Difference between revisions of "Part:BBa K2273002"
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__NOTOC__ | __NOTOC__ | ||
<partinfo>BBa_K2273002 short</partinfo> | <partinfo>BBa_K2273002 short</partinfo> | ||
| + | ===Usage and Biology=== | ||
| + | <html> | ||
| + | <figure> | ||
| + | <figure style="width:50%;float:right;"><img src="https://static.igem.org/mediawiki/2017/6/60/T--TU_Dresden--P_Communication_Figure1.jpeg" width="100%"/> | ||
| + | <figcaption><p align="justify"><b>Figure 1: The regulatory system for competence development in <i><b>Bacillus subtilis</b></i>.</b> The development of genetic competence is stimulated by quorum sensing due to the secretion of the ComX pheromone and its accumulation in the surrounding medium. (Modified from: <a target="_blank" href="https://www.ncbi.nlm.nih.gov/pubmed/22511326">Jung <i>et al.</i></a>)</p></figcation> | ||
| + | </figure> | ||
| + | <p align="justify">This promoter is part of the well-studied regulatory system for competence development in <i>Bacillus subtilis</i> (Figure 1). | ||
| + | <i>B. subtilis</i> constantly secretes the ComX pheromone (<a href="https://parts.igem.org/Part:BBa_K2273003">BBa_K2273003</a>), a 9- to 10-amino acid oligopeptide, as a signalling molecule <b>(a)</b>. By rising cell-density, the ComX-concentration in the surrounding medium increases until it reaches a threshold and activates ComP, a membrane-spanning protein kinase <b>(b)</b>. This mechanism is called quorum sensing. | ||
| + | The kinase reacts to the accumulation of ComX by phosphorylating the response regulator ComA <b>(c)</b> which then works as a transcription factor by binding to several promoters and enhancing their activity <b>(d)</b>. | ||
| + | </p> | ||
| + | <p align="justify"> | ||
| + | The most important promoter regulated by ComA and involved in competence development is the promoter of the <i>srfA</i> operon (P<sub><i>srfA</i></sub>). This operon contains not only genes for the production of the antibiotic surfactin <b>(e)</b> but also for ComS <b>(f)</b>, another small peptide that prevents the degradation of the autoregulated transcription factor ComK <b>(g)</b>. <br>Since the promoter is not only dependent on quorum sensing but also on nutritional conditions, we evaluated the influence of different media (Figure 2). We tested full medium (LB) as well as minimal media (MNGE, MCSE).</p> | ||
| − | + | </figure> | |
| − | < | + | <figure style="padding-right:15%; padding-left:15%;"> |
| − | === | + | |
| + | <img src="https://static.igem.org/mediawiki/parts/1/11/T--TU_Dresden--P_Communication_Psrf_Media.png" width="100%"/> | ||
| + | <figcaption><p align="justify"><b>Figure 2: Growth (A) and promoter activity of P<sub><i><b>srfA</b></i></sub> (B) in different media.</b> Wild type (WT, pink) and <i>comX</i>-knockout strains (Δ<i>comX</i>, dark blue) contain a P<sub><i>srfA</i></sub>-<i>luxABCDE</i> fusion.</p></figcation> | ||
| + | </figure> | ||
| + | |||
| + | <p align="justify">Furthermore, we studied the influence of ComX-overproduction on the promoter activity by integrating an inducible copy of <i>comX</i>.</p> | ||
| + | |||
| + | <figure style="padding-left:35%; padding-right:35%;"><img src="https://static.igem.org/mediawiki/parts/a/a3/T--TU_Dresden--P_Communication_Psrf_Induction.png" width="100%"/><figcaption><p align="justify"><b>Growth (A) and promoter activity of P<sub><i><b>srfA</b></i></sub> (B) in dependence of xylose-induction.</b> Wild type (WT, pink and light blue) and <i>comX</i>-knockout strain (Δ<i>comX</i>, black and dark blue) contain a P<sub><i>srfA</i></sub>-<i>luxABCDE</i> fusion. After 1 hour of incubation (dashed line) the cultures were induced with 1% xylose (light blue and black) or with distilled water (dH2O, pink and dark blue).</p></figcation> | ||
| + | </figure> | ||
| + | |||
| + | |||
| + | |||
| + | <p align="justify"> | ||
| + | <b>The promoter was created and used by <a target="_blank" href="http://2017.igem.org/Team:TU_Dresden/Project/Communication">TU_Dresden 2017</a>.</b> All assays have been conducted with <i>B. subtilis</i> W168 carrying the promoter fused to the <i>lux</i> operon. | ||
| + | Other promoters that haven been evaluated regarding their ComX-dependent activity: | ||
| + | <table> | ||
| + | <tr><td>P<sub><i>srfA</i></sub></td><td><a href="https://parts.igem.org/Part:BBa_K2273000">BBa_K2273000</a></td></tr> | ||
| + | <tr><td>P<sub><i>rapA</i></sub></td><td><a href="https://parts.igem.org/Part:BBa_K2273001">BBa_K2273001</a></td></tr> | ||
| + | <tr><td>P<sub><i>rapF</i></sub></td><td><a href="https://parts.igem.org/Part:BBa_K2273002">BBa_K2273002</a></td></tr> | ||
| + | <tr><td>P<sub><i>comG</i></sub></td><td><a href="https://parts.igem.org/Part:BBa_K2273012">BBa_K2273012</a></td></tr> | ||
| + | <tr><td>P<sub><i>comK</i></sub> mut</td><td><a href="https://parts.igem.org/Part:BBa_K2273013">BBa_K2273013</a></td></tr> | ||
| + | </table> | ||
| + | <br></p> | ||
| + | </html> | ||
| − | |||
<span class='h3bb'>Sequence and Features</span> | <span class='h3bb'>Sequence and Features</span> | ||
<partinfo>BBa_K2273002 SequenceAndFeatures</partinfo> | <partinfo>BBa_K2273002 SequenceAndFeatures</partinfo> | ||
Revision as of 13:05, 31 October 2017
PrapF: a ComA-dependent promoter of Bacillus subtilis
Usage and Biology
Figure 1: The regulatory system for competence development in Bacillus subtilis. The development of genetic competence is stimulated by quorum sensing due to the secretion of the ComX pheromone and its accumulation in the surrounding medium. (Modified from: Jung et al.) This promoter is part of the well-studied regulatory system for competence development in Bacillus subtilis (Figure 1).
B. subtilis constantly secretes the ComX pheromone (BBa_K2273003), a 9- to 10-amino acid oligopeptide, as a signalling molecule (a). By rising cell-density, the ComX-concentration in the surrounding medium increases until it reaches a threshold and activates ComP, a membrane-spanning protein kinase (b). This mechanism is called quorum sensing.
The kinase reacts to the accumulation of ComX by phosphorylating the response regulator ComA (c) which then works as a transcription factor by binding to several promoters and enhancing their activity (d).
The most important promoter regulated by ComA and involved in competence development is the promoter of the srfA operon (PsrfA). This operon contains not only genes for the production of the antibiotic surfactin (e) but also for ComS (f), another small peptide that prevents the degradation of the autoregulated transcription factor ComK (g). Figure 2: Growth (A) and promoter activity of PsrfA (B) in different media. Wild type (WT, pink) and comX-knockout strains (ΔcomX, dark blue) contain a PsrfA-luxABCDE fusion.
Since the promoter is not only dependent on quorum sensing but also on nutritional conditions, we evaluated the influence of different media (Figure 2). We tested full medium (LB) as well as minimal media (MNGE, MCSE).
Furthermore, we studied the influence of ComX-overproduction on the promoter activity by integrating an inducible copy of comX.
Growth (A) and promoter activity of PsrfA (B) in dependence of xylose-induction. Wild type (WT, pink and light blue) and comX-knockout strain (ΔcomX, black and dark blue) contain a PsrfA-luxABCDE fusion. After 1 hour of incubation (dashed line) the cultures were induced with 1% xylose (light blue and black) or with distilled water (dH2O, pink and dark blue).
The promoter was created and used by TU_Dresden 2017. All assays have been conducted with B. subtilis W168 carrying the promoter fused to the lux operon. Other promoters that haven been evaluated regarding their ComX-dependent activity:
| PsrfA | BBa_K2273000 |
| PrapA | BBa_K2273001 |
| PrapF | BBa_K2273002 |
| PcomG | BBa_K2273012 |
| PcomK mut | BBa_K2273013 |
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]