Difference between revisions of "Part:BBa K4307008"
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===Usage and Biology=== | ===Usage and Biology=== | ||
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| + | <span class='h3bb'>Sequence and Features</span> | ||
| + | <partinfo>BBa_K4307008 SequenceAndFeatures</partinfo> | ||
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| + | <!-- Uncomment this to enable Functional Parameter display | ||
| + | ===Functional Parameters=== | ||
| + | <partinfo>BBa_K4307000 parameters</partinfo> | ||
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<h2>Characterization</h2> | <h2>Characterization</h2> | ||
| − | <p>The following figure demonstrates our successful construction.</p > | + | <p>The following figure demonstrates our successful construction.</p> |
<br> | <br> | ||
| − | + | https://static.igem.wiki/teams/4307/wiki/wiki-parts-files/mainpage/bba-k4307008-gel.png<br> | |
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<b>Figure 1: </b> <b>The construction results of NisR.</b> | <b>Figure 1: </b> <b>The construction results of NisR.</b> | ||
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<h3>SDS-PAGE assay was done to characterize the biobrick.</h3> | <h3>SDS-PAGE assay was done to characterize the biobrick.</h3> | ||
| − | <p>To determine the expression of NisR, we used T7 promoter and lac operator to control the overexpression of NisR by IPTG. NisR in bacterial lysate was detected by SDS-PAGE(Figure 2), indicating that NisR can be successfully expressed in <i>E.coli</i>. </p > | + | <p>To determine the expression of NisR, we used T7 promoter and lac operator to control the overexpression of NisR by IPTG. NisR in bacterial lysate was detected by SDS-PAGE(Figure 2), indicating that NisR can be successfully expressed in <i>E.coli</i>. </p> |
<p> | <p> | ||
| − | <br> | + | <br>https://static.igem.wiki/teams/4307/wiki/wiki-parts-files/mainpage/bba-k4307008.png<br> |
| − | https://static.igem.wiki/teams/4307/wiki/wiki-parts-files/mainpage/bba-k4307008.png<br> | + | |
<b>Figure 2: </b> <b> SDS-PAGE result of NisR expression.</b> | <b>Figure 2: </b> <b> SDS-PAGE result of NisR expression.</b> | ||
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| − | </p > | + | |
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<h2>Conclusion</h2> | <h2>Conclusion</h2> | ||
| − | <p>According to the SDS-PAGE result, it can be found that NisR can be successfully expressed in <i>E.coli</i>, which indicates that it can act as the downstream regulator of NisK to pass down nisin signal functionally in <i>E.coli</i> as in <i>Lactococcus lactis</i>. This ensures the effectiveness of nisin TCS in <i>E.coli</i>.</p | + | <p>According to the SDS-PAGE result, it can be found that NisR can be successfully expressed in <i>E.coli</i>, which indicates that it can act as the downstream regulator of NisK to pass down nisin signal functionally in <i>E.coli</i> as in <i>Lactococcus lactis</i>. This ensures the effectiveness of nisin TCS in <i>E.coli</i>.</p> |
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Revision as of 15:15, 12 October 2022
NisR
Originated from Lactococcus lactis subsp. Lactis, NisR is a member of the two-component regulatory system NisK/NisR involved in the regulation of the biosynthesis of lantibiotic nisin. NisR functions as a regulatory protein which is phosphorylated by NisK in response to environmental nisin signals. We managed to express NisR in E.coli strain.
Functions together with NisK(BBa_K4307007), PnisA(BBa_K4307009), J23100 and EGFP, it forms composite part J23100-nisK-nisR+PnisA-EGFP(BBa_K4307013) to express fluorescence signal induced by nisin.
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000COMPATIBLE WITH RFC[1000]
Characterization
The following figure demonstrates our successful construction.
Figure 1: The construction results of NisR.
SDS-PAGE assay was done to characterize the biobrick.
To determine the expression of NisR, we used T7 promoter and lac operator to control the overexpression of NisR by IPTG. NisR in bacterial lysate was detected by SDS-PAGE(Figure 2), indicating that NisR can be successfully expressed in E.coli.
Figure 2: SDS-PAGE result of NisR expression.
Conclusion
According to the SDS-PAGE result, it can be found that NisR can be successfully expressed in E.coli, which indicates that it can act as the downstream regulator of NisK to pass down nisin signal functionally in E.coli as in Lactococcus lactis. This ensures the effectiveness of nisin TCS in E.coli.