Part:BBa_K1925002

Tetracycline-responsed U6 promoter

If you want to use tet-off or tet-on system to selectively express a RNA(lincRNA\shRNA) rather than a protein-coding gene,this Tetracycline-responsed promoter can be an best choice cuz it's a pol III type promoter.Comparing with tradtional Tetracycline-responsed CMV promoter,it's more suitable for RNA expression.

Sequence and Features

Assembly Compatibility:

• 10
  COMPATIBLE WITH RFC[10]
• 12
  COMPATIBLE WITH RFC[12]
• 21
  COMPATIBLE WITH RFC[21]
• 23
  COMPATIBLE WITH RFC[23]
• 25
  COMPATIBLE WITH RFC[25]
• 1000
  COMPATIBLE WITH RFC[1000]

Usage and Biology

Tetracycline-Controlled Transcriptional Activation is a method of inducible gene expression where transcription is reversibly turned on or off in the presence of the antibiotic tetracycline or one of its derivatives (e.g. doxycycline).

Tetracycline-controlled gene expression is based upon the mechanism of resistance to tetracycline antibiotic treatment found in gram-negative bacteria. In nature, the Ptet promoter expresses TetR, the repressor, and TetA, the protein that pumps tetracycline antibiotic out of the cell.

The difference between Tet-On and Tet-Off is not whether the transactivator turns a gene on or off, as the name might suggest; rather, both proteins activate expression. The difference relates to their respective response to tetracycline or doxycycline (Dox, a more stable tetracycline analogue); Tet-Off activates expression in the absence of Dox, whereas Tet-On activates in the presence of Dox.

References

(1)Gossen M, Freundlieb S, Bender G, Müller G, Hillen W, Bujard H (1995). "Transcriptional activation by tetracyclines in mammalian cells". Science (journal). 268 (5218): 1766–1769. doi:10.1126/science.7792603. PMID 7792603. (2)Orth P, Schnappinger D, Hillen W, Saenger W, Hinrichs W (2000). "Structural basis of gene regulation by the tetracycline inducible Tet repressor-operator system" (PDF). Nature Structural & Molecular Biology. 7 (3): 215–219. PMID 10700280. (3)Tet-On and Tet-Off are registered trademarks of Clontech Laboratories, Inc. in the United States. Bujard, Hermann; M. Gossen (1992). (4)"Tight Control of Gene Expression in Mammalian Cells by Tetracycline-Responsive Promoters.". Proc. Natl. Acad. Sci. U.S.A. 89 (12): 5547–51. doi:10.1073/pnas.89.12.5547. PMC 49329free to read. PMID 1319065. Allen, Nicholas D.; Antonius Plagge; Gavin Kelsey (2000). (5)"Directed Mutagenesis in Embryonic Stem Cells". Mouse Genetics and Transgenics: 259–263.Urlinger, Stefanie; Baron, Udo; Thellmann, Marion; Hasan, Mazahir T.; Bujard, Herman; Hillen, Wolfgang (2000). (6)"Exploring the sequence space for tetracycline-dependent transcriptional activators: Novel mutations yield expanded range and sensitivity.". Proc. Natl. Acad. Sci. U.S.A. 97 (14): 7963–8. doi:10.1073/pnas.130192197. PMC 16653free to read. PMID 10859354.Zhou, X.; Vink, M.; Klave, B.; Berkhout, B.; Das, A.T. (2006). (7)"Optimization of the Tet-On system for regulated gene expression through viral evolution.". Gene Ther. 13 (19): 1382–1390. doi:10.1038/sj.gt.3302780. PMID 16724096. http://www.lifetechnologies.com/ca/en/home/references/protocols/proteins-expression-isolation-and-analysis/protein-expression-protocol/inducible-protein-expression-using-the-trex-system.reg.us.html/ Sohal DS, Ngheim M, Crackower MA, Witt SA, Kimball TR, Tymitz KM, Penniger JM, Molkentin JD (2001) (8) Temporally regulated and tissue-specific gene manipulations in the adult and embryonic heart using tamoxifen-inducible Cre protein. Circ Res 89:20-25Moullan N, Mouchiroud L, Wang X, Ryu D, Williams EG, Mottis A, Jovaisaite V, Frochaux MV, Quiros PM, Deplancke B, Houtkooper RH, Auwerx J (2015). (9)"Tetracyclines Disturb Mitochondrial Function across Eukaryotic Models: A Call for Caution in Biomedical Research.". Celll Reports. 10 (10): 1681–91. doi:10.1016/j.celrep.2015.02.034. PMID 25772356.Chatzispyrou IA, Held NM, Mouchiroud L, Auwerx J, Houtkooper RH (2015). (10) "Tetracycline antibiotics impair mitochondrial function and its experimental use confounds research.". Cancer Research. 72 (21): 4446–9. doi:10.1158/0008-5472.CAN-15-1626. PMID 26475870.