霍乱弧菌宿主整合因子亚单位的可溶性克隆表达

李晶 王洪霞 阚飙 梁未丽

李晶, 王洪霞, 阚飙, 梁未丽. 霍乱弧菌宿主整合因子亚单位的可溶性克隆表达[J]. 疾病监测, 2016, 31(4): 278-281. doi: 10.3784/j.issn.1003-9961.2016.04.005
引用本文: 李晶, 王洪霞, 阚飙, 梁未丽. 霍乱弧菌宿主整合因子亚单位的可溶性克隆表达[J]. 疾病监测, 2016, 31(4): 278-281. doi: 10.3784/j.issn.1003-9961.2016.04.005
LI Jing, WANG Hong-xia, KAN Biao, LIANG Wei-li. Cloning and expression of soluble integration host factor subunit of Vibrio cholerae[J]. Disease Surveillance, 2016, 31(4): 278-281. doi: 10.3784/j.issn.1003-9961.2016.04.005
Citation: LI Jing, WANG Hong-xia, KAN Biao, LIANG Wei-li. Cloning and expression of soluble integration host factor subunit of Vibrio cholerae[J]. Disease Surveillance, 2016, 31(4): 278-281. doi: 10.3784/j.issn.1003-9961.2016.04.005

霍乱弧菌宿主整合因子亚单位的可溶性克隆表达

doi: 10.3784/j.issn.1003-9961.2016.04.005
基金项目: 

国家自然科学基金(No.81171640)

北京市科学技术委员会项目(No.D131100005313016)

Cloning and expression of soluble integration host factor subunit of Vibrio cholerae

Funds: 

This study was supported by the fund for the National Natural Science Foundation of China (No. 81171640) and the project from Beijing Municipal Science Technology Commission (No.D131100005313016)

  • 摘要: 目的 克隆表达霍乱弧菌全局性调节子宿主整合因子(integration host factor, IHF)亚单位基因ihfB。方法 以C7258染色体为模板, 聚合酶链反应(polymerase chain reaction, PCR)扩增ihfB基因, 扩增产物经NheⅠ和SapⅠ酶切后克隆入表达载体pXTB1;将ihfB的N端和亚单位基因ihfA的C端通过搭桥PCR重组, 重组片段N-ihfB-C-ihfA经NheⅠ和SapⅠ双酶切后克隆入表达载体pXTB1。重组质粒导入大肠埃希菌ER2566, 0.4 mmol/L IPTG诱导表达, 超声裂解, 上清经CBD柱吸附纯化, 二硫苏糖醇(DTT)柱上裂解洗脱。结果 成功构建表达野生型ihfB基因和嵌合体N-ihfB-C-ihfA的重组表达质粒pXTB1-ihfB和pXTB1-N-ihfB-C-ihfA, 并在宿主菌ER2566中诱导表达。pXTB1-ihfB为不溶性的包涵体表达, pXTB1-N-ihfB-C-ihfA为可溶性表达, 并获得高纯度的表达产物。结论 克隆表达了IHF 亚单位基因ihfB, IHF-的C端可显著提高IHF-的可溶性, 获得可溶性表达的纯化IHF-嵌合体蛋白, 为后续调控功能研究奠定了基础。
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出版历程
  • 收稿日期:  2016-02-03
  • 刊出日期:  2016-04-20

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