The invention belongs to but not limited to the technical field of
genetic engineering technology, and discloses a construction method and application of a BspE
gene deletion strain of
Brucella effect
protein, the BspE
gene deletion strain is constructed by using
homologous recombination and SacB reverse screening technology,
Brucella abortus A19
vaccine strain liquid is used as a template, PCR amplification is carried out to obtain upstream and downstream homologous arms of BspE
gene, and the BspE
gene deletion strain is constructed by using the upstream and downstream homologous arms of the BspE gene. The method comprises the following steps: carrying out fusion PCR (
Polymerase Chain Reaction) amplification on upstream and downstream homologous arms of a BspE gene subjected to gel
recovery, and connecting and transforming an obtained fusion fragment and a pUC19-SacB
plasmid to obtain a recombinant
plasmid pUC19-
delta BspE. The invention relates to a bovine
brucella BspE deletion
vaccine strain, which is characterized in that a
plasmid is obtained, the plasmid is electrically transformed into a bovine
brucella A19
competent cell, and the positive transformant is a bovine
brucella BspE deletion strain through
kanamycin resistance and
sucrose pressure screening, PCR (
polymerase chain reaction) identification and sequencing, and the deletion
vaccine strain improves the levels of antibacterial cytokines and specific
brucella antibody IgG, and can be used for preparing the bovine brucella BspE deletion vaccine strain. The
Brucella attenuated vaccine strain is a novel Brucella candidate
attenuated vaccine strain and has a good market prospect.