| 重组大肠杆菌表达降氰酶的诱导条件优化 |
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| 引用本文： | 陈焕基,李青云,尹业兴,刘幽燕,甘雨.重组大肠杆菌表达降氰酶的诱导条件优化[J].广西农业生物科学,2014(4):737-743. |
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| 作者姓名： | 陈焕基 李青云 尹业兴 刘幽燕 甘雨 |
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| 作者单位： | 广西大学化学化工学院,南宁530004 |
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| 基金项目： | 国家自然科学基金（51108098）; 广西大学大学生实验技能和科技创新能力训练基金资助项目（SYJN20120321）共同资助 |
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| 摘 要： | 采用积分光密度法对产碱杆菌（Alcaligenes sp.）降氰酶基因在大肠杆菌中的高效表达进行了诱导条件优化,以工程菌BL21（DE3）-pET28a-cdE的诱导表达条件进行优化,以提高目的蛋白的产量和活性。结果显示,重组菌表达降氰酶的最佳诱导条件为温度28℃,初始菌体浓度OD600值为0.6,IPTG浓度为0.8mmol/L,诱导时间12h。研究发现,诱导剂的添加浓度对外源蛋白的表达水平具有显著影响,使用终浓度为0.8mmol/L的IPTG,诱导表达12h,降氰酶蛋白收率达到143.27mg/L,比活力达到11560 U/mg。与不添加诱导剂相比,重组蛋白的收率和比活力分别提高了64倍和9倍。
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| 关 键 词： | 降氰酶 产碱杆菌 诱导表达 积分光密度法 重组大肠杆菌 |
Optimization of Induction Conditions of Recombinant Cyanide-degrading Enzyme Expressed in E. coli |
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| Chen Huanji,Li Qingyun,Yin Yexing,Liu Youyan,Gan Yu.Optimization of Induction Conditions of Recombinant Cyanide-degrading Enzyme Expressed in E. coli[J].Journal of Guangxi Agricultural and Biological Science,2014(4):737-743. |
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| Authors: | Chen Huanji Li Qingyun Yin Yexing Liu Youyan Gan Yu |
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| Institution: | (College of Chemistry and Chemical Engineering, Guangxi University, Nanning, 530004) |
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| Abstract: | Cyanide-degrading enzyme from Alcaligenes sp. expressed in E. coli were optimized by using integral optical density method to increase the production and activity of the recombinant protein. The results showed that the cyanide-degrading enzyme could be expressed optimally by adding 0.8 mmol/L IPTG at an initial OD600 of 0.6and induced for 12 hours at 28℃. Further study found that the concentration of IPTG had a significant impact on the expression level of recombinant cyanide-degrading enzyme. The yield of recombinant protein and its specific activity were 143.27 mg/L and 11 560 U/mg after induced by 0.8 mmol/L IPTG for 12 hours, which were increased by64 times and 9 times than those without IPTG, respectively. |
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| Keywords: | Cyanide-degrading enzyme Alcaligenes sp Inducible expression Integral optical density method Recombinant E coli |
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