研究生: |
英格麗 Magdiel - Inggrid Setyawati |
---|---|
論文名稱: |
Genetically Engineered Acetobacter xylinum: Bacterial Cellulose and Self Immobilization System Production Genetically Engineered Acetobacter xylinum: Bacterial Cellulose and Self Immobilization System Production |
指導教授: |
李振綱
Cheng-Kang Lee |
口試委員: |
王孟菊
Meng-Jiy Wang 劉懷勝 Hwai-Shen Liu |
學位類別: |
碩士 Master |
系所名稱: |
工程學院 - 化學工程系 Department of Chemical Engineering |
論文出版年: | 2007 |
畢業學年度: | 95 |
語文別: | 英文 |
論文頁數: | 140 |
中文關鍵詞: | Bacterial cellulose 、DAAO 、Acetobacter xylinum 、immobilization |
外文關鍵詞: | Bacterial cellulose, DAAO, Acetobacter xylinum, immobilization |
相關次數: | 點閱:169 下載:1 |
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Gram-negative, rod-shaped, obligate aerobic bacterium, Acetobacter xylinum is the most productive bacterial cellulose (BC) producer. In this study the A. xylinum host strain were genetically engineered for further enhancement of bacterial cellulose production and generate self immobilized cell system.
Vitreoscilla hemoglobin was constitutively expressed in Acetobacter xylinum to enhance bacterial cellulose (BC) production. The enhancement was much pronounced in static culture than in shaken culture. Reducing O2 tension in gaseous phase of the culture by tightly sealing the culture tube could also enhance BC production by 70 %. O2 tension reduced from 21 % to 15 % in the sealed and static culture of VHb-expressing A. xylinum after 7 days cultivation, while 7.36 g/l of BC with yield of 0.44 were obtained. BC pellicle production was successfully scaled-up in a sealed 4 l disposable zip lock plastic bag with BC yield of 0.35 and concentration of 6.31 g/l.
Along with the production of BC, the cell became entrapped inside the BC fibers produced. In order to take the advantage of these cells’ self-immobilization phenomenon for biotransformation, D-amino acid oxidase (DAAO) was chosen to be expressed in A. xylinum along with the BC production. A plasmid bearing the Rhodosporidium toruloides daao gene, placP-DAAO-122 was constructed and transformed into A. xylinum to enable the IPTG-induced A. xylinum’s DAAO production. The highest DAAO activity (~1.5 U/ml) was detected when 4% glycerol was supplemented in HS medium (HSG4). The detected DAAO activity was in the order of free enzyme > free cell > immobilized cell. Cell cultured in shaking condition produce more DAAO than in stationary condition. The operational stability of DAAO activity was in the order of free enzyme < free cell < immobilized cell.
Gram-negative, rod-shaped, obligate aerobic bacterium, Acetobacter xylinum is the most productive bacterial cellulose (BC) producer. In this study the A. xylinum host strain were genetically engineered for further enhancement of bacterial cellulose production and generate self immobilized cell system.
Vitreoscilla hemoglobin was constitutively expressed in Acetobacter xylinum to enhance bacterial cellulose (BC) production. The enhancement was much pronounced in static culture than in shaken culture. Reducing O2 tension in gaseous phase of the culture by tightly sealing the culture tube could also enhance BC production by 70 %. O2 tension reduced from 21 % to 15 % in the sealed and static culture of VHb-expressing A. xylinum after 7 days cultivation, while 7.36 g/l of BC with yield of 0.44 were obtained. BC pellicle production was successfully scaled-up in a sealed 4 l disposable zip lock plastic bag with BC yield of 0.35 and concentration of 6.31 g/l.
Along with the production of BC, the cell became entrapped inside the BC fibers produced. In order to take the advantage of these cells’ self-immobilization phenomenon for biotransformation, D-amino acid oxidase (DAAO) was chosen to be expressed in A. xylinum along with the BC production. A plasmid bearing the Rhodosporidium toruloides daao gene, placP-DAAO-122 was constructed and transformed into A. xylinum to enable the IPTG-induced A. xylinum’s DAAO production. The highest DAAO activity (~1.5 U/ml) was detected when 4% glycerol was supplemented in HS medium (HSG4). The detected DAAO activity was in the order of free enzyme > free cell > immobilized cell. Cell cultured in shaking condition produce more DAAO than in stationary condition. The operational stability of DAAO activity was in the order of free enzyme < free cell < immobilized cell.
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