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李志峰, 聂军, 戴迎春, 李建栋, 陈清, 俞守义. 副溶血弧菌不耐热溶血毒素的表达与纯化[J]. 中国公共卫生, 2004, 20(1): 27-28.
引用本文: 李志峰, 聂军, 戴迎春, 李建栋, 陈清, 俞守义. 副溶血弧菌不耐热溶血毒素的表达与纯化[J]. 中国公共卫生, 2004, 20(1): 27-28.
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LI Zhi-feng, NIE Jun, DAI Ying-chun, . Expression and purification of thermolabile hemolysin of vibrio parahaemolyticus[J]. Chinese Journal of Public Health, 2004, 20(1): 27-28.
Citation: LI Zhi-feng, NIE Jun, DAI Ying-chun, . Expression and purification of thermolabile hemolysin of vibrio parahaemolyticus[J]. Chinese Journal of Public Health, 2004, 20(1): 27-28.
shu

副溶血弧菌不耐热溶血毒素的表达与纯化

Expression and purification of thermolabile hemolysin of vibrio parahaemolyticus

    摘要
  • 摘要:
      目的   表达和纯化融合表达的副溶血弧菌不耐热溶血毒素.
      方法   将表达载体pET32a+-tlh转化大肠杆菌BL21(λDE3), 诱导表达副溶血弧菌不耐热溶血毒素, 表达产物用聚丙稀酰胺凝胶电泳(SDS-PAGE)鉴定, 8mmol/L的尿素溶解包涵体, 用6×his组氨酸蛋白质镍亲和层析树脂亲和层析纯化目的蛋白.
      结果   诱导表达的目的蛋白以包涵体形式存在, 其含量约占菌体总蛋白的10%, 纯化获得了高纯度的融合表达的副溶血弧菌不耐热溶血毒素.
      结论   转化有重组质粒pET32a+-tlh的BL21(λDE3)可稳定高效地表达目的蛋白, 采用低浓度的咪唑、β-巯基乙醇和Tritonx-100等方法减少杂蛋白污染, 亲和层析纯化目的蛋白, 可获得高纯度的目的蛋白.

     

    Abstract:
      Objective   To express and obtain the fused protein thermolabile hemolysin(TLH)of vibrio parahaemolyticus (VP).
      Methods   Prokaryotic expression plasmid p ET32a+-tlh was constructed and incorporated into E.coil BL21(λDE3) the engineered bacteria were inducted by IPTG to express TLH, which was identified by SDS-PAGE and purified by using 6×hisNi-NTA Resin.
      Results   the induced expression fusion protein was inclusion, which expression reached to 10% of the total protein of BL21(λDE3), the high pure fused protein TLH was obtained by using affinity chromatography method.
      Conclusion   the recombinant plasmid p ET32a+-tlh had stably and efficiently expression in BL21(λDE3), the high pure fused protein TLH could be obtained by using affinity chromatography method by using low concentration imidazole β,-ME and Triton x-100 to reduce contaminated protein.

     

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