Tet-One Inducible Expression System
Tet-One Inducible Expression System
Brand: Takara Bio.
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Tet-One Inducible Expression System
The Tet-One Inducible Expression System is a tetracycline-inducible gene expression system that allows you to establish a tightly inducible expression system for your gene of interest in mammalian cells using an all-in-one plasmid vector. The pTetOne Vector expresses the Tet-On 3G transactivator from the constitutive human PGK promoter in the forward orientation, and your gene of interest from the PTRE3GS promoter in the reverse orientation. In the presence of doxycycline (Dox), the Tet-On 3G transactivator specifically binds and activates high-level transcription from the inducible promoter that controls expression of your gene. In addition, the kit contains linear selection markers for resistance to puromycin and hygromycin, either of which can be cotransfected with the pTetOne Vector to obtain stable clones. It also includes Xfect Transfection Reagent and Tet System Approved FBS.
Overview
- All-in-one vector design provides all the power of Tet-On 3G technology in a single vector, making inducible expression as easy as constitutive expression
- Low background and high maximum induced expression
- High sensitivity to doxycycline: near maximum expression at 10 ng/ml doxycycline
- Inducible expression in a broad range of tissues, because the Tet-On 3G transactivator is expressed from the human PGK promoter
- High-titer lentivirus packaging system provided with Lenti-X Tet-One systems
- Retro-X systems incude our Retro-X Universal Packaging System
- The AAVpro system uses a helper-free packaging system and includes extraction reagents for efficient isolation of AAV particles
Applications
- Tightly controlled inducible expression in mammalian cells
1,000-fold induction of luciferase expression in transient transfections
1,000-fold induction of luciferase expression in transient transfections. 293T cells were transiently transfected with the pTetOne-Luc control plasmid and treated with 10-fold dilutions of doxycycline. 24 hr after treatment, the cells were analyzed for luciferase activity.
Tet-One System Mechanism — The Tet-On 3G transactivator protein is expressed constitutively from the human PGK promoter (PPGK) but is unable to bind to the TRE3G promoter (PTRE3G) in the absence of doxycycline (Dox)
Tet-One System Mechanism — The Tet-On 3G transactivator protein is expressed constitutively from the human PGK promoter (PPGK) but is unable to bind to the TRE3G promoter (PTRE3G) in the absence of doxycycline (Dox). When bound by Dox, supplied in the culture medium, the transactivator undergoes a conformational change, binds to PTRE3G and activates transcription of a transgene cloned downstream.
Heinz, N., Schambach, A., Galla, M., Maetzig, T., Baum, C., Loew R. & Schiedlmeier, B. Retroviral and transposon-based Tet-regulated all-in-one vectors with reduced background expression and improved dynamic range. Human Gene Ther. 22, 166–176 (2011).
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1,000-fold induction of luciferase expression in transient transfections
1,000-fold induction of luciferase expression in transient transfections. 293T cells were transiently transfected with the pTetOne-Luc control plasmid and treated with 10-fold dilutions of doxycycline. 24 hr after treatment, the cells were analyzed for luciferase activity.
Tet-One System Mechanism — The Tet-On 3G transactivator protein is expressed constitutively from the human PGK promoter (PPGK) but is unable to bind to the TRE3G promoter (PTRE3G) in the absence of doxycycline (Dox)
Tet-One System Mechanism — The Tet-On 3G transactivator protein is expressed constitutively from the human PGK promoter (PPGK) but is unable to bind to the TRE3G promoter (PTRE3G) in the absence of doxycycline (Dox). When bound by Dox, supplied in the culture medium, the transactivator undergoes a conformational change, binds to PTRE3G and activates transcription of a transgene cloned downstream.
