Part:BBa_K1431101
TetOn-3G, an ideal controller of mammalian gene expression with TRE-3G promoter+PolyA
Tet-On(Tetracycline-Controlled Transcriptional Activation,also known as rtTA2S-M2) is a system of inducible gene expression systems for mammalian cells. Tet-On 3G (also known as rtTA-V16[citation needed]) is similar to Tet-On but was derived from rtTA2S-S2 rather than rtTA2S-M2. The Tet-On 3G protein has 5 amino acid differences compared to Tet-On which appear to increase its sensitivity to doxycycline(Dox) even further. Tet-On 3G is sensitive to 100-fold less Dox and is 7-fold more active than the original Tet-On.[http://en.wikipedia.org/wiki/Tetracycline-controlled_transcriptional_activation]
Target cells that express the Tet-On 3G transactivator protein and contain a gene of interest (GOI) under the control of a TRE3G promoter (PTRE3G,BBa_K1431301[1]) will express high levels of GOI, but only when cultured in the presence of Dox, which is a synthetic tetracycline derivative(the mechanism of Tet-On 3G system show below). In the presence of Dox, Tet-On 3G binds specifically to PTRE3G and activates transcription of the downstream GOI. PTRE3G lacks binding sites for endogenous mammalian transcription factors, so it is virtually silent in the absence of induction.
The Dox concentrations required for induction of Tet-On Systems are far below cytotoxic levels for either cell culture or transgenic studies, and Tet-On 3G responds to even lower concentrations than its predecessors (Zhou et. al,. 2006). Note that Tet-On Systems respond well only to doxycycline, and not to tetracycline (Gossen & Bujard, 1995). The half-life of Dox in cell culture medium is 24 hours. To maintain continuous inducible GOI expression in cell culture, the medium should be replenished with Dox every 48 hours.(Source: Clontech)
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000COMPATIBLE WITH RFC[1000]
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