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Engineering of Thiamine Pyrophosphate Fluorescent Biosensors Based on Ribozyme Switches in Mammalian Cells

文献类型：期刊论文

作者Zhang Yuan-Yuan1,2,3; Cheng Hui3; Sun Yan3; Wang Jin-E3; Wu Zheng-Yan2; Pei Ren-Jun
刊名CHINESE JOURNAL OF ANALYTICAL CHEMISTRY
出版日期2017-02-01
卷号45期号:2页码:157-161
关键词Thiamine Pyrophosphate Ribozyme Switches Enhanced Green Fluorescence Protein Gene Expression Regulation Fluorescent Biosensors Mammalian Cells
DOI10.1016/S1872-2040(16)60992-1
文献子类Article
英文摘要As the main active form of intracellular thiamine (vitamin B1), thiamine pyrophosphate (TPP) is an essential cofactor in oxidative metabolism of the sugars, fatty acids and amino acids. By now, numerous TPP-dependent artificial riboswitch systems have been developed to regulate target gene expression but are limited in bacteria, fungi or plant cells. Herein, the activating (switch-on) and inhibiting (switch-off) TPP-dependent hammerhead ribozyme switches, inspired by the previously reported structures of prokaryotes screening, were investigated in mammalian cells. Such ribozyme switches were inserted into the 3 '-UTR of the enhanced green fluorescence protein (EGFP) gene through overlap extension PCR cloning and were further transfected into HEK293 cells at increasing concentrations of TPP. The EGFP gene-regulatory ability was analyzed with fluorescent microscope and flow cytometry. These resultant TPP-inducible gene regulation devices showed obvious ligand dose-dependency and excellent specificity. Upon the addition of 150 mu M TPP, two "switch-on" and one "switch-off" constructs demonstrated 3.1-fold or 1.9-fold increment and 2.3-fold reduction of EGFP level, respectively. The ligand-responsive ribozyme switches, by transforming the change of TPP concentration into the visual reporter genetic expression in cells, enable efficient development of label-free, noninvasive and high specific biosensors in living mammalian cells.
WOS关键词GENE-EXPRESSION ; MESSENGER-RNAS ; RIBOSWITCHES ; APTAMERS ; LIGANDS ; SYSTEM
WOS研究方向Chemistry
语种英语
WOS记录号WOS:000397209600003
资助机构National Natural Science Foundation of China(21628502 ; National Natural Science Foundation of China(21628502 ; National Natural Science Foundation of China(21628502 ; National Natural Science Foundation of China(21628502 ; National Natural Science Foundation of China(21628502 ; National Natural Science Foundation of China(21628502 ; National Natural Science Foundation of China(21628502 ; National Natural Science Foundation of China(21628502 ; Natural Science Foundation of Jiangsu Province, China(BK20130357 ; Natural Science Foundation of Jiangsu Province, China(BK20130357 ; Natural Science Foundation of Jiangsu Province, China(BK20130357 ; Natural Science Foundation of Jiangsu Province, China(BK20130357 ; Natural Science Foundation of Jiangsu Province, China(BK20130357 ; Natural Science Foundation of Jiangsu Province, China(BK20130357 ; Natural Science Foundation of Jiangsu Province, China(BK20130357 ; Natural Science Foundation of Jiangsu Province, China(BK20130357 ; 21507156 ; 21507156 ; 21507156 ; 21507156 ; 21507156 ; 21507156 ; 21507156 ; 21507156 ; BK20130351) ; BK20130351) ; BK20130351) ; BK20130351) ; BK20130351) ; BK20130351) ; BK20130351) ; BK20130351) ; 31300096) ; 31300096) ; 31300096) ; 31300096) ; 31300096) ; 31300096) ; 31300096) ; 31300096) ; National Natural Science Foundation of China(21628502 ; National Natural Science Foundation of China(21628502 ; National Natural Science Foundation of China(21628502 ; National Natural Science Foundation of China(21628502 ; National Natural Science Foundation of China(21628502 ; National Natural Science Foundation of China(21628502 ; National Natural Science Foundation of China(21628502 ; National Natural Science Foundation of China(21628502 ; Natural Science Foundation of Jiangsu Province, China(BK20130357 ; Natural Science Foundation of Jiangsu Province, China(BK20130357 ; Natural Science Foundation of Jiangsu Province, China(BK20130357 ; Natural Science Foundation of Jiangsu Province, China(BK20130357 ; Natural Science Foundation of Jiangsu Province, China(BK20130357 ; Natural Science Foundation of Jiangsu Province, China(BK20130357 ; Natural Science Foundation of Jiangsu Province, China(BK20130357 ; Natural Science Foundation of Jiangsu Province, China(BK20130357 ; 21507156 ; 21507156 ; 21507156 ; 21507156 ; 21507156 ; 21507156 ; 21507156 ; 21507156 ; BK20130351) ; BK20130351) ; BK20130351) ; BK20130351) ; BK20130351) ; BK20130351) ; BK20130351) ; BK20130351) ; 31300096) ; 31300096) ; 31300096) ; 31300096) ; 31300096) ; 31300096) ; 31300096) ; 31300096)
源URL[http://ir.hfcas.ac.cn:8080/handle/334002/33408]  
专题合肥物质科学研究院_技术生物与农业工程研究所
作者单位1.Anhui Med Univ, Sch Life Sci, Hefei 230032, Peoples R China
2.Chinese Acad Sci, Hefei Inst Phys Sci, Hefei 3, Peoples R China
3.Suzhou Inst Nano Tech & Nano Bion, Chinese Acad Sci, Suzhou 215123, Peoples R China
推荐引用方式
GB/T 7714		 Zhang Yuan-Yuan,Cheng Hui,Sun Yan,et al. Engineering of Thiamine Pyrophosphate Fluorescent Biosensors Based on Ribozyme Switches in Mammalian Cells[J]. CHINESE JOURNAL OF ANALYTICAL CHEMISTRY,2017,45(2):157-161.
APA Zhang Yuan-Yuan,Cheng Hui,Sun Yan,Wang Jin-E,Wu Zheng-Yan,&Pei Ren-Jun.(2017).Engineering of Thiamine Pyrophosphate Fluorescent Biosensors Based on Ribozyme Switches in Mammalian Cells.CHINESE JOURNAL OF ANALYTICAL CHEMISTRY,45(2),157-161.
MLA Zhang Yuan-Yuan,et al."Engineering of Thiamine Pyrophosphate Fluorescent Biosensors Based on Ribozyme Switches in Mammalian Cells".CHINESE JOURNAL OF ANALYTICAL CHEMISTRY 45.2(2017):157-161.

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来源：合肥物质科学研究院

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