[1]张锋,王期,李桂兰,等.溶氧反馈分批补料高密度培养枯草杆菌谷氨酰胺合成酶工程菌[J].南京师大学报(自然科学版),2010,33(02):96-103.
 Zhang Feng,Wang Qi,Li Guilan,et al.High Density Fed-Batch Culture of Escherichia coli BL21/pET28b-glnA With DO Feed-back Control of Nutrient Feeding[J].Journal of Nanjing Normal University(Natural Science Edition),2010,33(02):96-103.
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溶氧反馈分批补料高密度培养枯草杆菌谷氨酰胺合成酶工程菌()
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《南京师大学报(自然科学版)》[ISSN:1001-4616/CN:32-1239/N]

卷:
第33卷
期数:
2010年02期
页码:
96-103
栏目:
生命科学
出版日期:
2010-06-20

文章信息/Info

Title:
High Density Fed-Batch Culture of Escherichia coli BL21/pET28b-glnA With DO Feed-back Control of Nutrient Feeding
作者:
张锋;王期;李桂兰;徐健红;洪素丽;岳玲;冯志勇;殷志敏;
南京师范大学生命科学学院, 江苏南京210046
Author(s):
Zhang FengWang QiLi GuilanXu JianhongHong SuliYue LingFeng ZhiyongYin Zhimin
School of Life Science,Nanjing Normal University,Nanjing 210046,China
关键词:
谷氨酰胺合成酶 重组大肠杆菌 分批补料培养 高密度培养
Keywords:
G lutam ine synthetase recomb inan tE. co li fed -batch cu lture h igh cell dens ity cu lture
分类号:
TQ925
摘要:
为了建立枯草杆菌谷氨酰胺合成酶基因重组工程菌BL21/pET28b-glnA的高密度培养工艺,首先以摇瓶培养为基础,对影响工程菌生长及目的蛋白表达的因素如培养基、pH值、诱导剂浓度及时间等进行优化,再扩大至B IOTECH-5BG(5 L)自动玻璃发酵罐培养,利用溶氧反馈补料策略进行分批补料,并在培养过程中保持20%~50%的溶解氧,7.0~7.5的pH,以及1.0 g/L乳糖诱导12 h,成功地进行了工程菌的高密度培养,最终细菌干重达到48.1 g/L,目的蛋白的表达量占菌体蛋白总量的55%,粗提物酶比活为10.35 U/mg,整个补料过程细菌比生长率稳定的控制在0.1 h-1左右.
Abstract:
To investigate the optim a l h igh cell density culture conditions of recomb inant BL21 /pET28b-g lnA to produce recom b inant G lutam ine Synthetase ( GS) of Bac illus Subtilis, based on the exper im enta l resu lts obtained in shake flask cu ltivation, the effects of the composition o f the cu lturem edium, the range o f pH, induce r concen tration and induced tim e on the g row th o f bac teria and the expression leve l o f GS we re ana lyzed. Then the bacter ia we re transferred to B IOTECH- 5BG( 5 L) DO feedback fed ba tch cu lture system. By keep ing dissolved oxygen at 20% ~ 50%, pH at 7.0~ 7.5, cultu ring in Optim izedM9M edium and induc ing for 12 h in the presence o f 1 g /L lactose at 37℃ during fedbatch cu ltiva tion, we perform ed a three tim e repeated culture. Ce ll density of recomb inan t E. co li w as reached to 48.1 g /L under constant ( 0.10 h - 1 ) specific g row th rates. The express ion leve l o f recom binant prote in GS w as over 55% o f the to tal pro te in in E. coli. Specific ac tiv ity o f crude ex trac t enzyme w as 10.35 U /mg. A h igh expression and stab ilized cu lture process o f recomb inan t Glutam ine Synthe tase of Bac illus Subtilis has been established, wh ich founds the basis for further larg e sca le produc tion o f G lutam ine Synthe tase.

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备注/Memo

备注/Memo:
通讯联系人: 殷志敏, 博士, 教授, 研究方向: 生物化学及细胞生物学. E-mail:yinzhimin@ njnu. edu. cn
更新日期/Last Update: 2013-04-08