Multivalent carrier vaccine for shrimp and application thereof

A vector vaccine and shrimp technology, applied in the field of genetic bioengineering, can solve the problems of insufficiency, inconvenient administration, and difficulty in the protection period of vaccine immunity exceeding 28 days, achieving the effect of long protection period and long polyvalent cross-protection

Active Publication Date: 2014-03-12
马悦 +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0008] However, there are still many shortcomings in the existing research work: 1) The administration is inconvenient, mainly by injection, and it is difficult to carry out productive breeding applications; 2) The immune protection period of the existing vaccines is difficult to exceed 28 days , cannot meet the actual needs of the immune protection period (more than 40 days) in shrimp production and breeding, so there is no production application value and development prospect
Before the present invention was made, such a vector vaccine had not been seen in any similar reports or invention patents so far.

Method used

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  • Multivalent carrier vaccine for shrimp and application thereof
  • Multivalent carrier vaccine for shrimp and application thereof
  • Multivalent carrier vaccine for shrimp and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0027] Example 1: Preparation of Multivalent Vector Vaccine Against White Spot Syndrome and Vibriosis of Shrimp

[0028] 1. Medium preparation

[0029] (1) DSM liquid medium: Bacterial nutrient broth (Difco) 8g, 10% (w / v) KCl 10ml, 1.2% (w / v) MgSO 4 ·7H 2 O 10ml, 1M NaOH 1.5ml, adjust pH to 7.6, ddH 2 Set the volume of O to 1000ml. After autoclaving, cool to 50℃, and add sterilized 1M Ca(NO 3 ) 2 , 0.01MMnCl 2 , 1mMFeSO 4 1ml each.

[0030] (2) PBS buffer: Na 2 HPO 4 ·2H 2 O 2.74g / L, NaH 2 PO 4 ·H 2 O 0.63g / L, dissolve the ingredients in 1000ml of distilled water, and sterilize at 121°C for 20 minutes.

[0031] (3) LB medium: Tryptone 10g / L, Yeast extract 5g / L, Sodium chloride (NaCl) 10g / L, adjust the pH to 7.0.

[0032] 2. Resuscitation of plate medium: Streak the seeds of the recombinant Bacillus subtilis strain stored at -80°C with the preservation number of CCTCCNo: M 2012395 on LB solid plates and cultivate overnight at 37°C to rejuvenate the recombinant Bacillus subtilis strain....

Embodiment 2

[0037] Example 2: Evaluation of the immunoprotective power of oral immunization of Penaeus vannamei against white spot syndrome

[0038] Breeding of Penaeus vannamei: The purchased experimental shrimp (Penaeus vannamei) were temporarily raised in a 3000L aquaculture tank after being tested without WSSV, with filtered fresh seawater (seawater salinity of 25g / L), and the culture water temperature was maintained at 26 ±2℃, after 1 week of holding, unhealthy individuals are eliminated. After continuing to breed for a week (average weight 10±1.2g / tail), they were randomly divided into groups, each with 30 tails, and each group paralleled 3 tanks (100L / tank).

[0039] Preparation of WSSV virus suspension: Dissecting the gills and pancreas of the sick shrimp, weighing 1g, cutting and grinding under liquid nitrogen, adding TN buffer (50 mmol / L Tris- HCl, pH 7.6, 0.4 mol / L NaCl), homogenize with a glass homogenizer in an ice bath, centrifuge the homogenate at 4°C, 7000 g for 20 min, disca...

Embodiment 3

[0045] Example 3: Evaluation of immune protection of Penaeus chinensis by oral immunization against white spot syndrome

[0046] Breeding of Penaeus chinensis: The experimental prawns (Penaeus chinensis) purchased from Tianjin are temporarily raised in a 3000L aquaculture tank after being tested for WSSV-free, with filtered fresh seawater (seawater salinity of 35g / L), and the culture water temperature is maintained at 25 ±2℃, after 1 week of holding, unhealthy individuals are eliminated. After continuing to breed for a week (average weight 10g±0.9g / tail), they were randomly divided into 100L tanks, each group of 30 fish, each group paralleled 3 tanks (100L / tank).

[0047] Preparation of WSSV virus suspension: Dissecting the gills and pancreas of the sick shrimp, weighing 1g, cutting and grinding under liquid nitrogen, adding TN buffer (50 mmol / L Tris- HCl, pH 7.6, 0.4 mol / L NaCl), homogenize with a glass homogenizer in an ice bath, centrifuge the homogenate at 4°C, 7000 g for 20 ...

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Abstract

The invention relates to a novel multivalent carrier vaccine for a shrimp. The vaccine is characterized by comprising a bacillus subtilis recombination strain obtained by a genetic engineering technique, the strain belongs to the category of bacillus subtilis HT5303 and is preserved in China Center for Type Culture Collection, and the preservation number is CCTCC NO.: M 2012395. When spores are formed by the bacillus subtilis vaccine strain, white spot syndrome virus VP281 protein can be demonstrated on the surfaces of the spores, and fusion protein of cell-penetrating peptides, white spot syndrome virus VP19 and vibrio harveyi 3-glyceraldehyde-phosphate dehydrogenase (GAPDH) can be secreted and expressed when vegetative cells are formed. Compared with other vaccines, the multivalent carrier vaccine has the advantages of cross protection and long protection period. Moreover, the vaccine can be added into feeds and has the advantages of resistance against high-temperature setting of feeds and broad prospects for commercial development.

Description

technical field [0001] The invention belongs to the technical field of genetic bioengineering, and relates to a novel polyvalent vector vaccine for prawn white spot syndrome virus (WSSV) and vibriosis and its application, in particular to a polyvalent vaccine for preventing and treating prawn white spot syndrome and vibriosis. Valence carrier vaccines and their applications. Background technique [0002] Shrimp is a high-value aquatic product mainly produced in Asia and Latin America for export purposes. At present, the main cultured species in my country are Penaeus vannamei, Penaeus monodon and Penaeus sinensis. However, in the past 10 years, there have been quite serious problems in shrimp farming, among which white spot syndrome virus (White spot syndrome virus, WSSV), taura virus (Taura Syndrome Virus, TSV) and Vibrio are the main culprits of the disease . Shrimp white spot syndrome (WSS) is an fulminant infectious disease caused by shrimp white spot syndrome virus (W...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): A61K48/00A61K39/385A61K39/295A61P31/20A61P31/04C12N1/21C12N15/34C12R1/125
Inventor 不公告发明人
Owner 马悦
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