Protective antigen protein of porcine toxigenic pasteurella multocida as well as application and vaccine of protective antigen protein
A technology of protective antigen and Pasteurella, which is applied in the field of immunology, can solve the problems of low production capacity, unfavorable scale-up production, unsatisfactory protein expression, and increased production cost of the production line, so as to increase production practicability and good immunity Originality, conducive to the effect of production amplification
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Embodiment 1
[0032] Construction of embodiment 1 recombinant strain
[0033] 1.1 Construction of expression engineering bacteria
[0034] According to literature research: the full length of PMT is toxic, and PMT-N is more immunogenic than the C-terminal of PMT. Afterwards, we constructed 3 strains expressing the full-length of PMT, PMT-N (PMT N-terminal 1~ 520 fragment), PMT-C protein (PMT 520-1295 fragment), PET-PMT full-length (BL21), PET-PMT-N (BL21), PET-PMT-C (BL21) expression engineering bacteria .
[0035] 1.2 Full-length PMT, PMT-N, PMT-C protein expression, verification and purification quantification
[0036]Express engineering bacteria PET-PMT full-length (BL21), PET-PMT-N (BL21), PET-PMT-C (BL21) were streaked on LB (corresponding to resistance) solid plates and cultured at 37°C for 16 hours, and single The colonies were inoculated in 5ml LB liquid medium containing 100μg / ml corresponding resistance, cultured on a shaker (150r / min) at 37°C for 16h, and then inoculated at 1%...
Embodiment 2
[0038] Example 2. Vaccine Preparation
[0039] 2.1 Preparation of PMT full-length vaccine, PMT-N vaccine, PMT-C vaccine, PMT-N+PMT-C vaccine
[0040] Take a certain amount of PMT full-length protein (1), PMT-N protein (2), PMT-C protein (3) and PMT-N+PMT-C protein (4) prepared in Example 1 and add a certain amount of Bowers' protein Whole bacteria inactivated liquid, add a certain proportion of water-based adjuvant, stir evenly with a magnetic stirrer, and then sub-package. After passing the inspection, it is the porcine atrophic rhinitis subunit vaccine, wherein the antigen content in the vaccine is 50-200 μg / ml, The ratio of antigen and adjuvant in the vaccine is as follows:
[0041] Table 1 Composition of vaccines 1, 2, 3, 4
[0042]
[0043]
[0044] Note: "-" means no addition.
Embodiment 3
[0045] Embodiment 3. Vaccine sterility test
[0046] The sterility test was performed on the above three subunit vaccines in accordance with the appendix of the current "Chinese Veterinary Pharmacopoeia". The results were sterile growth.
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