Avibacterium paragallinarum antigen protein, vaccine composition containing Avibacterium paragallinarum antigen, and preparation method and application of composition
A vaccine composition and antigen protein technology, applied in the field of Avian Bacillus paragallina antigen protein, can solve the problems that monovalent vaccine or bivalent vaccine cannot produce protective effect, and the protective effect is not very ideal, and achieve good immunogenicity and broad spectrum. The effect of immunogenicity
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Embodiment 1
[0061] Embodiment 1 Construction of Avibacterium paragallinarum HMTp210 2 District 2 Gene Recombination Plasmid
[0062] 1.1 Primer design
[0063] According to the HMTp210 region 2 and region 2 gene sequences of Apg types A, B and C, a pair of universal primers were designed using Primer5.0 software, and the 5' ends of the upstream and downstream primers were respectively introduced with BamH I, Hind III restriction sites and protective bases. The base was used to amplify the full-length sequence of the HMTp210 region 2 gene. The sequences of the primers are shown in Table 1, and the underlined part is the introduced restriction site. The primers were synthesized by Jinweizhi Company.
[0064] Table 1 Primer Sequence
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[0066] 1.2 Extraction of Genomic DNA of Avibacterium paragallinarum
[0067] Apg type A HN3 strain, B type HN5 strain and C type SD3 strain were streaked and inoculated on the TSA plate, at 37°C, 5% CO 2 Cultivate in an incubator for 24-36 hours,...
Embodiment 2
[0075]Example 2 Construction of recombinant expression strains BL21-A, BL21-B and BL21-C
[0076] 2.1 Transformation of E.coli BL-21 Competent Cells with Recombinant Plasmid and Expression of Recombinant Protein
[0077] In Example 1, the recombinant plasmid with the correct sequence identified by sequencing was transformed into E. coli BL-21 competent cells, and the recombinant expression strains BL21-A, BL21-B, and BL21-C were respectively obtained. At the same time, the empty plasmid pET-32a was treated with the same Methods Transformed E.coli BL-21 competent cells, coated with ampicillin-containing LB solid medium, cultured at 37°C for 16 hours, picked well-growing single colonies and inoculated them in 10ml of ampicillin-containing LB liquid medium, 37°C, Shake culture at 200rpm for 2h. OD of bacteria solution 600 When the value reaches 0.6-0.8, take 1ml of uninduced expression bacteria solution and empty plasmid bacteria solution as samples before induction, add IPTG w...
Embodiment 3
[0086] The preparation of embodiment 3 chicken infectious rhinitis vaccines
[0087] 3.1 Preparation of recombinant chicken infectious rhinitis subunit vaccine
[0088] Combine p-A, p-B, and p-C recombinant proteins according to a certain ratio, add Tween-80 to prepare a water phase, and then mix and emulsify the water phase with imported white oil at a volume of 1:2 to make chicken containing three recombinant protein components with different concentrations For the infectious rhinitis subunit vaccine, the concentrations of the three recombinant protein components in each vaccine are shown in Table 2.
[0089] Table 2 Protein content of different vaccines
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