Difference between revisions of "Part:BBa K3788021"
 
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<u> Introduction </u> :
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<p>The Part BBa_K3788021 is a part composed from the parts BBa_K3788017 (lexA-repressed promoter), BBa_K3788015, BBa_E1010 (RFP) fused to <i>cal</i> gene and BBa_E0040 (GFP)  fused to <i>caa</i> gene.</p>
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<p>With this design, this part integrates a part of the <i>cal</i> gene, coding for the lysis protein CaL. It is also composed from <i>caa</i> and <i>cai</i> genes, respectively for the Colicin A toxin and the immunity protein Cal.</p>
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<p>In normal conditions, <i>caa</i> and <i>cai</i> genes are positioned as an operon, and the <i>cai</i> gene is located on the antisense strand and expressed in a constitutive manner.</p>
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<p>This system’s regulation is constituted from many regulation elements, which permits it to obtain a fine-tuned regulation.</p>
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<p>This system is located on a plasmid present in some <i>E. Coli</i> populations. It allows Colicin A accumulation on the host cell, and then induces its death due to CaL action, which provokes Colicin A liberation on the extracellular medium, which induces the death of bacteria that don’t produce Cal protein. </p>
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<p><i>cal</i> gene codes for CaL protein, a lipopeptide with a length between 28 and 48 amino acid residues. This protein is produced as a precursor and then processed. Once the protein is processed, it can form pores in the bacterial membrane, which leads to the quasi-lysis of the host cell, and provokes a leak from the cellular component into the medium.</p>
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<u>Design</u> :
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<p>This construction is used to study the time-shift between ColA and Cal production after induction of the promoter, by measuring the fluorescence kinetics.</p>
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Induced by the SOS response (LexA dependent) upon DNA damages (UV, mitomycin…). This promoter allow the expression of the operon caa and cal. caa gene is coding for Colicin A (ColA) a toxin toxin produced by E. coli strains, and active against other non-immune E.coli strains. In this contruction GFP permit to understand the mechanism, and ColA isn't able to have his toxin activity. To protect themselfs, E. coli have an imumnity protein coded by cai gene (-). cal gene is codding for a lysis protein, when this protein is produce ColA can be delivered in the extracellular medium after the E. coli cells lysis. In this contruction RFP permit to understand the mechanism, and Cal isn't able to have his lysis activity. The regulation is complex and involves many regulatory elements: terminator (+) and (-), promoters, antisense coding sequence, etc.
 
  
  

Latest revision as of 00:01, 22 October 2021


Expression of GFP and of the timer device fused to the RFP protein


Introduction  :

The Part BBa_K3788021 is a part composed from the parts BBa_K3788017 (lexA-repressed promoter), BBa_K3788015, BBa_E1010 (RFP) fused to cal gene and BBa_E0040 (GFP) fused to caa gene.

With this design, this part integrates a part of the cal gene, coding for the lysis protein CaL. It is also composed from caa and cai genes, respectively for the Colicin A toxin and the immunity protein Cal.

In normal conditions, caa and cai genes are positioned as an operon, and the cai gene is located on the antisense strand and expressed in a constitutive manner.

This system’s regulation is constituted from many regulation elements, which permits it to obtain a fine-tuned regulation.

This system is located on a plasmid present in some E. Coli populations. It allows Colicin A accumulation on the host cell, and then induces its death due to CaL action, which provokes Colicin A liberation on the extracellular medium, which induces the death of bacteria that don’t produce Cal protein.

cal gene codes for CaL protein, a lipopeptide with a length between 28 and 48 amino acid residues. This protein is produced as a precursor and then processed. Once the protein is processed, it can form pores in the bacterial membrane, which leads to the quasi-lysis of the host cell, and provokes a leak from the cellular component into the medium.


Design :

This construction is used to study the time-shift between ColA and Cal production after induction of the promoter, by measuring the fluorescence kinetics.



Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal NheI site found at 1528
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal BglII site found at 1848
    Illegal BamHI site found at 1521
    Illegal XhoI site found at 1397
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal NgoMIV site found at 1597
    Illegal AgeI site found at 2849
    Illegal AgeI site found at 2961
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI.rc site found at 867
    Illegal SapI.rc site found at 1351