Genetic engineering subunit oral vaccine strain for preventing porcine epidemic diarrhea as well as construction method and application thereof

A porcine epidemic diarrhea and genetic engineering technology, applied in the field of veterinary medicine, can solve the problems of biosafety consequences, unfavorable large-scale production, restricting the popularization and application of recombinant lactic acid bacteria preparations, etc., and achieve the effect of good probiotics

Inactive Publication Date: 2018-04-27
NORTHEAST AGRICULTURAL UNIVERSITY
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  • Abstract
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  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

In addition, antibiotic resistance is often used as a selection marker when constructing lactic acid bacteria expression systems. With the practical application of such genetically engineered lactic acid bacteria preparations, the transfer of resistance factors will bring serious biosafety consequences
In addition, the recombinant lactic acid bacteria system constructed in this research field is mainly inducible, that is, the expression of the vaccine antigen needs to be induced by an inducer, and in the absence of an inducer, the recombinant lactic acid bacteria will stop the secretion of the vaccine antigen, which is not conducive to large-scale production. production, which further restricts the popularization and application of the recombinant lactic acid bacteria preparation

Method used

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  • Genetic engineering subunit oral vaccine strain for preventing porcine epidemic diarrhea as well as construction method and application thereof
  • Genetic engineering subunit oral vaccine strain for preventing porcine epidemic diarrhea as well as construction method and application thereof
  • Genetic engineering subunit oral vaccine strain for preventing porcine epidemic diarrhea as well as construction method and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0066] The probiotic comparison of embodiment 1 Lactobacillus reuteri LJ-15 strain and Lactobacillus casei

[0067] The inventor of the present invention isolated a strain of Lactobacillus reuteri from the intestinal tract of pigs, named it Lactobacillus reuteri LJ-15 strain, and preserved it in the China Type Culture Collection Center, and its strain preservation number is CCTCC NO.M2017570 . In order to use Lactobacillus reuteri LJ-15 strain as a carrier for delivering oral vaccine immune antigens, a systematic comparative analysis of its probiotics and Lactobacillus casei commonly used in this field was carried out.

[0068] 1. Inhibit diarrhea in weaned piglets

[0069] The weaned piglets were divided into 3 groups with 10 piglets in each group. The piglets in the experimental group I were fed the diet containing L. Feed the normal diet, observe the diarrhea situation of the piglets caused by weaning stress regularly every day, and continue to observe for 30 days. The s...

Embodiment 2

[0097] Example 2 Construction of a genetically engineered subunit oral vaccine strain for preventing porcine epidemic diarrhea based on porcine-derived Lactobacillus reuteri

[0098] 1. Construction of lactic acid bacteria constitutive expression plasmid pPG-T7g10-PPT

[0099] Based on the transformation of plasmid pPG612, the plasmid pPG612 was double digested with XbaI and XhoI, and the vector fragment obtained after gel recovery was connected with the HCE-T7g10-PgsA anchor-MCS-rrnBT1T2 sequence to construct a constitutive secretory expression lactic acid bacterium Vector pPG-T7g10-PPT, wherein the nucleotide sequence of the Bacillus Geobacillus toebii constitutive secretory expression promoter HCE is shown in SEQ ID NO.1, and the nucleotide sequence of the T7g10 translation enhancer is shown in SEQ ID NO.2, The nucleotide sequence of PgsAanchor is shown in SEQ ID NO.3, the transcription terminator sequence of Escherichia coli rrnBT1T2 is shown in SEQ ID NO.4, and MCS is a m...

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Abstract

The invention discloses a genetic engineering subunit oral vaccine strain for preventing porcine epidemic diarrhea as well as a construction method and an application thereof. Autochthonous lactic acid bacteria strains which are separated from intestinal tract of pigs are used as transferring active carriers of vaccine antigens, PEDV S protein neutralizing antigen core epitope domain COE is selected as an immunogen, and at the same time transport speed of the vaccine antigen is effectively improved, immune response time of bodies is shortened, an antigen transporting cell M cell targeting peptide Col and dendritic cell targeting peptide DCpep for antigen processing and presentation is introduced, a constitutive expression carrier is used for constructing a recombined lactic acid bacteria preparation for expressing the PEDV S protein neutralizing antigen core epitope domain COE and a M cell targeting peptide Col and a dendritic cell targeting peptide DCpep, so that the vaccine antigen secretes along with breeding of genetic engineering bacterial strains, and disadvantages of induction type recombined lactic acid bacteria are avoided. An effective technical means is provided for developing the genetic engineering subunit oral vaccine strain for preventing porcine epidemic diarrhea is provided.

Description

technical field [0001] The present invention relates to a vaccine strain and a construction method thereof, in particular to a genetically engineered subunit oral vaccine strain capable of preventing porcine epidemic diarrhea and a construction method thereof, and also relates to the preparation of the vaccine strain for oral administration of porcine epidemic diarrhea. application in vaccines. The invention belongs to the technical field of veterinary medicine. Background technique [0002] Porcine epidemic diarrhea (porcine epidemic diarrhea, PED) is an acute highly contagious infectious disease of pigs caused by porcine epidemic diarrhea virus (porcine epidemic diarrhea virus, PEDV), and is an important infectious disease that endangers the healthy development of my country's pig industry. One, the disease is mainly characterized by watery diarrhea, vomiting and dehydration, and it is distributed worldwide, causing serious harm and bringing great economic losses to the pi...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K39/215A61P31/14C12N1/20C12N15/74C12R1/225
Inventor 徐义刚王丽马孙婷唐丽杰李一经姜艳平
Owner NORTHEAST AGRICULTURAL UNIVERSITY
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