Production method of porcine foot-and-mouth disease type O genetically engineered composite epitope protein vaccine
A genetic engineering, swine foot-and-mouth disease technology, applied in the field of immunization, can solve the problems of small unit output, high production cost, unsuitable for industrial production, etc., and achieve the effects of short production cycle, fewer personnel, and less reagent consumption.
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[0032] Example 1
[0033] Engineering bacteria culture and induced expression:
[0034] Take 500 mL of the overnight cultured engineering bacteria seed solution and inoculate it in 50 L of LB culture solution (containing 75 μg / mL kanamycin), ferment at 37° C., pH 7.2, dissolved oxygen 40%, and add it by exponential flow. After 5 hours of fermentation, IPTG was added to a final concentration of 1.0 mmol / L, and expression was induced for 5 hours to obtain the recombinant protein after induced expression.
[0035] Bacteria collection and fragmentation:
[0036] The collected fermentation broth was concentrated using a 750KD hollow fiber column, and stopped when it reached 5L. Using a high pressure homogenizer, the concentrate was crushed under the conditions of 900 bar and two cycles.
[0037] Inclusion body washing and denaturation solubilization:
[0038] The broken fermentation broth was washed with a 750KD hollow fiber column, and the buffer was 1×IBWashBuffer (EDTA 10mmo...
Example Embodiment
[0043] Embodiment 2
[0044] Engineering bacteria culture and induced expression:
[0045] Take 400 mL of the overnight cultured engineering bacteria seed solution and inoculate it into 50 L of LB culture solution (containing 75 μg / mL kanamycin), ferment at 37° C., pH 7.2, and dissolved oxygen 40%, and add it by exponential flow. After 5 hours of fermentation, IPTG was added to a final concentration of 1.0 mmol / L, and expression was induced for 5 hours to obtain the recombinant protein after induced expression.
[0046] Bacteria collection and fragmentation:
[0047] The collected fermentation broth was concentrated using a 750KD hollow fiber column, and stopped when it reached 5L. Using a high pressure homogenizer, the concentrate was crushed under the conditions of 900 bar and two cycles.
[0048] Inclusion body washing and denaturation solubilization:
[0049] The broken fermentation broth was washed with a 750KD hollow fiber column, and the buffer was 1×IBWashBuffer (E...
Example Embodiment
[0054] Embodiment 3
[0055] Engineering bacteria culture and induced expression:
[0056] Take 300 mL of the overnight cultured engineering bacteria seed solution and inoculate it in 50 L of LB culture solution (containing 75 μg / mL kanamycin), ferment at 37° C., pH 7.2, and dissolved oxygen 40%, and add by exponential flow After 5 hours of fermentation, IPTG was added to a final concentration of 1.0 mmol / L, and expression was induced for 5 hours to obtain the recombinant protein after induced expression.
[0057] Bacteria collection and fragmentation:
[0058] The collected fermentation broth was concentrated using a 750KD hollow fiber column, and stopped when it reached 5L. Using a high pressure homogenizer, the concentrate was crushed under the conditions of 900 bar and two cycles.
[0059] Inclusion body washing and denaturation solubilization:
[0060] The broken fermentation broth was washed with a 750KD hollow fiber column, and the buffer was 1×IBWashBuffer (EDTA 10...
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