Difference between revisions of "Part:BBa K2549028"
(→‎Biology)
Line 12: Line 12:
 
<!-- Add more about the biology of this part here -->
 
<!-- Add more about the biology of this part here -->
 
===Biology===
 
===Biology===
−
=====It works as we designed =====
 
−
interaction between ZF43.8-VP64 and 8*ZF43.8-minCMV promotor
 
−
 
−
@@@@
 
−
 
 
=====Synthetic promotor operators regulated by artificial zinc finger-based transcription factors=====
 
=====Synthetic promotor operators regulated by artificial zinc finger-based transcription factors=====
 
Khalil AS et al have reported several synthetic promotor operators which can interact with artificial zinc finger-based transcription factors with high specificity and high orthogonality<ref>A synthetic biology framework for programming eukaryotic transcription functions. Khalil AS, Lu TK, Bashor CJ, ..., Joung JK, Collins JJ. Cell, 2012 Aug;150(3):647-58  PMID: 22863014; DOI: 10.1016/j.cell.2012.05.045</ref>.
 
Khalil AS et al have reported several synthetic promotor operators which can interact with artificial zinc finger-based transcription factors with high specificity and high orthogonality<ref>A synthetic biology framework for programming eukaryotic transcription functions. Khalil AS, Lu TK, Bashor CJ, ..., Joung JK, Collins JJ. Cell, 2012 Aug;150(3):647-58  PMID: 22863014; DOI: 10.1016/j.cell.2012.05.045</ref>.
Line 22: Line 17:
 
[[File:ZF2.jpg|none|240px|thumb|Khalil AS et al stated:''sTFs constructed from OPEN-engineered ZFs are orthogonal to one another. sTF43-8 activated noncognate Promoter21-16 due to the fortuitous creation of a sequence that is significantly similar to the binding sequence of 43-8, when the downstream BamHI restriction site is considered.'']]
 
[[File:ZF2.jpg|none|240px|thumb|Khalil AS et al stated:''sTFs constructed from OPEN-engineered ZFs are orthogonal to one another. sTF43-8 activated noncognate Promoter21-16 due to the fortuitous creation of a sequence that is significantly similar to the binding sequence of 43-8, when the downstream BamHI restriction site is considered.'']]
  
 +
===Characterization===
 +
=====It works as we designed.=====
 +
[[File:aTF-test.png|none|480px|thumb|'''Interaction between transcriptional activators and their binding sites. A degradable EGFP (d2EGFP) is linked downstream the promoter to indicate the expression level of it. DBD, DNA binding domain which is zinc finger in our assay. AD or SD, activating- or silencing-form transcriptional domain. RE, responsive elements. MFI, median fluorescence intensity.''']]
 +
 +
Flow cytometry results suggest that the transcriptional activators can effectively activate the responsive promoters with high specificity and high orthogonality.
  
 
<!-- Uncomment this to enable Functional Parameter display  
 
<!-- Uncomment this to enable Functional Parameter display  

Revision as of 13:45, 17 October 2018


8*ZF43.8-minCMV

This part is one of the response elements of our amplifier, also executing the combiner function. 8*ZF43.8 binding (Part:BBa_K2446013) sites is assembled using two 4*ZF43.8 binding sites (Part:BBa_K2446006) with a biobrick scar between them. Minimal CMV (Part:BBa_K2549049) is a promotor providing very low basal expression and high maximal expression after induction. This part can switch on the expression of gene downstream after induced by our zinc finger-based transcription activator.

Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    COMPATIBLE WITH RFC[1000]


Biology

Synthetic promotor operators regulated by artificial zinc finger-based transcription factors

Khalil AS et al have reported several synthetic promotor operators which can interact with artificial zinc finger-based transcription factors with high specificity and high orthogonality[1].

Khalil AS et al stated:sTFs constructed from OPEN-engineered ZFs are orthogonal to one another. sTF43-8 activated noncognate Promoter21-16 due to the fortuitous creation of a sequence that is significantly similar to the binding sequence of 43-8, when the downstream BamHI restriction site is considered.

Characterization

It works as we designed.
Interaction between transcriptional activators and their binding sites. A degradable EGFP (d2EGFP) is linked downstream the promoter to indicate the expression level of it. DBD, DNA binding domain which is zinc finger in our assay. AD or SD, activating- or silencing-form transcriptional domain. RE, responsive elements. MFI, median fluorescence intensity.

Flow cytometry results suggest that the transcriptional activators can effectively activate the responsive promoters with high specificity and high orthogonality.


References

  1. ↑ A synthetic biology framework for programming eukaryotic transcription functions. Khalil AS, Lu TK, Bashor CJ, ..., Joung JK, Collins JJ. Cell, 2012 Aug;150(3):647-58 PMID: 22863014; DOI: 10.1016/j.cell.2012.05.045