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.
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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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