Difference between revisions of "Part:BBa K782007"

(Characterization)
(Introduction)
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TAL effectors (TALEs) are bacterial plant pathogen transcription factors, that bind to DNA by specifically recognizing one base pair with a single tandem repeat in their DNA-binding domain. A tandem TALE repeat contains 33 to 35 amino acids, where the 12th and 13th amino acid, called a “repeat variable diresidue” (RVD), are responsible for specific interactions with the corresponding base pair (Scholze and Boch, 2011).
 
TAL effectors (TALEs) are bacterial plant pathogen transcription factors, that bind to DNA by specifically recognizing one base pair with a single tandem repeat in their DNA-binding domain. A tandem TALE repeat contains 33 to 35 amino acids, where the 12th and 13th amino acid, called a “repeat variable diresidue” (RVD), are responsible for specific interactions with the corresponding base pair (Scholze and Boch, 2011).
  
[[Image:Svn_12_NicTAL12_.png]]
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[[Image:Svn_12_NicTAL12--.png]]
  
 
'''Figure 1:''' Schematic representation of the construct.
 
'''Figure 1:''' Schematic representation of the construct.

Revision as of 08:41, 26 September 2012

NicTAL12:NLS DNA binding domain

Introduction

TAL effectors (TALEs) are bacterial plant pathogen transcription factors, that bind to DNA by specifically recognizing one base pair with a single tandem repeat in their DNA-binding domain. A tandem TALE repeat contains 33 to 35 amino acids, where the 12th and 13th amino acid, called a “repeat variable diresidue” (RVD), are responsible for specific interactions with the corresponding base pair (Scholze and Boch, 2011).

Svn 12 NicTAL12--.png

Figure 1: Schematic representation of the construct.

Single binding sequence for NicTAL:TCTATCAATGATAGA

Characterization

We discovered that NicTAL part deposited in the Registry by the Slovenian iGEM2010 team was missing subdomain in DNA-binding domain, so [http://2012.igem.org/Team:Slovenia our team] added this missing part on N terminal sequence. Apart from adding N terminal sequence,we added HIS tag on N terminal end and NLS on terminal C end of NicTAL12 (Figure 2). This construct was later used for designing TAL-based activator and repressor by adding VP16 and KRAB domain.

Svn 12 NicTAL10vs12.PNG

Figure 2:Sequence alignment of NicTAL10 and NicTAL12 showing differences between constructs.


Results obtained by testing NicTAL12:KRAB repressor and activator, proved that NicTAL12 binds to its binding sites better than NicTAL10.

Svn 12 NicTAL graf.png

Figure 3: Results of testing NicTAL:KRAB repressor demonstrated better binding ability of NicTAL12.

References

Scholze, H., and Boch, J. (2011) TAL effectors are remote controls for gene activation. Curr. Opin. Microbiol. 14, 47-53.

Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal BamHI site found at 2133
    Illegal XhoI site found at 1224
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    COMPATIBLE WITH RFC[1000]