Full-Text Search:
Home|Journal Papers|About CNKI|User Service|FAQ|Contact Us|中文
《Acta Medicinae Universitatis Scientiae et Technologiae Huazhong》 2018-03
Add to Favorite Get Latest Update

Construction,Expression and Characteristic Analysis of Recombinant Human TIM-4-EGFP Fusion Protein

Chen Zhizhong;Hu Lihua;Qin Jilin;Department of Laboratory,the People's Hospital of Guangxi Zhuang Autonomous Region;Department of Laboratory,Union Hospital,Tongji Medical College,Huazhong University of Science and Technology;Center for Reproductive Medicine and Genetics,the People's Hospital of Guangxi Zhuang Autonomous Region;  
Objective The present study aimed to explore the expression and purification of a fusion protein of human TIM-4 and EGFP in Escherichia coli(E.coli)and evaluate its bioactivity.Methods The cDNA fragments of human TIM-4 and EGFP genes were respectively amplified by RT-PCR and cloned into prokaryotic expression vector pET-28 a.The constructed recombinant plasmid pET-28 a-TIM-4-EGFP was transformed to E.coli BL21(DE3)for the expression under the induction of IPTG.The expressed protein was purified by Ni-NTA resin.Recombinant protein was analyzed by SDS-PAGE and Western blotting,and its binding activity to the apoptotic cells was detected under the fluorescence microscope.Results The TIM-4-EGFP vector was constructed and expressed in E.coli.The TIM-4-EGFP fusion protein was identified and verified by SDS-PAGE and Western blotting.Our results demonstrated that all the TIM-4-EGFP fusion proteins recognize and bind directly to apoptotic cells,but not to viable cells.We further verified that the interactions of TIM-4-EGFP with apoptotic cells were blocked by TIM-4-Ig fusion proteins.Conclusion We successfully constructed a fusion protein encoding human TIM-4 and EGFP,and expressed it in E.coli.The fusion protein shows a readily obtainable source of biologically active TIM-4,which has considerable potential for further studies on human TIM-4 and its receptor.
【Fund】: 国家自然科学基金资助项目(No.81260007);; 广西自然科学基金资助项目(No.2017GXNSFAA198074);; 广西卫生厅课题资助项目(No.Z2012316 No.Z2013403)
【CateGory Index】: R392
Download(CAJ format) Download(PDF format)
CAJViewer7.0 supports all the CNKI file formats; AdobeReader only supports the PDF format.
©2006 Tsinghua Tongfang Knowledge Network Technology Co., Ltd.(Beijing)(TTKN) All rights reserved