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Salmonella typhimurium aroA and luxS double-gene-deleted strain and attenuated vaccine prepared by same

A technology of Salmonella typhimurium and attenuated vaccine, which is applied in the biological field, can solve the problems of reducing bacterial virulence, and achieve the effects of reducing proliferation ability, reducing virulence, and maintaining good immunity

Active Publication Date: 2017-08-11
SHANGHAI VETERINARY RES INST CHINESE ACAD OF AGRI SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] Studies at home and abroad have shown that the deletion, transformation and modification of regulatory genes and metabolism-related genes can reduce the virulence of bacteria, such as aroA and luxS, among which the aroA gene encodes 3-enolacetone in the biosynthesis pathway of aromatic amino acids Acylshikimate-5-phosphate synthase, the loss of function of this gene leads to the growth of bacteria requiring tyrosine, phenylalanine, tryptophan, etc. In the absence of these metabolites, the growth of bacteria will be inhibited

Method used

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  • Salmonella typhimurium aroA and luxS double-gene-deleted strain and attenuated vaccine prepared by same
  • Salmonella typhimurium aroA and luxS double-gene-deleted strain and attenuated vaccine prepared by same
  • Salmonella typhimurium aroA and luxS double-gene-deleted strain and attenuated vaccine prepared by same

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Embodiment 1

[0018] 1. Salmonella typhimurium aroA and luxS double gene deletion strain and its construction.

[0019] The Salmonella typhimurium aroA and luxS double gene deletion strain provided by the present invention is constructed by using the Red homologous recombination method, which specifically includes the following steps:

[0020] Step 1: Construction of the aroA targeting fragment. The first primer pair was designed according to the pKD3 sequence. The 5' ends of the two primers in the first primer pair respectively contained the upstream gene of the aroA gene and the downstream gene of the aroA gene. Using pKD3 as a template, the The first primer pair amplifies the chloramphenicol resistance gene by PCR to obtain the aroA targeting fragment containing the upstream gene of the aroA gene and the downstream gene of the aroA gene,

[0021] As shown in Table 1, in this embodiment, the names of the first primer pair are aroA deletion-F and aroA deletion-R, and their sequences are sh...

Embodiment 2

[0051] This embodiment provides a Salmonella typhimurium double gene deletion strain attenuated vaccine prepared according to Example 1 to obtain the Salmonella typhimurium double gene deletion strain, and the following methods are used to evaluate its immune effect and optimal immune dose:

[0052] Divide 40 BALB / c mice into 5 groups, 8 mice in each group, with immunization dose of 1×10 7 CFU / piece, 1×10 6 CFU / piece, 1×10 5 CFU / piece, 1×10 4 CFU / only and PBS blank control group. After 2 weeks of immunization, use 1×10 6 The CFU wild strain SAT52 strain was challenged, the number of survival and death in each group was observed, and the immune protection effect of different immunization doses was evaluated.

[0053] The results showed that all the PBS control group died, and the immune dose was 1×10 7 CFU / piece, 1×10 6 The protection rate of CFU / only was 100%, and the immune dose was 1×10 5 CFU / piece, 1×10 4 The protection rates of CFU / only were 87.5% and 62.5% respect...

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Abstract

The invention provides a salmonella typhimurium aroA and luxS double-gene-deleted strain and an attenuated vaccine prepared by the same. The salmonella typhimurium aroA and luxS double-gene-deleted strain is collected at China General Microbiological Culture Collection Center, and the collection number of the strain is CGMCC NO. 13736. The double-gene-deleted strain is obtained by deleting aroA and luxS genes by a Red homologous recombination method, so that toxicity is remarkably reduced, the prepared salmonella typhimurium double-gene-deleted attenuated vaccine is good in immunity maintenance, and the strain can be used for preparing the salmonella typhimurium double-gene-deleted attenuated vaccine for preventing and treating salmonella typhimurium infection.

Description

technical field [0001] The invention belongs to the field of biotechnology, and in particular relates to a Salmonella typhimurium aroA and luxS double gene deletion strain and an attenuated vaccine prepared therefrom. Background technique [0002] Salmonella is an important food-borne zoonotic pathogen, which can not only cause a variety of livestock and poultry diseases (fowl pullorum, chicken typhoid, chicken paratyphoid, swine paratyphoid, etc.), cause systemic sepsis and enteritis, and give Aquaculture causes serious economic losses, and can cause human food poisoning by contaminating food, posing a serious threat to human health. Salmonella typhimurium can infect many poultry, livestock, rats, birds and cold-blooded animals, and flies and fleas can carry the bacteria. Humans are usually infected with Salmonella due to ingestion of contaminated eggs or undercooked meat, and the incidence of Salmonella typhimurium infection ranks first among Salmonella infections, which ...

Claims

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Application Information

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IPC IPC(8): C12N1/20A61K39/112A61P31/04C12R1/42
CPCA61K39/0275A61K2039/522A61K2039/552C12N9/1092C12N9/88C12Y205/01019C12Y404/01021Y02A50/30
Inventor 于圣青王少辉杨登辉吴晓君丁铲
Owner SHANGHAI VETERINARY RES INST CHINESE ACAD OF AGRI SCI
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