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A kind of small RNA relevant to the virulence of Brucella and its application in the preparation of attenuated Brucella

A Brucella, virulence technology, applied in the direction of DNA / RNA fragments, recombinant DNA technology, medical preparations containing active ingredients, etc., can solve the problem of less research

Active Publication Date: 2022-04-22
HARBIN VETERINARY RES INST CHINESE ACADEMY OF AGRI SCI
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

According to speculation, Brucella should theoretically have 100 to 200 sRNAs. Currently, only two sRNAs, AbsR1 and AbsR2, have been identified, and there are relatively few studies

Method used

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  • A kind of small RNA relevant to the virulence of Brucella and its application in the preparation of attenuated Brucella
  • A kind of small RNA relevant to the virulence of Brucella and its application in the preparation of attenuated Brucella
  • A kind of small RNA relevant to the virulence of Brucella and its application in the preparation of attenuated Brucella

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Embodiment 1

[0046] 1 Materials and methods

[0047] 1.1 Strains, plasmids and vectors

[0048] Brucella M28 is preserved by the biosafety level 3 laboratory of Harbin Veterinary Research Institute, Chinese Academy of Agricultural Sciences. Plasmids pSP72 and pBlue-Kan (recorded in "Construction of ERG6 gene deletion mutant strain of S. Period, Lei Jie et al., which are preserved and provided by the Harbin Veterinary Research Institute of the Chinese Academy of Agricultural Sciences.) are preserved by this laboratory. DH5α competent cells were purchased from Nanjing Nuoweizan. 6-7-week-old SPF grade BALB / c mice were purchased from Beijing Weitong Lihua Experimental Animal Technology Co., Ltd.

[0049] All experiments involving Brucella strains and their pathogenicity were performed in biosafety level 3 laboratories.

[0050] 1.2 Reagents

[0051] Brucella Selective Supplement was purchased from OXOID, and TSB and TSA were purchased from BD Company. Super-Fidelity DNA Polymerase, Mu...

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Abstract

The invention discloses a small RNA related to the virulence of Brucella and its application in preparing attenuated Brucella. The present invention uses Northern blot to prove the existence of sRNA Clu7 in the virulent Brucella strain M28, and uses the method of homologous recombination to construct the sRNA deletion strain M28ΔClu7, using mouse macrophage RAW264.7 and Balb / c mice as examples The model assessed the effect of this sRNA on M28 virulence. The results showed that the deletion of the sRNA could significantly down-regulate the ability of the virulent strain M28 to replicate and survive in the mouse macrophage RAW264.7 and mouse spleen. At the 5th week of inoculation, M28ΔClu7 was completely cleared from the mice, and the virulence was significantly reduced compared with M28. The mouse challenge protection experiment showed that the protective effect of immunizing M28ΔClu7 on mice against virulent M28 infection reached the protective power of vaccine strain M5‑90. The above results indicate that sRNA Clu7 has a huge decisive role in the virulence of Brucella. The proposal of the present invention has played a huge role in promoting the discovery of the virulence mechanism of Brucella. It provides new technical means for the research and development of new candidate vaccine strains of Brucella.

Description

technical field [0001] The invention relates to a small RNA related to the virulence of Brucella, and also relates to the application of the small RNA in preparing attenuated Brucella. The invention belongs to the field of biotechnology. Background technique [0002] Brucellosis (brucellosis), referred to as brucellosis, is a zoonotic chronic bacterial infectious disease caused by Brucella. It is widely distributed all over the world. my country is also a serious epidemic area of ​​brucellosis. The number of new cases of human per year ranks first in the world. Brucellosis has caused great damage to the health of our people and agriculture. People infected with Brucella show symptoms such as undulating fever, endocarditis, and arthritis, and animals infected by it can lead to miscarriage. Unlike enteric pathogens, Brucella virulence does not depend on the expression of specific virulence factors, but on their ability to replicate and survive in host cells. Animal brucello...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/113C12N1/21A61K39/10A61P31/04C12R1/01
CPCA61K39/098A61P31/04C12N15/113A61K2039/522C12N2310/10
Inventor 步志高胡森许达
Owner HARBIN VETERINARY RES INST CHINESE ACADEMY OF AGRI SCI
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