Difference between revisions of "Part:BBa K2610035"

 
 
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<partinfo>BBa_K2610035 short</partinfo>
 
<partinfo>BBa_K2610035 short</partinfo>
  
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This composite part features twice the regulatory part promoter SoxS (BBa_K2610030) and two times 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 twice the regulatory part promoter SoxS ([https://parts.igem.org/Part:BBa_K2610030 BBa_K2610030]) and two times the fluorescent protein GFP ([https://parts.igem.org/Part:BBa_E0040 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 superoxide pathway in Escherichia coli. It acts a superactivator of downstream stress genes.
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Regulatory protein SoxS is involved in the oxidative stress signaling pathway in <i> Escherichia coli </i>. 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===
 
===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. We have created this part in order to amplify the fluorescent signal measured after pSoxS-GFP activation.
 
  
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As was demonstrated in Figure 1 pSoxS-GFP-GFP-pSoxS-GFP-GFP responds to nalidixic acid in a dose-dependent response curve.  
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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. We have created this part in order to amplify the fluorescent signal measured after pSoxS-GFP activation.  
  
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[[File:T--Leiden--amplification3.png|750px]]  
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As can be observed in Figure 1 the pSoxS-GFP-pSoxS-GFP construct leads to an increased mean fluorescence intensity compared to pSoxS-GFP ([https://parts.igem.org/Part:BBa_K2610031 BBa_K2610031]), but not compared to pSoXS-GFP-pSoxS-GFP ([https://parts.igem.org/Part:BBa_K2610034 BBa_K2610034]).
  
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<span style="font-size:1em"><b>Figure 1.</b> Dose-response curve as a result of treatment with nalidixic acid. </span>  
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We also assessed the dose-dependency of this reporter. We found that an increase in nalidixic acid concentration of nalidixic acid leads to an increasing mean fluorescence intensity (MFI). A decrease in detected GFP expression at higher concentrations can be attributed to the lethality of the stressor.
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[[File:T--Leiden--amplificA.png|200px]] [[File:T--Leiden--amplificB.jpeg|200px]] [[File:T--Leiden--amplficC.jpeg|200px]]<br><br>
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<span style="font-size:1em"><b>Figure 1.</b> Median fluorescence intensity (MFI) as a result of treatment with nalidixic acid for pSoxS-GFP-GFP-pSoxS-GFP-GFP compared to pSoxS-GFP-pSoxS-GFP and pSoxS-GFP . It can also be seen that there is a dose response curve to nalidixic acid. The decrease in detected GFP expression at higher concentrations can be attributed to lethality of the stressor. </span>  
  
  

Latest revision as of 02:22, 18 October 2018


pSoxS-GFP-GFP-pSoxS-GFP-GFP

This composite part features twice the regulatory part promoter SoxS (BBa_K2610030) and two times 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

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. We have created this part in order to amplify the fluorescent signal measured after pSoxS-GFP activation.

As can be observed in Figure 1 the pSoxS-GFP-pSoxS-GFP construct leads to an increased mean fluorescence intensity compared to pSoxS-GFP (BBa_K2610031), but not compared to pSoXS-GFP-pSoxS-GFP (BBa_K2610034).

We also assessed the dose-dependency of this reporter. We found that an increase in nalidixic acid concentration of nalidixic acid leads to an increasing mean fluorescence intensity (MFI). A decrease in detected GFP expression at higher concentrations can be attributed to the lethality of the stressor.

T--Leiden--amplificA.png T--Leiden--amplificB.jpeg T--Leiden--amplficC.jpeg

Figure 1. Median fluorescence intensity (MFI) as a result of treatment with nalidixic acid for pSoxS-GFP-GFP-pSoxS-GFP-GFP compared to pSoxS-GFP-pSoxS-GFP and pSoxS-GFP . It can also be seen that there is a dose response curve to nalidixic acid. The decrease in detected GFP expression at higher concentrations can be attributed to lethality of the stressor.


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
    Illegal BsaI.rc site found at 1673
    Illegal BsaI.rc site found at 2704
    Illegal BsaI.rc site found at 3430