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昆虫细胞-杆状病毒培养体系的研究
常韶华
Subtype博士
Thesis Advisor李佐虎
1998-03-01
Degree Grantor中国科学院研究生院
Abstract本文从生化工程的角度出发,以提高产率、降低生产成本为目标,系统研究了昆虫细胞-杆状病毒培养体系。论文验证了昆虫细胞生长过程中内源性生长因子的存在,并采用低血清驯化和补加磷酸胰蛋白胨肉汤(TPB)相结合的方法,使培养成本降低50%。发现采用流加TPB可明显提高细胞密度和产物表达水平。感染指数和感染时间共同影响病毒在细胞内的复制水平,改进营养水平有利于提高产物表达水平。创造性地将周期温度变化操作引入培养体系,使体系效率提高30%以上。还首次建立了统一描述细胞生长全过程动力学的唯象模型,细胞培养过程与病毒感染过程均与模型吻合较好。这些研究结果可为进一步深入研究和大规模生产提供基础数据和科学依据。
Other AbstractInsect cell culture was used to produce wild baculovirus and pro-uerokinase (pro-UK) in this study. The main research was divided into 5 parts: 1 Insect cell growth was studied for serum was verified through the test of inoculum effect and conditional medium. Sf9 cells was adapted in 2% serum medium. Trypotse phosphate broth (TPB) was added as a serum substitute. Compared to 10% serum medium, adapted cells obtained similar growth in 2% serum medium with 2% TPB supplemented, while the culture cost reduced 50%. 2 Sf9 cell can grow in various culture vessels. 50-100 rpm was suitable for suspension culture. For batch culture, the maximum cell density was 18.6 * 10~5/ml in spinner flask, 17.5 * 10~5/ml in 2-L agitator bioreactor, 23.5 * 10~5/ml in air-liquid-lift bioreactor. Cells raised to 23.8 * 10~5/ml in half medium replacement and to 25.4 * 10~5/ml in TPB feeding culture. 3 Effects of multiplicity of infection (MOI) was dependent on time of infection. Low MOI required early infection and long culture time, while high MOI was reverse. Pro-UK yields were 420 IU/ml in spinner flask and 530 IU/ml in 2-L agitator bioreactor with MOI=3. Medium replacement and fed-batch can enhanced baculovirus and pro-UK yields. The pro-UK yield reached 820 IU/ml in 1% serum medium with 2% TPB supplemented. 4 Temperature oscillation was introduced to insect cell-baculovirus culture system. The optimum temperature oscillation in 12 hr for baculovirus propagation was 24-28 ℃. Compared to 28 ℃ culture, the baculovirus yields raised about 30%. 5 Phenomenological model of cell growth was established on experimental work. This model could describe the whole process of insect cell baculovirus culture system and coincide well with the experimental result.
Pages111
Language中文
Document Type学位论文
Identifierhttp://ir.ipe.ac.cn/handle/122111/8335
Collection研究所(批量导入)
Recommended Citation
GB/T 7714
常韶华. 昆虫细胞-杆状病毒培养体系的研究[D]. 中国科学院研究生院,1998.
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