Difference between revisions of "Part:BBa K3482016"
Line 3: Line 3:
 
<partinfo>BBa_K3482016 short</partinfo>
 
<partinfo>BBa_K3482016 short</partinfo>
  
−
This part allows for IPTG inducible expression of the ccdB toxin in E. coli which lead to cell death by acting on the DNA gyrase. the action of ccdB leads to the degradation of DNA , which leads to cell death. It can therefore be induced by IPTG.
+
This part allows for IPTG inducible expression of the CcdB toxin in E. coli which lead to cell death by acting on the DNA gyrase. the action of CcdB leads to the degradation of DNA , which leads to cell death. It can therefore be induced by IPTG.
 +
The CcdB toxin taget the GyrA subunit of the DNA gyrase. The CcdB toxin block the GyrA subunit and leads to a break into the double stranded DNA.
 +
 
 +
This part will be used in a kill-switch system with anohter part containing the antitoxin <html><a style="padding: 0px; margin: 0px;" href="https://parts.igem.org/Part:BBa_K3482020"> BBa_K3482020 </a></html>.
 +
 
 +
===Characterization===
 +
Functional killswitch assay with IPTG and aTc gradients on agar plate
 +
 
 +
[[File:BBa K3482016 kill-switch plate.png|200px|]]
 +
 
 +
E.coli Nissle 1917 were transformed with a plasmid containing the part BBa K3482016 and the part <html><a style="padding: 0px; margin: 0px;" href="https://parts.igem.org/Part:BBa_K3482020"> BBa_K3482020 </a></html> (CcdA antitoxin part) and plated with a gradient of aTc and IPTG on agar plate.
 +
The plate shows a small stimulation/activity of the CcdA antitoxin with aTc induction, also IPTG induction shows induction of the CcdB toxin, and a number of surving cells (probable mutants) proportional to the dilution of the plated culture.
  
−
<!-- Add more about the biology of this part here
 
−
===Usage and Biology===
 
  
 
<!-- -->
 
<!-- -->

Revision as of 15:14, 22 October 2020


IPTG-inducible ccdb toxin

This part allows for IPTG inducible expression of the CcdB toxin in E. coli which lead to cell death by acting on the DNA gyrase. the action of CcdB leads to the degradation of DNA , which leads to cell death. It can therefore be induced by IPTG. The CcdB toxin taget the GyrA subunit of the DNA gyrase. The CcdB toxin block the GyrA subunit and leads to a break into the double stranded DNA.

This part will be used in a kill-switch system with anohter part containing the antitoxin BBa_K3482020 .

Characterization

Functional killswitch assay with IPTG and aTc gradients on agar plate

BBa K3482016 kill-switch plate.png

E.coli Nissle 1917 were transformed with a plasmid containing the part BBa K3482016 and the part BBa_K3482020 (CcdA antitoxin part) and plated with a gradient of aTc and IPTG on agar plate. The plate shows a small stimulation/activity of the CcdA antitoxin with aTc induction, also IPTG induction shows induction of the CcdB toxin, and a number of surving cells (probable mutants) proportional to the dilution of the plated culture.


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
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI site found at 307