Part:BBa_K2918010
T7sp1 promoter
A T7 promoter that should give 50% of wild type T7 promoter expression and with a binding site for Transcription Activator like Effector protein (TALE) repressor.
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]
Usage and Biology
The (<a target=”_blank” href=”https://parts.igem.org/Part:BBa_K2918006”>0.5 T7 promoter variant</a>) was engineered to contain a binding site for a Transcriptional Activator like Effector protein (TALE). TALE proteins consist of repeats where 12th and 13th amino acids can vary, the repeats are called the repeat variable diresidue (RVD) (Segall-Shapiro, Sontag et al. 2018). These RVDs have been shown to bind to DNA in a simple one-to-one binding code. A unique 18bp binding site was incorporated into the promoter, this binding site recruits a specific TALE protein called TALEsp1 that acts as a repressor (Segall-Shapiro, Sontag et al. 2018). The TALEsp1 protein was designed to bind protein specifically at the binding site and are predicted not be bind anywhere in the genome. The position of the binding site can be readjusted to alter the extent of repression by the TALE protein.
Strain Construction
Strain construction: Aim: To clone the promoter in a level 0 MoClo backbone (<a target=”_blank” href=”https://www.addgene.org/47984/”>pICH41233</a>). Procedure: The DNA sequence of the part was synthesized by IDT with flanking BpiI sites and respective MoClo compatible promoter overhangs. The cloning protocol can be found in the MoClo section below.
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