Difference between revisions of "Part:BBa K2610031"
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<partinfo>BBa_K2610031 short</partinfo>
 
<partinfo>BBa_K2610031 short</partinfo>
  
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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.  
  
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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===
  
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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.
 
  
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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>  
  
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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 </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