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Example of a T-REx system controlling the expression of shRNA. 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).
The tac promoter consists of the '–35' region of the trp promoter and the '–10' region of the lac promoter (and differs from a related trc promoter by 1 bp [3]). The tac promoter is, therefore, inducible by IPTG (Isopropyl β- D -1-thiogalactopyranoside), whilst also allowing higher maximum gene expression than either the lac or trp promoters.
To perform a run-off transcription assay, a gene of interest, including the promoter, is cloned into a plasmid. [4] The plasmid is digested at a known restriction enzyme cut site downstream from the transcription start site such that the expected mRNA run-off product would be easily separated by gel electrophoresis .
The promoter resembles an eukaryotic one, though much simplified. It contains BRE and TATA elements and are recognized by TFB and TBP. [3] Promoters represent critical elements that can work in concert with other regulatory regions (enhancers, silencers, boundary elements/insulators) to direct the level of transcription of a given gene. A ...
The promoter initiates the transcription and is therefore the point of control for the expression of the cloned gene. The promoters used in expression vector are normally inducible, meaning that protein synthesis is only initiated when required by the introduction of an inducer such as IPTG. Gene expression however may also be constitutive (i.e ...
Soon, the lab was able to clone the T7 RNA polymerase and use it, along with the powerful T7 promoter, to transcribe copious amounts of almost any gene. [4] The development of the T7 expression system has been considered the most successful biotechnology developed at the Brookhaven National Laboratory, being licensed by over 900 companies which ...
A transcription factor is a protein that binds to specific DNA sequences (enhancer or promoter), either alone or with other proteins in a complex, to control the rate of transcription of genetic information from DNA to messenger RNA by promoting (serving as an activator) or blocking (serving as a repressor) the recruitment of RNA polymerase.
The measure of the promoter activity has a broad meaning. The promoter activity could be measured for different situations or research questions, [4] such as: estimation of the level of expression in comparison (relative) to some known value; how fast a gene is expressed after induction; the timing of expression relative to others genes