Difference between revisions of "Part:BBa K2610031"

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<partinfo>BBa_K2610031 short</partinfo>
 
<partinfo>BBa_K2610031 short</partinfo>
  
This composite part features the regulatory part promoter SoxS (BBa_K2610030) and the fluorescent protein GFP (BBa_E0040). It can be used to visualize upregulation of SoxS as a result of superoxide stress.  
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This composite part features the regulatory part promoter pSoxS (BBa_K2610030) and the fluorescent protein GFP (BBa_E0040). It can be used to visualize upregulation of SoxS as a result of superoxide stress.
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Regulatory protein SoxS is involved in the oxidative stress signaling pathway in Escherichia coli. Intracellular superoxide-generating compounds cause the SoxR to activate transcription of SoxS, which then triggers a set of defense and repair genes that form the oxidative response system.  
  
Regulatory protein SoxS is involved in the superoxide pathway in Escherichia coli. It acts a superactivator of downstream stress genes.
 
  
 
===Usage and Biology===
 
===Usage and Biology===
  
We, iGEM Leiden 2018, have designed this composite part as part of our project Fifty Shades of Stress. This reporter part allowed us to detect stress-induced changes in SoxS transcription using flow cytometry.
 
  
As can be seen in figure 1, treatment with increasing concentrations of nalidixic acid leads to an increasing median fluorescence. Moreover, we have integrated this promoter into a heatmap with many stress-related promoters (Figure 2). SoxS is activated specifically by nalidixic acid and hydrogen peroxide.
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iGEM Leiden 2018 has created a screening system for the detection of new antibiotics. This composite part which allowed us to detect stress-induced changes in SoxS transcription, signaling that a certain compound causes superoxide stress.
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We have tested of these several stress-activated promoters against a repertoire of known antibiotics using flow cytometry.  As can be seen in the heatmap below, treatment with both nalidixic acid and hydrogen peroxide causes a significant increase in fluorescent signal of pSoxS-GFP.
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[[File:T--Leiden--heatmap.png|750px]] <br>
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<span style="font-size:1em"><b>Figure 2.</b> Heatmap with stress-activated promoters. Values for each promoter are normalised to its respective negative control (no stressor, not shown). </span>
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[[File:T--Leiden--SoxS-GFP1.png|750px]] <br>
 
[[File:T--Leiden--SoxS-GFP1.png|750px]] <br>
 
<span style="font-size:1em"><b>Figure 1.</b> Median Fluorescence Intensity (MFI) in AU after 4 hour incubation with nalidixic acid in various concentrations. </span>  
 
<span style="font-size:1em"><b>Figure 1.</b> Median Fluorescence Intensity (MFI) in AU after 4 hour incubation with nalidixic acid in various concentrations. </span>  
  
[[File:T--Leiden--heatmap.png|750px]] <br>
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<span style="font-size:1em"><b>Figure 2.</b> Heatmap with stress-activated promoters </span>
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<!-- Add more about the biology of this part here
 
<!-- Add more about the biology of this part here

Revision as of 23:36, 16 October 2018


pSoxS-GFP

This composite part features the regulatory part promoter pSoxS (BBa_K2610030) and the fluorescent protein GFP (BBa_E0040). It can be used to visualize upregulation of SoxS as a result of superoxide stress.

Regulatory protein SoxS is involved in the oxidative stress signaling pathway in Escherichia coli. Intracellular superoxide-generating compounds cause the SoxR to activate transcription of SoxS, which then triggers a set of defense and repair genes that form the oxidative response system.


Usage and Biology

iGEM Leiden 2018 has created a screening system for the detection of new antibiotics. This composite part which allowed us to detect stress-induced changes in SoxS transcription, signaling that a certain compound causes superoxide stress.

We have tested of these several stress-activated promoters against a repertoire of known antibiotics using flow cytometry. As can be seen in the heatmap below, treatment with both nalidixic acid and hydrogen peroxide causes a significant increase in fluorescent signal of pSoxS-GFP.

T--Leiden--heatmap.png
Figure 2. Heatmap with stress-activated promoters. Values for each promoter are normalised to its respective negative control (no stressor, not shown).


T--Leiden--SoxS-GFP1.png
Figure 1. Median Fluorescence Intensity (MFI) in AU after 4 hour incubation with nalidixic acid in various concentrations.


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.rc site found at 947