DesUSP; Seed train + fermentation
2015-10-28 20:00:43 0 举报
DesUSP是一种用于制备高纯度、低内毒素的蛋白质的方法。该方法包括种子培养和发酵两个步骤。在种子培养阶段，菌种被接种到含有适当营养物质的培养基中，通过控制温度、pH值和氧气供应等条件，使菌种快速繁殖并产生大量的蛋白质。接下来是发酵阶段，将种子培养物接种到大规模的发酵罐中，继续提供适当的营养物质和环境条件，促进菌种的生长和蛋白质的合成。通过控制发酵过程的温度、pH值、搅拌速度等参数，可以优化蛋白质的产量和质量。最终，经过分离纯化步骤，可以获得高纯度的蛋白质产物。DesUSP方法具有操作简单、成本低、效率高等优点，广泛应用于生物医药、食品工业等领域。
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大纲/内容
FERMENTATION(cell growth and expression)
FILTRATION(Pre-capture step treatment)
CELL LYSIS & HEAT TREATMENT(Protein release)
CENTRIFUGATION(Cell debris removal)
SEED TRAIN 1+2(inoculation and expression)
CENTRIFUGATION(Cell harvest)
Choice of strain: soluble periplasmic expression Expression vector: lac Promotor Mode of cultivation: fed-batch Fermentation temperature: 37 oC Medium composition: filter-sterilised glucose Time and duration of induction: 3~4 days General production titre: 0.5 ~ 2.4 g/L Estimated production titre: 1.5 g/L Fed-batch cell concentration: ~130 g DCW/L
Production host: Genetic modified E.coli Production titre: 15 mg/L Doubling time time: 20~30 min Operation: Shaking flask inoculumn at 37ºC and 200 rpm for 13 hrs. Resistance/Selection Marker: • Bactericidal antibiotics kanamycin and tetracycline combination Ideal Expression System: • Single host cell for both polypeptides • Single expression plasmid • Soluble periplasmic expression Periplasmic expression: • Avoiding release of large quantities of DNA RNA and cytoplasmic proteins • Easy to concentrate via centrifugation
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