Tetracycline (Tet)-induction system is a powerful tool to control the expression of target genes in mammalian cells.
Figure1: Tet-On system (Gene regulation systems for gene therapy applications in the central nervous system,2012)
A classic and enhanced Tet-On system mediated by regular plasmid has been constructed at Creative BioMart. Both systems are transient because the plasmid vector is hardly integrated into the host genome. The system can deliver foreign fragments to mammalian cells with simple operation and good efficiency in various cell types, so it is expected to provide a wide range of services for biomedical researchers.
Our Tet-On plasmid vector was designed to silence the system actively through tTS protein without tetracycline. tTS and rtTA are expressed as fusion proteins and as gene activation switches. In the presence of tetracycline, it is expressed rapidly by the strong activation of rtTA protein. Our vector has been optimized to replicate high copy numbers in E.coli and to be transfected efficiently in many mammalian cell lines.
Unlike the Tet-on vector expressing only rtTA, our Tet-On vector system acts as a real tetracycline-regulated on/off switch for controlling gene expression, which can minimize background expression without induction, and lead to high sensitivity and high dynamic range induced by tetracycline. Delivery of plasmid vectors into cells by conventional transfection is technically straightforward and much easier than virus-based vectors, which need to package virus vectors to survive.
The expression mechanism of our enhanced Tet-On system is the same as that of the standard version. The difference is that the enhanced version is designed to allow tissue-specific induction of target transgenes in the presence of tetracycline, while minimizing leakage expression of non-target tissues in the absence of tetracycline. Instead of fusion expression, the GOI, tTS gene and rtTA gene are driven by three different promoters respectively. In the absence of tetracycline, tTS, which is widely expressed in all tissues, binds to TRE promoter with high affinity, thus inhibiting GOI expression in all tissues. In the presence of tetracycline, rtTA is specifically expressed in the target tissue.
TRE promoter can drive the expression of GOI at a very high level. In addition, conventional transfection of plasmids usually results in very high copy numbers (up to thousands of copies per cell). This can lead to a very high level of expression of the genes carried on the vector.
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