1986 Fiscal Year Final Research Report Summary
Highly Efficient Production of Recombinant Gene by Repeated Fed-Batch Culture
| Project/Area Number |
60470118
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| Research Category |
Grant-in-Aid for General Scientific Research (B)
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| Allocation Type | Single-year Grants |
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| Research Field |
反応工学
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| Research Institution | Nagoya University |
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Principal Investigator |
KOBAYASHI Takeshi Nagoya University, 工学部, 教授 (10043324)
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| Co-Investigator(Kenkyū-buntansha) |
IIJIMA Shinji Nagoya University, 工学部, 助教授 (00168056)
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| Project Period (FY) |
1985 – 1986
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| Keywords | Copy number / Transcription efficiency / Plasmid / Stability / Nutrient concentration / Fed-batch culture / Tryptophan promoter / グルコースセンサー |
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| Research Abstract |
For overproduction of useful proteins, control of gene-copy number, stability of plasmid, control of transcription efficiency and control of nutrients are important in fed-batch culture of gene-engineered microorganisms. The recombinant E. coli harboring a runaway-replication plasmid was cultivated at 25゜C and then the temperature was shifted to 37゜C. The copy number increased drastically and the gene product was amplified almost 40-fold.
Effects of nutrients on <alpha> -amylase production and plasmid stability were studied in fed-batch culture of recombinant E. coli. Plasmid was maintained stably in a semi-synthetic medium but plasmid-free segregants were observed in a synthetic medium.
To control transcriptional efficiency, fundamental properties of the trp promoter were also investigated in fed-batch culture using a recombinant containing the lacZ gene controlled by this promoter. On-off regulation of the promoter was achieved by controlling the tryptophan level.
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