K39/12A61K39/39A61P31/20C12N15/70C12N15/66C12N15/34C07K14/01
CPCA61K39/12A61K39/39A61K2039/55505C07K14/005C12N15/66C12N15/70C12N2710/14022
Inventor 秦启伟黄晓红周胜魏京广黄友华
Owner SOUTH CHINA AGRI UNIV
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