中南医学科学杂志2016，Vol.44Issue(2)：121-124,129,5.DOI:10.15972/j.cnki.43-1509/r.2016.02.001
梅毒螺旋体粘附素Tp0751重组菌影的构建与鉴定
Construction and Identification of Recombinant E.coli Bacterial Ghosts Expressing Treponema Pallidum Adhesin Tp0751
摘要
Abstract
Objective To construct the recombinant E.coli bacterial ghosts (EBG) expressing Treponema pallidum ( Tp) adhesin Tp0751 and to further investigate its potential immunoprotection against Tp infection. Methods E.coli DH5α cells transformed with the plasmids pHH43,containing phagephiX174 lysis E gene,were induced at 42 ℃ to produce EBG. The lysis rate was calculated and EBG were identified by DNA agarose gel electrophoresis and Transmission Electron Microscope ( TEM ) . E. coli BL21 cells with prokaryotic double expression recombinant plasmids pET28a-E′-Tp0751-E-box,containing lysis gene cassette (E-box),membrane anchor sequence E’-linker and target gene Tp0751,were induced to express recombinant prtotein Tp0751 at 28℃ and to generate EBG at 42℃.Recombinant Tp0751 was identified by western blot.The lysis rate of recombinant EBG( rEBG) was calculated,DNA agarose gel electrophoresis and TEM were used to identify the EBG. Results The lysis rate of EBG was 98.13%.No DNA ladder was observed by DNA gel electrophoresis and TEM showed that the vast majority of bacteria were lysed into empty cell envelopes lacking cy-toplasmic content yet retaining unaltered morphological features of their living counterparts.At 28 ℃,rEBG effectively ex-pressed Tp0751 that were specifically reactive with syphilitic sera by Western blot.At 42℃,the lysis rate of rEBG was 96. 37%.and similar to EBG tested by electrophoresis and TEM. Conclusion The recombinant EBG expressing Tp0751 are successfully constructed and expressed Tp0751 shows good immunoreactivity.关键词
梅毒螺旋体/Tp0751/粘附素/菌影/大肠埃希菌Key words
Treponema pallidum/Tp0751/adhesion/bacterial ghost/Esherichia coli分类
医药卫生引用本文复制引用
江银波,曹二龙,朱洪,赵飞骏,甄红娇,张佳俐,余坚,曾铁兵..梅毒螺旋体粘附素Tp0751重组菌影的构建与鉴定[J].中南医学科学杂志,2016,44(2):121-124,129,5.基金项目
国家自然科学基金(No.81273322)；湖南省教育厅开放创新平台基金(No.15K110)；湖南省教育厅资助科研项目(No.15C1256)；湖南省分子靶标新药研究协同创新中心(湘教通[2014])405号)；湖南省高校科技创新团队支持计划资助(湘教通[2010]53号) （No.81273322）
