口腔医学研究

口腔医学研究 ›› 2017, Vol. 33 ›› Issue (2): 140-144.DOI: 10.13701/j.cnki.kqyxyj.2017.02.006

• 基础研究论著 • 上一篇    下一篇

IPTG浓度对大肠杆菌中重组GroEL表达的影响

肖玲玲, 杨洁, 张瑞, 陈小凤, 刘玉, 吴娟, 苗雷英, 孙卫斌*   

  1. 南京大学医学院附属口腔医院,南京市口腔医院牙周病科 江苏 南京 210008
  • 收稿日期:2016-09-21 出版日期:2017-02-20 发布日期:2017-02-22
  • 通讯作者: 孙卫斌,电话:025-83620138
  • 作者简介:肖玲玲(1990~ ),女,江苏人,硕士在读,主要从事牙周病相关研究。
  • 基金资助:
    国家自然科学基金(编号:81271155、51472115)南京市科技发展计划项目(编号:201402036)南京市医学科技发展项目(编号:YKK14107)

Effect of IPTG Concentrations on Recombinant Protein GroEL Production in Escherichia Coli.

XIAO Ling-ling, YANG Jie, ZHANG Rui, CHEN Xiao-feng, LIU Yu, WU Juan, MIAO Lei-ying, SUN Wei-bin*   

  1. Nanjing Stomatological Hospital, Medical School of Nanjing University, Nanjing 210008, China
  • Received:2016-09-21 Online:2017-02-20 Published:2017-02-22

摘要： 目的:探究诱导重组GroEL(牙周炎相关动脉粥样硬化疫苗)的最佳IPTG浓度,并分析IPTG浓度影响重组GroEL表达量的原因。方法:构建GroEL的大肠杆菌( Escherichia Coli, E.coli)表达体系,使用不同浓度IPTG(0,10,20,30,50,100 μmol/L)诱导构建的E.coli (GroEL-E.coli),运用SDS-PAGE及相关软件定量分析GroEL表达量;透射电子显微镜及A值测定法分别检测IPTG浓度对GroEL-E.coli内部包涵体(IB)形成及GroEL-E.coli生长的影响。结果:重组GroEL主要以可溶性蛋白的形式存在。IPTG的加入可增加GroEL表达量,当IPTG浓度为30 μmol/L时GroEL表达量最多。IPTG可促进IB形成并抑制GroEL-E.coli生长,且影响效果随IPTG浓度增加而增强。结论:IPTG浓度影响GroEL-E.coli中重组GroEL的表达,最佳IPTG浓度为30 μmol/L;IPTG可通过诱导IB形成及抑制细菌增殖影响GroEL表达量。

关键词: 大肠杆菌, 重组蛋白, Isopropyl-β-D-thiogalactopyranoside

Abstract: Objective: To explore the effect of IPTG concentrations on recombinant GroEL production in Escherichia Coli (E.coli) and analyze the reasons. Methods: The E. coli expression system for GroEL was constructed and induced by different IPTG concentrations (0, 10, 20, 30, 50, 100 μmol/L) for GroEL expression, and the SDS-PAGE and gel analysis software were used to determine the amount of expressed GroEL. The transmission electron microscopy (TEM) and OD-measuring were used to detect IB formation and bacterial growth. Results: GroEL mainly exists in the soluble form. The addition of IPTG can obviously increase the GroEL expression, and 30 μmol/L IPTG leaded to the most GroEL expression. Conclusion: The recombinant GroEL expression in E. coli differed among IPTG concentrations, and the optimal IPTG concentration was 30μmol/L. The main reasons for IPTG effect on GroEL yield were IB formation and growth inhibition.

Key words: Escherichia coli , Recombinant protein Isopropyl-β-D-thiogalactopyranoside

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