Part:BBa_K782014
12x[TALD] operator_CMV promoter_mCitrine
- TALD label represents TAL effector 1295 from zebrafish experiments (Sander et al., 2011).
Introduction
Transcription activation like (TAL) effectors are bacterial plant pathogen transcription factors that bind to DNA by recognizing a specific DNA sequence in which each base pair binds a single tandem repeat in in the TAL DNA-binding domain. A tandem TAL repeat contains 33 to 35 amino acids, where the 12th and the 13th amino acid, called a “repeat variable diresidue” (RVD) are responsible for specific interactions with the corresponding base pair (Scholze and Boch, 2011). All TAL repeats have almost identical sequences, differing only in the RVDs. This modularity of TAL effector binding domains therefore makes them a perfect tool to target specific DNA sequences by designing specific binding domains for a selected TAL effector. We designed twelve specific binding sites for TALD:KRAB upstream of CMV promoter (Figure 1). After binding of TALD:KRAB on binding sites, a repression of reporter protein mCitrine occurs. mCitrine is yellow fluorescent protein.
Figure 1. Shematic representation of twelve specific binding site for TALD upstream of CMV promoter and reporter protein mCitrine.
Single binding sequence for TALD is: TCGTCCAATAGCTTCTC
- mCitrine was provided from host lab.
- Binding sites for TAL effectors were ordered from IDT.
References
Scholze, H., and Boch, J. (2011) TAL effectors are remote controls for gene activation. Curr. Opin. Microbiol. 14, 47-53.
Sander, J. D., Cade, L., Khayter, C., Reyon, D., Peterson, R. T., Joung, J. K., and Yeh, J.-R. J. (2011) Targeted gene disruption in somatic zebrafish cells using engineered TALENs. Nature Biotechnology 29, 697–698
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21INCOMPATIBLE WITH RFC[21]Illegal BamHI site found at 465
Illegal XhoI site found at 1095 - 23COMPATIBLE WITH RFC[23]
- 25INCOMPATIBLE WITH RFC[25]Illegal AgeI site found at 214
Illegal AgeI site found at 446 - 1000COMPATIBLE WITH RFC[1000]
None |