| 用毕赤酵母的GAP启动子调控表达L-阿拉伯糖异构酶 |
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| 引用本文： | 符仙,陈丽萍,张爱联,崔艳艳.用毕赤酵母的GAP启动子调控表达L-阿拉伯糖异构酶[J].工业微生物,2014,44(1):51-54. |
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| 作者姓名： | 符仙 陈丽萍 张爱联 崔艳艳 |
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| 作者单位： | 符仙 (海南大学农学院,海口市,570228)； 陈丽萍(中国热带农业科学院热带生物技术研究所,农业部热带作物生物学与遗传资源利用重点实验室,海口市,571101)；张爱联(中国热带农业科学院热带生物技术研究所,农业部热带作物生物学与遗传资源利用重点实验室,海口市,571101)；崔艳艳(中国热带农业科学院热带生物技术研究所,农业部热带作物生物学与遗传资源利用重点实验室,海口市,571101)； |
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| 基金项目： | 中央级公益性科研院所基本科研业务费专项资金资助项目(项目编号：ITBB120503) |
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| 摘 要： | 应用PCR从大肠杆菌基因组中扩增L-阿拉伯糖异构酶基因,用EcoR I和Not I双酶切将其克隆进P.pastoris表达载体,获得重组表达载体pGAP9K-L-ai。通过电转法将pGAP9K—L-ai转化毕赤酵母GS115,筛选高G418抗性和高表达L-阿拉伯糖异构酶的重组工程菌。用葡萄糖作为碳源在摇瓶中发酵48 h,表达重组L-ai 53 mg/L。用毕赤酵母的GAP启动子调控表达的重组L-ai具有异构D-半乳糖生成D-塔格糖的生物学活性。
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| 关 键 词： | L-阿拉伯糖异构酶 毕赤酵母 pGAP 基因表达 生物活性 |
Expression of L-arabinose isomerase in P. pastoris by using its GAP promoter |
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| FU Xian,CHEN Li-ping,ZHANG Ai-lian,CUI Yan-yan.Expression of L-arabinose isomerase in P. pastoris by using its GAP promoter[J].Industrial Microbiology,2014,44(1):51-54. |
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| Authors: | FU Xian CHEN Li-ping ZHANG Ai-lian CUI Yan-yan |
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| Institution: | College of Life Science and Agriculture, Hainan University, Haikou 570228 ; 2. Key Laboratory of Biology and Genetic Resources of Tropical Crop, Institute of Tropical Bioscienee and Bioteehnology, Chinese Academy of Tropical Agricultural Sciences, Haikou 571101, China |
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| Abstract: | L-arabinose isomerase gene was amplified from E. coli by PCR technique according to the L-ai sequence and cloned directly into plasmid pGAP9K to obtain P. pastoris expression vector pGAP-L-ai which was then transformed into the P. pastoris GS115 by electroporation method. A high gene copied and high L-ai expressed strain was used as engineer- ing strain and selected by replicating the transformants on G418 containing plates and expressing them. After two days fer- mentation in shaking flask using glucose as carbon source, 53 mg/L of recombinant L-ai was secreted in the medium. The recombinant L-arabinose isomerase showed its biological activity in changing D-galactose to D-tagatose. |
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| Keywords: | L-arabinose isomerase Pichia pastoris pGAP gene expression biological activity |
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