1. An Improved Tet-On System for Gene Expression in Neurons Delivered by a Single Lentiviral Vector.
- Author
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Xuebi Tian, Gongming Wang, Ying Xu, Ping Wang, Shasha Chen, Hui Yang, Feng Gao, Aijun Xu, Fei Cao, Xiaogao Jin, Anne Manyande, and Yuke Tian
- Subjects
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GENETIC regulation , *NERVOUS system , *TRANSGENE expression , *LYASES - Abstract
AbstractIn most cases, the successful application of gene therapy requires the development of vectors that can provide regulated control of therapeutic gene expression. We have reconstituted the Tet-On (tetracycline-regulated transgene expression) two-component system in a single lentiviral vector with insertion of a chicken chromatin insulator (cHS4) element between the two expression cassettes. Optimization of this vector included an improved reverse tetracycline-dependent trans-activator (rtTA) sequence developed through HIV viral evolution, and an rtTA-dependent, Tet-responsive element containing modifications of the tetOsequence (TRE-tight1) to improve leaky basal transcription. Transduction of HeLa cells with these lentiviral vectors resulted in a high level of rtTA expression in the presence of doxycycline. In neuronal cells, rtTA expression driven by a neuron-specific enolase (NSE) promoter was more efficient than gene expression from a murine cytomegalovirus promoter. Transgene expression from the NSE promoter also provided tightly regulated gene expression in neurons in vivo. [ABSTRACT FROM AUTHOR]
- Published
- 2009
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