Brucella melitensis recombination strain M5-delta bp26-delta znuA as well as preparation method and application thereof

Abstract

The invention discloses a brucella melitensis recombination strain M5-delta bp26-delta znuA as well as a preparation method and application thereof. The preparation method comprises the steps: with a brucella melitensis M5 genome as a template, carrying out PCR (Polymerase Chain Reaction) to obtain upstream and downstream homologous arms of a znuA gene, and connecting the upstream and downstream homologous arms to obtain a fragment delta znuA; cloning the fragment delta znuA to a carrier pRE112 to obtain a recombinant plasmid pRE-delta znuA; inducing the plasmid into a brucella melitensis M5-delta bp26 cell, and respectively screening through chloramphenicol and saccharose selective mediums to obtain double homologous recombinant recons, namely the recombination strain M5-delta bp26-delta znuA. Compared with an M5 parent strain, the strain is consistent in growth characteristic, stable in heredity, not easy to cause throwback, low in toxicity, favorable in immune effect retention and capable of being applied to the distinguishing and detection of natural infection or artificial immunity because of bp26 gene deletion markers. Therefore, the strain can be used as a marker vaccine with a better effect to be applied clinically.

Description

technical field [0001] The invention belongs to the field of recombinant bacterial vaccines, and in particular relates to a Brucella melitensis recombinant strain M5-Δbp26-ΔznuA, its preparation method and application. The strain is obtained by deleting the znuA gene on the basis of the deletion of the bp26 gene of the sheep type 5 attenuated strain (M5). Background technique [0002] Brucellosis is a zoonotic infectious disease caused by Brucella and characterized by abortion and fever. Due to the characteristics of intracellular parasitism of Brucella, the prevention and control of this disease has always been based on attenuated vaccine immunization. At present, the attenuated strains used for the prevention and control of brucellosis mainly include foreign bovine type 19 (S19) attenuated strain and sheep type ReV. No. 2 (S2) attenuated strain. These classic attenuated vaccine strains have played an important role in the prevention and control of the spread and spread o...

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Problems solved by technology

However, due to the very small amount of free zinc ions in the host, in order to absorb enough zinc to ensure the metabolism of the bacteria themselves and proliferate in the host, various bacteria such as Brucella have evolved several types of proteins to participate in the uptake and transport of zinc. Ions, these protein manipulation systems for transporting zinc ions are named ZnuABC, this system includes a variety of proteins encoded by znuA, znuB, and znuC genes, among which ZnuA is a periplasmic-solute binding protein, and the mechanism of this gene is currently being studied Still less, it is only speculated that the ZnuA protein is related to Brucella virulence factors

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