Statistical optimization of medium composition and culture condition for the production of recombinant anti-lipopolysaccharide factor of Eriocheir sinensis in Escherichia coli
文献类型：期刊论文
| 作者 | Jiang Shan1,2; Liu Mei1; Wang Baojie1; Jiang Keyong1; Wang Lei1 |
| 刊名 | CHINESE JOURNAL OF OCEANOLOGY AND LIMNOLOGY
 |
| 出版日期 | 2011-11-01 |
| 卷号 | 29期号:6页码:1249-1259 |
| 关键词 | Anti-lipopolysaccharide Factor Optimization Medium Composition Culture Conditions Statistical Methodology |
| ISSN号 | 0254-4059 |
| DOI | 10.1007/s00343-011-0282-5 |
| 文献子类 | Article |
| 英文摘要 | Anti-lipopolysaccharide factors (ALFs) are important antimicrobial peptides that are isolated from some aquatic species. In a previous study, we isolated ALF genes from Chinese mitten crab, Eriocheir sinensis. In this study, we optimized the production of a recombinant ALF by expressing E. sinensis ALF genes in Escherichia coli maintained in shake-flasks. In particular, we focused on optimization of both the medium composition and the culture condition. Various medium components were analyzed by the Plackett-Burman design, and two significant screened factors, (NH(4))(2)SO(4) and KH(2)PO(4), were further optimized via the central composite design (CCD). Based on the CCD analysis, we investigated the induction start-up time, the isopropylthio-D-galactoside (IPTG) concentration, the post-induction time, and the temperature by response surface methodology. We found that the highest level of ALF fusion protein was achieved in the medium containing 1.89 g/L (NH(4))(2)SO(4) and 3.18 g/L KH(2)PO(4), with a cell optical density of 0.8 at 600 nm before induction, an IPTG concentration of 0.5 mmol/L, a post-induction temperature of 32.7 degrees C, and a post-induction time of 4 h. Applying the whole optimization strategy using all optimal factors improved the target protein content from 6.1% (without optimization) to 13.2%. We further applied the optimized medium and conditions in high cell density cultivation, and determined that the soluble target protein constituted 10.5% of the total protein. Our identification of the economic medium composition, optimal culture conditions, and details of the fermentation process should facilitate the potential application of ALF for further research.; Anti-lipopolysaccharide factors (ALFs) are important antimicrobial peptides that are isolated from some aquatic species. In a previous study, we isolated ALF genes from Chinese mitten crab, Eriocheir sinensis. In this study, we optimized the production of a recombinant ALF by expressing E. sinensis ALF genes in Escherichia coli maintained in shake-flasks. In particular, we focused on optimization of both the medium composition and the culture condition. Various medium components were analyzed by the Plackett-Burman design, and two significant screened factors, (NH(4))(2)SO(4) and KH(2)PO(4), were further optimized via the central composite design (CCD). Based on the CCD analysis, we investigated the induction start-up time, the isopropylthio-D-galactoside (IPTG) concentration, the post-induction time, and the temperature by response surface methodology. We found that the highest level of ALF fusion protein was achieved in the medium containing 1.89 g/L (NH(4))(2)SO(4) and 3.18 g/L KH(2)PO(4), with a cell optical density of 0.8 at 600 nm before induction, an IPTG concentration of 0.5 mmol/L, a post-induction temperature of 32.7 degrees C, and a post-induction time of 4 h. Applying the whole optimization strategy using all optimal factors improved the target protein content from 6.1% (without optimization) to 13.2%. We further applied the optimized medium and conditions in high cell density cultivation, and determined that the soluble target protein constituted 10.5% of the total protein. Our identification of the economic medium composition, optimal culture conditions, and details of the fermentation process should facilitate the potential application of ALF for further research. |
| 学科主题 | Marine & Freshwater Biology ; Oceanography |
| URL标识 | 查看原文 |
| 语种 | 英语 |
| WOS记录号 | WOS:000299039400012 |
| 公开日期 | 2012-07-03 |
| 源URL | [http://ir.qdio.ac.cn/handle/337002/12040]  |
| 专题 | 海洋研究所_海洋生物技术研发中心 |
| 作者单位 | 1.Chinese Acad Sci, R&D Ctr Marine Biotechnol, Inst Oceanol, Qingdao 266071, Peoples R China 2.Chinese Acad Sci, Grad Univ, Beijing 100049, Peoples R China |
| 推荐引用方式 GB/T 7714 | Jiang Shan,Liu Mei,Wang Baojie,et al. Statistical optimization of medium composition and culture condition for the production of recombinant anti-lipopolysaccharide factor of Eriocheir sinensis in Escherichia coli[J]. CHINESE JOURNAL OF OCEANOLOGY AND LIMNOLOGY,2011,29(6):1249-1259. |
| APA | Jiang Shan,Liu Mei,Wang Baojie,Jiang Keyong,&Wang Lei.(2011).Statistical optimization of medium composition and culture condition for the production of recombinant anti-lipopolysaccharide factor of Eriocheir sinensis in Escherichia coli.CHINESE JOURNAL OF OCEANOLOGY AND LIMNOLOGY,29(6),1249-1259. |
| MLA | Jiang Shan,et al."Statistical optimization of medium composition and culture condition for the production of recombinant anti-lipopolysaccharide factor of Eriocheir sinensis in Escherichia coli".CHINESE JOURNAL OF OCEANOLOGY AND LIMNOLOGY 29.6(2011):1249-1259. |
入库方式： OAI收割
来源：海洋研究所
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