Difference between revisions of "Part:BBa K4137000"
 
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<partinfo>BBa_K4137000 short</partinfo>
 
<partinfo>BBa_K4137000 short</partinfo>
  
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BBa_K4137000, nicknamed ccdB for the purpose of our project, encodes for the production of CcdB protein which is a toxin to many bacteria. CcdB loses its toxicity by binding with CcdA, which prevents it from binding and interact with DNA Gyrase. The Myc tag was added to the CcdB coding sequence during the part design, this allows us to characterize the protein and go through purification.  
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BBa_K4137000 codes for the CcdA protein, a component of the CcdA-CcdB toxin-antitoxin system. CcdA binds with CcdB which inhibits CcdB from targeting DNA gyrase thus preventing DNA double-strand breaks.  
  
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===Usage and Biology===
 
  
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[[Image:ccda-his.png|800px|thumb|center|Fig.1 ccdA with C-Terminal 6x His-Tag.]]
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===Characterization===
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We obtained the sequence for ccdA from IDT through gblock synthesis. Since we aimed to develop determine how different combinations of our toxin-antitoxin system components (ccdA, ccdB, mleR) affect one another's activity to verify our kill switch theory, we designed our constructs with biobrick restriction sites that bracket our genetic components--to insert one of our components into our backbone pSB1C3, we would cut at EcoRI and SpeI for both the insert and the vector. We also flanked each component with a purification tag to observe their expression levels.
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[[Image:ccda-cut.png|800px|thumb|center|Fig.2 ccdA with E-S cut sites.]]
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===Sequence and Features===
 
<span class='h3bb'>Sequence and Features</span>
 
<span class='h3bb'>Sequence and Features</span>
 
<partinfo>BBa_K4137000 SequenceAndFeatures</partinfo>
 
<partinfo>BBa_K4137000 SequenceAndFeatures</partinfo>
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Latest revision as of 02:54, 10 October 2022


CcdA-6xHis Antitoxin

BBa_K4137000 codes for the CcdA protein, a component of the CcdA-CcdB toxin-antitoxin system. CcdA binds with CcdB which inhibits CcdB from targeting DNA gyrase thus preventing DNA double-strand breaks.


Fig.1 ccdA with C-Terminal 6x His-Tag.

Characterization

We obtained the sequence for ccdA from IDT through gblock synthesis. Since we aimed to develop determine how different combinations of our toxin-antitoxin system components (ccdA, ccdB, mleR) affect one another's activity to verify our kill switch theory, we designed our constructs with biobrick restriction sites that bracket our genetic components--to insert one of our components into our backbone pSB1C3, we would cut at EcoRI and SpeI for both the insert and the vector. We also flanked each component with a purification tag to observe their expression levels.

Fig.2 ccdA with E-S cut sites.

Sequence and Features

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]