Difference between revisions of "Part:BBa K782024"

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Transcription activation like (TAL) effectors are proteins able to specifically bind desired DNA sequence. The central domain of the protein is constructed from variable number of tandem repeats differing only in two amino acids. The 12th and the 13th amino acid are called a “repeat variable diresidue” (RVD) and are responsible for specific interactions with the corresponding base pair (Scholze and Boch, 2011). This modularity of TAL effector binding domains therefore makes them a perfect tool to target specific DNA sequences.
 
Transcription activation like (TAL) effectors are proteins able to specifically bind desired DNA sequence. The central domain of the protein is constructed from variable number of tandem repeats differing only in two amino acids. The 12th and the 13th amino acid are called a “repeat variable diresidue” (RVD) and are responsible for specific interactions with the corresponding base pair (Scholze and Boch, 2011). This modularity of TAL effector binding domains therefore makes them a perfect tool to target specific DNA sequences.
  
Our construct contains [https://parts.igem.org/Part:BBa_K782067 two specific binding sites] for [https://parts.igem.org/wiki/index.php?title=Part:BBa_K782007 NicTAL12] and [https://parts.igem.org/wiki/index.php?title=Part:BBa_K782005 TALD] upstream of CMV promoter. Downstream of CMV promoter we cloned yellow fluorescent protein mCitrine an easy detectable monomer with excitation maximum at 516 nm and emission maximum at 529 nm. (Figure 1).  
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Our construct contains [https://parts.igem.org/Part:BBa_K782067 two specific binding sites] for [https://parts.igem.org/wiki/index.php?title=Part:BBa_K782007 NicTAL12] and [https://parts.igem.org/wiki/index.php?title=Part:BBa_K782005 TALD] upstream of minimal promoter. Downstream of minimal promoter we cloned yellow fluorescent protein mCitrine an easy detectable monomer with excitation maximum at 516 nm and emission maximum at 529 nm (Figure 1).  
After binding of [https://parts.igem.org/wiki/index.php?title=Part:BBa_K782011 NicTAL12:KRAB] or [https://parts.igem.org/wiki/index.php?title=Part:BBa_K782009 TALD:KRAB] on binding sites, a repression of reporter protein mCitrine occurs.  
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After binding of [https://parts.igem.org/wiki/index.php?title=Part:BBa_K782066 NicTAL12:VP16] or [https://parts.igem.org/wiki/index.php?title=Part:BBa_K782009 TALD:KRAB] on binding sites, an activation of reporter protein mCitrine occurs.  
  
 
Single binding sequence for NicTAL12 is: TCTATCAATGATAGA
 
Single binding sequence for NicTAL12 is: TCTATCAATGATAGA
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Single binding sequence for TALD is: TCGTCCAATAGCTTCTC
 
Single binding sequence for TALD is: TCGTCCAATAGCTTCTC
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Figure 1:
  
 
[[Image:Svn_12_2n-2d-pmin-mCit.png‎]]
 
[[Image:Svn_12_2n-2d-pmin-mCit.png‎]]

Revision as of 14:08, 26 September 2012

2x[NicTAL]+2x[TALD] operator_minimal promoter_mCitrine

  • TALD label represents TAL effector 1295 from zebrafish experiments (Sander et al., 2011).

Introduction

Transcription activation like (TAL) effectors are proteins able to specifically bind desired DNA sequence. The central domain of the protein is constructed from variable number of tandem repeats differing only in two amino acids. The 12th and the 13th amino acid are called a “repeat variable diresidue” (RVD) and are responsible for specific interactions with the corresponding base pair (Scholze and Boch, 2011). This modularity of TAL effector binding domains therefore makes them a perfect tool to target specific DNA sequences.

Our construct contains two specific binding sites for NicTAL12 and TALD upstream of minimal promoter. Downstream of minimal promoter we cloned yellow fluorescent protein mCitrine an easy detectable monomer with excitation maximum at 516 nm and emission maximum at 529 nm (Figure 1). After binding of NicTAL12:VP16 or TALD:KRAB on binding sites, an activation of reporter protein mCitrine occurs.

Single binding sequence for NicTAL12 is: TCTATCAATGATAGA

Single binding sequence for TALD is: TCGTCCAATAGCTTCTC

Figure 1:

Svn 12 2n-2d-pmin-mCit.png

Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal XhoI site found at 178
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal AgeI site found at 35
    Illegal AgeI site found at 105
  • 1000
    COMPATIBLE WITH RFC[1000]