Method for preparing recombinant spore for surface display of prawn white spot syndrome virus Vp28

A technology of white spot syndrome and surface display, applied in the direction of botanical equipment and methods, microorganism-based methods, biochemical equipment and methods, etc., to achieve good stability

Inactive Publication Date: 2011-05-04
JIANGSU UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0004] The purpose of the present invention is to display the Vp28 protein on the surface of the Bacillus subtilis spore, construct the Bacillus subtilis recombinant spore displaying the shrimp white spot syndrome virus protein Vp28 on the surface, make the Vp28 protein obtain good stress resistance, and be suitable for oral inoculation of prawns, but so far No similar reports or invention patents have been seen yet

Method used

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  • Method for preparing recombinant spore for surface display of prawn white spot syndrome virus Vp28
  • Method for preparing recombinant spore for surface display of prawn white spot syndrome virus Vp28
  • Method for preparing recombinant spore for surface display of prawn white spot syndrome virus Vp28

Examples

Experimental program
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Effect test

Embodiment 1

[0041] Preparation of recombinant spores of Bacillus subtilis displaying Vp28 on the surface using CotC as carrier protein

[0042] 1. Molecular Biology Operations

[0043] 1.1 Extraction of Bacillus subtilis chromosome

[0044] Collect 10mL Bacillus subtilis168 (trp - ) (Bacillus Genetic Stock Center, Department of Biochemistry The Ohio State University 484 West Twelfth Avenue Columbus, Ohio 43210, USA) culture, add 0.5ml TE to suspend the precipitate. Add 30μl lysozyme (100mg / ml) to each microcentrifuge tube, react at 37°C for 1 hour; add 50μl SDS (10%) and 20μl proteinase K (20mg / ml), shake evenly, react at 37°C for 2 hours, add etc. Take the supernatant after extraction with phenol / chloroform, add 2 times the volume of ethanol, precipitate the DNA for more than 2 hours at room temperature, centrifuge at 12,000g for 10 minutes, discard the supernatant and wash the DNA precipitate with 500μl of 75% ethanol to Remove inorganic salt ions. After the DNA precipitate is dry, ...

Embodiment 2

[0074] Preparation of recombinant spores of Bacillus subtilis displaying Vp28 on the surface using CotG as carrier protein

[0075] 1. Molecular Biology Operations

[0076] 1.1 Extraction of virus chromosome

[0077] Concrete operation method is with the extraction of embodiment 1 of the present invention, 1.1 virus chromosome

[0078] 1.2 Extraction of Bacillus subtilis chromosome

[0079] Concrete operation method is with the extraction of the embodiment of the present invention 1,1.2 Bacillus subtilis chromosome

[0080] 1.3 Molecular biology techniques

[0081] Concrete operation method is with the embodiment of the present invention 1,1.3 molecular biology technology

[0082] 1.4PCR amplification

[0083] Concrete operation method is with the embodiment of the present invention 1,1.4 molecular biology technology

[0084] 2. Plasmid construction

[0085] 2.1 Basic plasmid construction

[0086] Concrete operation method is the same as embodiment 1, the construction ...

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Abstract

The invention relates to a method for preparing a recombining spore of surface display foreign protein, in particular to a recombining spore for surface display prawn white spot syndrome virus Vp28 protein of a bacillus subtilis spore. In the method, a bacillus subtilis spore capsid protein gene is used as a molecule vector and is recombined with an antigen protein gene on the surface of the prawn white spot syndrome virus to construct an integral recombining plasmid in fusion expression; the bacillus subtilis is converted to a bacillus subtilis recombining strain; and the recombining spore of the spore surface display prawn white spot syndrome virus protein produced by the recombining strain is induced.

Description

technical field [0001] The invention relates to a preparation method of a recombinant spore displaying foreign protein on the surface, in particular to a recombinant spore displaying the Vp28 protein of shrimp white spot syndrome virus on the surface of the bacillus subtilis spore. Background technique [0002] Shrimp white spot syndrome is a comprehensive disease caused by white spot syndrome virus (WSSV), which causes huge economic losses to shrimp farming. Shrimp white spot syndrome virus vaccine is the most effective measure to control virus infection. Monodon shrimp were orally inoculated with recombinant WSSV envelope protein rVp28 expressed in Escherichia coli, and the protection rate reached 77% (WitteveldtJ, et al. 2004. J. Virol. 78(4): 2057-61). However, recombinant Vp28 or Vp19 protein vaccines have poor stability and low utilization in water environments; oral soluble antigen (or recombinant antigen) vaccines are easily degraded by proteases in the digestive tra...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/62C12N15/40C12N15/31C12N15/75C12N1/21C12R1/125
Inventor 宁德刚徐卫东李倩吴春笃
Owner JIANGSU UNIV
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