Part:BBa_K5490027

TALE BINDING SITE FOR SYNTHETIC NFAT

TALEs can be designed to bind almost any sequence of choice. Furthermore, one TALE sequence can target multiple TALE binding sites, provided the proximity between the sites is sufficiently low. These binding sites can be positioned anywhere, whether in central or peripheral DNA regions.

In this particular case, a TALE sequence is fused with the NFAT transcription factor to bind to 10 TALE binding site upstream of a minimal promoter. Upon receiving a stimulus, this setup activates transcription with high specificity

Sequence and Features

Assembly Compatibility:

• 10
  COMPATIBLE WITH RFC[10]
• 12
  INCOMPATIBLE WITH RFC[12]

  Illegal NotI site found at 141
  Illegal NotI site found at 211
  Illegal NotI site found at 506
  Illegal NotI site found at 576
  
• 21
  COMPATIBLE WITH RFC[21]
• 23
  COMPATIBLE WITH RFC[23]
• 25
  INCOMPATIBLE WITH RFC[25]

  Illegal AgeI site found at 7
  Illegal AgeI site found at 347
  Illegal AgeI site found at 372
  Illegal AgeI site found at 712
  
• 1000
  COMPATIBLE WITH RFC[1000]

TALEs (Transcription Activator-Like Effectors) are a fascinating system, first characterized in plant pathogens, and in many ways, they share similarities with the CRISPR-Cas9 system, serving as its predecessor. TALEs are modular and can be customized to target any DNA sequence by modifying specific amino acids. By fusing TALEs with various proteins, such as nucleases, they can be used as powerful genetic engineering tools to cut target DNA, or as activators or inhibitors to regulate specific regions of the genome.

The core of the TALE system consists of 33 to 35 amino acid repeats, with the specificity for DNA binding determined by the 12th and 13th residues of each repeat. These two residues form the Repeat-Variable Diresidue (RVD), which dictates which nucleotide the repeat will bind to:

NI → binds to Adenine (A)

HD → binds to Cytosine (C)

NG → binds to Thymine (T)

NN → binds to Guanine (G) (and sometimes Adenine)

By assembling these repeats, each of which targets a specific nucleotide, TALEs can be designed to bind almost any sequence of choice. Furthermore, one TALE sequence can target multiple TALE binding sites, provided the proximity between the sites is sufficiently low. These binding sites can be positioned anywhere, whether in central or peripheral DNA regions.

In this particular case, a TALE sequence is fused with the NFAT transcription factor to bind to 10 TALE binding sites upstream of a minimal promoter. Upon receiving a stimulus, this setup activates transcription with high specificity Zhang S, Chen H, Wang J. Generate TALE/TALEN as Easily and Rapidly as Generating CRISPR. Mol Ther Methods Clin Dev. 2019 Feb 19;13:310-320. doi: 10.1016/j.omtm.2019.02.004. PMID: 30923728; PMCID: PMC6423989.