Difference between revisions of "Part:BBa K4307023"

 
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This part is derived from PnisA, which is originated from <i>Lactococcus lactis subsp. Lactis</i>, by the improvement done by our own software tool to enhance the promoter.
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<span class='h3bb'>Sequence and Features</span>
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<h2>Sequence and Features</h2>
 
<partinfo>BBa_K4307023 SequenceAndFeatures</partinfo>
 
<partinfo>BBa_K4307023 SequenceAndFeatures</partinfo>
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===Functional Parameters===
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<partinfo>BBa_K4307000 parameters</partinfo>
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<h2>Characterization </h2>
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<p>The following figure demonstrates our successful construction.</p>
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<br>
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<div class="center">
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    <div class="thumb tnone">
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        <div class="thumbinner" style="width:50%;">
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            <a href="https://static.igem.wiki/teams/4307/wiki/wiki-parts-files/mainpage/bba-k4307023-gel.png" class="image">
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                <img alt="" src="https://static.igem.wiki/teams/4307/wiki/wiki-parts-files/mainpage/bba-k4307023-gel.png" width="100%" height=auto class="thumbimage" /></a>                  <div class="thumbcaption">
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                <div class="magnify">
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                    <a href="https://static.igem.wiki/teams/4307/wiki/wiki-parts-files/mainpage/bba-k4307023-gel.png" class="internal" title="Enlarge"></a>
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                </div>
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                <b>Figure 1: </b> <b>The construction results of PnisRRH09.</b>
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            </div>
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        </div>
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    </div>
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</div>
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<h3>Fluorescence assay was done to characterize the bio brick.    </h3>
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<p>We introduced these 10 promoters into our system into MACH1-T1 strain to detect EGFP fluorescence signal with and without nisin induction by microplate reader. Results demonstrated that fluorescence signals were enhanced both with and without nisin induction except for PnisRRH07 and PnisRRH10(Figure 2).  </p>
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<p>
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  <br>
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<div class="center">
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        <div class="thumbinner" style="width:50%;">
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            <a href="https://static.igem.wiki/teams/4307/wiki/wiki-parts-files/mainpage/bba-k4307023-1.png" class="image">
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                <img alt="" src="https://static.igem.wiki/teams/4307/wiki/wiki-parts-files/mainpage/bba-k4307023-1.png" width="100%" height=auto class="thumbimage" /></a>                  <div class="thumbcaption">
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                <div class="magnify">
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                    <a href="https://static.igem.wiki/teams/4307/wiki/wiki-parts-files/mainpage/bba-k4307023-1.png" class="internal" title="Enlarge"></a>
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                </div>
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                <b>Figure 2: </b> <b> Fluorescence spectrophotometry results of the 10 mutant promoters with the highest predictive strength.</b>
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            </div>
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        </div>
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    </div>
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<br>
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Furthermore, flow cytometry results confirmed that percentages of positive fluorescence signal bacteria also increased both with and without nisin induction(Figure 3), indicating that the improvement of promoter PnisA by our software tool was practical.
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<br>
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<div class="center">
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    <div class="thumb tnone">
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        <div class="thumbinner" style="width:50%;">
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            <a href="https://static.igem.wiki/teams/4307/wiki/wiki-parts-files/mainpage/bba-k4307023-2.png" class="image">
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                <img alt="" src="https://static.igem.wiki/teams/4307/wiki/wiki-parts-files/mainpage/bba-k4307015-2.png" width="100%" height=auto class="thumbimage" /></a>                  <div class="thumbcaption">
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                <div class="magnify">
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                    <a href="https://static.igem.wiki/teams/4307/wiki/wiki-parts-files/mainpage/bba-k4307023-2.png" class="internal" title="Enlarge"></a>
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                </div>
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                <b>Figure 3: </b> <b> Flow cytometry results of the 10 mutant promoters with the highest predictive strength. </b>
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            </div>
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        </div>
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    </div>
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</div>
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<br>
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</p>
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<h2>Conclusion</h2>
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<p>According to fluorescence assay result, it can be found that the improvement of promoter PnisA by our software tool is a great success, though the effectiveness of PnisRRH07 and PnisRRH10 is relatively low. This helps us a lot in providing an improve to the composite part J23100-nisK-nisR+PnisA-EGFP(BBa_K4307045) and create better nisin induced expression system in <i>E.coli</i>.</p>
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Latest revision as of 16:44, 13 October 2022


PnisRRH09

This part is derived from PnisA, which is originated from Lactococcus lactis subsp. Lactis, by the improvement done by our own software tool to enhance the promoter.


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]



Characterization

The following figure demonstrates our successful construction.


Figure 1: The construction results of PnisRRH09.

Fluorescence assay was done to characterize the bio brick.

We introduced these 10 promoters into our system into MACH1-T1 strain to detect EGFP fluorescence signal with and without nisin induction by microplate reader. Results demonstrated that fluorescence signals were enhanced both with and without nisin induction except for PnisRRH07 and PnisRRH10(Figure 2).


Figure 2: Fluorescence spectrophotometry results of the 10 mutant promoters with the highest predictive strength.

Furthermore, flow cytometry results confirmed that percentages of positive fluorescence signal bacteria also increased both with and without nisin induction(Figure 3), indicating that the improvement of promoter PnisA by our software tool was practical.
Figure 3: Flow cytometry results of the 10 mutant promoters with the highest predictive strength.

Conclusion

According to fluorescence assay result, it can be found that the improvement of promoter PnisA by our software tool is a great success, though the effectiveness of PnisRRH07 and PnisRRH10 is relatively low. This helps us a lot in providing an improve to the composite part J23100-nisK-nisR+PnisA-EGFP(BBa_K4307045) and create better nisin induced expression system in E.coli.