Difference between revisions of "Team:Paris Bettencourt/Notebook/Idli and micro-organisms"
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+ | |||
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+ | <div id="notebookMenu"> | ||
+ | <ul> | ||
+ | <li><a href="#july">July</a></li> | ||
+ | <li><a href="#august">August</a></li> | ||
+ | <li><a href="#september">September</a></li> | ||
+ | </ul> | ||
+ | </div> | ||
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<a name="august" class="anchor"><h1></h1></a> | <a name="august" class="anchor"><h1></h1></a> | ||
− | <h2 class="date | + | <h2 class="date two">August 10th</h2><br><br> |
We did electrocompetent cells of ''Lactobacillus plantarum'' with <a href="https://2015.igem.org/Team:Paris_Bettencourt/Protocols#FSElectrocompetentLactobacillus"> a particular protocol</a> <br> | We did electrocompetent cells of ''Lactobacillus plantarum'' with <a href="https://2015.igem.org/Team:Paris_Bettencourt/Protocols#FSElectrocompetentLactobacillus"> a particular protocol</a> <br> | ||
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<a name="august" class="anchor"><h1></h1></a> | <a name="august" class="anchor"><h1></h1></a> | ||
− | <h2 class="date | + | <h2 class="date two">August 14th</h2><br><br> |
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<a name="august" class="anchor"><h1></h1></a> | <a name="august" class="anchor"><h1></h1></a> | ||
− | <h2 class="date | + | <h2 class="date two">August 19th and 20th</h2><br><br> |
We started an <a href="https://2015.igem.org/Team:Paris_Bettencourt/Protocols#FSIdli">idli</a> and We put for the fermentation Sc SK1, ScBY4743 mCherry, Sc SK1 with PHO 80 avoid by geneticinR, Sc SK1 with PHO 85 avoid by geneticinR and Sc SK1 with PHO 85 avoid by FRT-geneticinR-FRT. <br> | We started an <a href="https://2015.igem.org/Team:Paris_Bettencourt/Protocols#FSIdli">idli</a> and We put for the fermentation Sc SK1, ScBY4743 mCherry, Sc SK1 with PHO 80 avoid by geneticinR, Sc SK1 with PHO 85 avoid by geneticinR and Sc SK1 with PHO 85 avoid by FRT-geneticinR-FRT. <br> | ||
− | After the right time of fermentation, we took samples from each idli in each conditions (different µorganisms) from 3 diffenrent area of the idli, and we used the <a href="https://2015.igem.org/Team:Paris_Bettencourt/Protocols#FSTitrationacidphytic">Phytic acid assay kit</a> on the samples. | + | After the right time of fermentation, we took samples from each idli in each conditions (different µorganisms) from 3 diffenrent area of the idli, and we used the <a href="https://2015.igem.org/Team:Paris_Bettencourt/Protocols#FSTitrationacidphytic">Phytic acid assay kit</a> on the samples.<br> |
+ | <br> | ||
+ | The analysis of the phyticacid with the kit said us that ther are no phytic acid in the samples. (cf. <a href="https://2015.igem.org/Team:Paris_Bettencourt/Notebook/Phytase">Phytase Notebook</a>) | ||
− | <!-- | + | <!-- 20 Août --> |
− | < | + | <a name="august" class="anchor"><h1></h1></a> |
+ | <h2 class="date two">August 20th</h2><br><br> | ||
− | <!-- --> | + | We did again the experiment where we added 10<sup>8</sup> cells for the fermentation step of the idli recipe. We used <i>S. cerevisiae</i> BY4743 mCherry and <i>S. cerevisiae</i> SK1 with the gene PHO80 or PHO85 were avoid, <i>Lactobacillus plantarum</i> NC8, and <i>Lactococcus lactis</i> MG1363. With a plate counter, we obtained this numbers of colonies in function of the dilution for the plating. <br> |
+ | From the media in the top of the idli :<br> | ||
+ | |||
+ | <table> | ||
+ | <tr> <td><b>Media</b></td><td><b>Dilutions</b> </td><td> <b>Number of colonies of Yeast </b></td> </tr> | ||
+ | <tr> <td>YPD+gen</td> <td>10<sup>-1</sup> </td><td> 79</td> </tr> | ||
+ | <tr> <td>YPD+gen</td> <td> 10<sup>-2</sup> </td><td> 32 </td> </tr> | ||
+ | <tr> <td>YPD+gen</td> <td> 10<sup>-3</sup> </td><td> 10 </td> </tr> | ||
+ | <tr> <td>MRS</td> <td> 10<sup>-1</sup> </td><td> 1 </td> </tr> | ||
+ | <tr> <td>MRS</td> <td> 10<sup>-2</sup> </td><td> 0 </td> </tr> | ||
+ | <tr> <td>MRS</td> <td> 10<sup>-3</sup> </td><td> 0 </td> </tr> | ||
+ | <tr> <td>LB</td> <td> 10<sup>-1</sup> </td><td> 0 </td> </tr> | ||
+ | <tr> <td>LB</td> <td> 10<sup>-2</sup> </td><td> 0 </td> </tr> | ||
+ | <tr> <td>LB</td> <td> 10<sup>-3</sup> </td><td> 1 </td> </tr> | ||
+ | <tr> <td>M17</td> <td> 10<sup>-1</sup> </td><td> 0 </td> </tr> | ||
+ | <tr> <td>M17</td> <td> 10<sup>-2</sup> </td><td> 0</td> </tr> | ||
+ | <tr> <td>M17</td> <td> 10<sup>-3</sup> </td><td> 0 </td> </tr> | ||
+ | </table> | ||
+ | <br> | ||
+ | |||
+ | <table> | ||
+ | <tr> <td><b>Media</b></td><td><b>Dilutions</b> </td><td> <b>Number of colonies of <i>L. Plantarum</i>/<i>L. Lactis</i> </b></td> </tr> | ||
+ | <tr> <td>YPD+gen</td> <td>10<sup>-1</sup> </td><td> 0</td> </tr> | ||
+ | <tr> <td>YPD+gen</td> <td> 10<sup>-2</sup> </td><td> 0 </td> </tr> | ||
+ | <tr> <td>YPD+gen</td> <td> 10<sup>-3</sup> </td><td> 0 </td> </tr> | ||
+ | <tr> <td>MRS</td> <td> 10<sup>-1</sup> </td><td> 465 </td> </tr> | ||
+ | <tr> <td>MRS</td> <td> 10<sup>-2</sup> </td><td> 99 </td> </tr> | ||
+ | <tr> <td>MRS</td> <td> 10<sup>-3</sup> </td><td> 6 </td> </tr> | ||
+ | <tr> <td>LB</td> <td> 10<sup>-1</sup> </td><td> 96 </td> </tr> | ||
+ | <tr> <td>LB</td> <td> 10<sup>-2</sup> </td><td> 51 </td> </tr> | ||
+ | <tr> <td>LB</td> <td> 10<sup>-3</sup> </td><td> 15 </td> </tr> | ||
+ | <tr> <td>M17</td> <td> 10<sup>-1</sup> </td><td> 123 </td> </tr> | ||
+ | <tr> <td>M17</td> <td> 10<sup>-2</sup> </td><td> 3</td> </tr> | ||
+ | <tr> <td>M17</td> <td> 10<sup>-3</sup> </td><td> 7 </td> </tr> | ||
+ | </table> | ||
+ | <br> | ||
+ | |||
+ | From the batter phase of the idli:<br> | ||
+ | |||
+ | |||
+ | <table> | ||
+ | <tr> <td><b>Media</b></td><td><b>Dilutions</b> </td><td> <b>Number of colonies of Yeast </b></td> </tr> | ||
+ | <tr> <td>YPD+gen</td> <td>10<sup>-1</sup> </td><td> 217</td> </tr> | ||
+ | <tr> <td>YPD+gen</td> <td> 10<sup>-2</sup> </td><td> 150 </td> </tr> | ||
+ | <tr> <td>YPD+gen</td> <td> 10<sup>-3</sup> </td><td> 19 </td> </tr> | ||
+ | <tr> <td>MRS</td> <td> 10<sup>-1</sup> </td><td> 6 </td> </tr> | ||
+ | <tr> <td>MRS</td> <td> 10<sup>-2</sup> </td><td> 1 </td> </tr> | ||
+ | <tr> <td>MRS</td> <td> 10<sup>-3</sup> </td><td> 1 </td> </tr> | ||
+ | <tr> <td>LB</td> <td> 10<sup>-1</sup> </td><td> 2 </td> </tr> | ||
+ | <tr> <td>LB</td> <td> 10<sup>-2</sup> </td><td> 0 </td> </tr> | ||
+ | <tr> <td>LB</td> <td> 10<sup>-3</sup> </td><td> 0 </td> </tr> | ||
+ | <tr> <td>M17</td> <td> 10<sup>-1</sup> </td><td> 0 </td> </tr> | ||
+ | <tr> <td>M17</td> <td> 10<sup>-2</sup> </td><td> 0</td> </tr> | ||
+ | <tr> <td>M17</td> <td> 10<sup>-3</sup> </td><td> 0 </td> </tr> | ||
+ | </table> | ||
+ | <br> | ||
+ | |||
+ | <table> | ||
+ | <tr> <td><b>Media</b></td><td><b>Dilutions</b> </td><td> <b>Number of colonies of <i>L. Plantarum</i>/<i>L. Lactis</i> </b></td> </tr> | ||
+ | <tr> <td>YPD+gen</td> <td>10<sup>-1</sup> </td><td> 0</td> </tr> | ||
+ | <tr> <td>YPD+gen</td> <td> 10<sup>-2</sup> </td><td> 0 </td> </tr> | ||
+ | <tr> <td>YPD+gen</td> <td> 10<sup>-3</sup> </td><td> 0 </td> </tr> | ||
+ | <tr> <td>MRS</td> <td> 10<sup>-1</sup> </td><td> 396 </td> </tr> | ||
+ | <tr> <td>MRS</td> <td> 10<sup>-2</sup> </td><td> 51 </td> </tr> | ||
+ | <tr> <td>MRS</td> <td> 10<sup>-3</sup> </td><td> 6 </td> </tr> | ||
+ | <tr> <td>LB</td> <td> 10<sup>-1</sup> </td><td> 125 </td> </tr> | ||
+ | <tr> <td>LB</td> <td> 10<sup>-2</sup> </td><td> 13 </td> </tr> | ||
+ | <tr> <td>LB</td> <td> 10<sup>-3</sup> </td><td> 8 </td> </tr> | ||
+ | <tr> <td>M17</td> <td> 10<sup>-1</sup> </td><td> 503 </td> </tr> | ||
+ | <tr> <td>M17</td> <td> 10<sup>-2</sup> </td><td> 111</td> </tr> | ||
+ | <tr> <td>M17</td> <td> 10<sup>-3</sup> </td><td> 29 </td> </tr> | ||
+ | </table> | ||
+ | <br> | ||
+ | |||
+ | We osberved that the µorganisms was in higher number near in the batter of the idli than in the idli water or in the grind rice and dall phase, where nothing has grew. | ||
+ | |||
+ | <br><br> | ||
+ | <!-- 27 Août --> | ||
+ | |||
+ | <a name="august" class="anchor"><h1></h1></a> | ||
+ | <h2 class="date two">August 27th</h2><br><br> | ||
+ | |||
+ | |||
+ | We did idli with <i>S. cerevisiae</i> SK1 where PHO85 were avoid by a genetic system composed by two FRT sequences and the gene of the geneticin resistance. And also, the <i>S. cerevisiae</i> SK1 PHO80 avoid, the <i>S. cerevisiae</i> SK1 PHO85 avoid, Sc BY4743 mCherry and the first <i>S. cerevisiae</i> SK1 together with the <i>S. cerevisiae</i> with the mCherry gene. <br> | ||
+ | To plate on YPD and geneticin (at 200 µg.µl<sup>-1</sup>), We took the sample from 3 area of the idli : top, middle and bottom of the idli, and we did 3 dilutions (from 10<sup>-1</sup> to 10<sup>-3</sup>). Below, we can see the picture of each plates for each conditions of dilution, area and yeast strains that we used.<br><br> | ||
+ | |||
+ | <div class="column-right"align="center"> | ||
+ | <IMG SRC= "https://static.igem.org/mediawiki/2015/b/b0/Tube1%262.png” width=50%> | ||
+ | </div><br> | ||
+ | |||
+ | <div class="column-left"align="center"> | ||
+ | <IMG SRC= "https://static.igem.org/mediawiki/2015/a/a4/Tube3%264.png” width=50%> | ||
+ | </div> | ||
+ | <br><br> | ||
+ | |||
+ | <div align="left"> | ||
+ | <IMG SRC= "https://static.igem.org/mediawiki/2015/e/e6/Tube5.png" width=50%> | ||
+ | </div><br> | ||
+ | |||
+ | |||
+ | The number of yeast was around the same for each strains, and we obtained the next table, where we did a mean of the yeast that we counted, with a plate counter. | ||
+ | |||
+ | <table> | ||
+ | <tr> <td><b>Area</b></td><td><b>Dilutions</b> </td><td> <b>Number of colonies of Yeast </b></td> </tr> | ||
+ | <tr> <td>Top</td> <td>10<sup>-1</sup> </td><td> 285</td> </tr> | ||
+ | <tr> <td>Top</td> <td> 10<sup>-2</sup> </td><td> 11 </td> </tr> | ||
+ | <tr> <td>Top</td> <td> 10<sup>-3</sup> </td><td> 2 </td> </tr> | ||
+ | <tr> <td>Middle</td> <td> 10<sup>-1</sup> </td><td> 362 </td> </tr> | ||
+ | <tr> <td>Middle</td> <td> 10<sup>-2</sup> </td><td> 15 </td> </tr> | ||
+ | <tr> <td>Middle</td> <td> 10<sup>-3</sup> </td><td> 2 </td> </tr> | ||
+ | <tr> <td>Bottom</td> <td> 10<sup>-1</sup> </td><td> 357 </td> </tr> | ||
+ | <tr> <td>Bottom</td> <td> 10<sup>-2</sup> </td><td> 16 </td> </tr> | ||
+ | <tr> <td>Bottom</td> <td> 10<sup>-3</sup> </td><td> 1 </td> </tr> | ||
+ | </table> | ||
+ | <br> | ||
+ | |||
+ | |||
+ | |||
+ | <!-- 10 Septembre --> | ||
+ | |||
+ | <a name="september" class="anchor"><h1></h1></a> | ||
+ | <h2 class="date three">September 10th</h2><br><br> | ||
+ | |||
+ | We started, using a TECAN, to find any informations on our strains that we want to use in fine. We did in a first time an experiment at 30°C, with <i>S. cerevisiae</i> yPH151, <i>S. cerevisiae</i> SK1 PHO 80 deleted, <i>S. cerevisiae</i> SK1 PHO 85 deleted, <i>S. cerevisiae</i> SK1 PHO 80 and PHO 85 deleted, and <i>Propioni f.</i> on 8 different media (Water, Idli batter/100, Idli water, M17, MRS, URA3free media, YPD and LB. We obtained this growth curve : <br> | ||
+ | |||
+ | <div class="column-left"align="center"> | ||
+ | <IMG SRC= "https://static.igem.org/mediawiki/2015/b/be/Mean_Control_10sep.png" width=50%> | ||
+ | </div> | ||
+ | <div class="column-right"align="center"> | ||
+ | <IMG SRC= "https://static.igem.org/mediawiki/2015/6/6d/Mean_PHO80_10sep.png" width=50%> | ||
+ | </div><br> | ||
+ | <div class="column-left"align="center"> | ||
+ | <IMG SRC= "https://static.igem.org/mediawiki/2015/4/42/Mean_PHO85_10sep.png" width=50%> | ||
+ | </div> | ||
+ | <div class="column-right"align="center"> | ||
+ | <IMG SRC= "https://static.igem.org/mediawiki/2015/0/0a/Mean_PHOavoid_10sep.png" width=50%> | ||
+ | </div><br> | ||
+ | <div class="column-left"align="center"> | ||
+ | <IMG SRC= "https://static.igem.org/mediawiki/2015/c/ca/Mean_Propioni_10sep.png" width=50%> | ||
+ | </div> | ||
+ | <div class="column-right"align="center"> | ||
+ | <IMG SRC= "https://static.igem.org/mediawiki/2015/f/fd/Sc_yPH151_10sep.png" width=50%> | ||
+ | </div><br><br> | ||
+ | |||
+ | And particulary, the growth curve of the strains on idli water :<br> | ||
+ | <IMG SRC= "https://static.igem.org/mediawiki/2015/9/97/Mean_Idliwater_10sep.png" width=50%> | ||
+ | <br><br> | ||
+ | |||
+ | <!-- 11 Septembre --> | ||
+ | |||
+ | <a name="september" class="anchor"><h1></h1></a> | ||
+ | <h2 class="date three">September 11th</h2><br><br> | ||
+ | |||
+ | We did in a first time an experiment at 37°C, with <i>E. coli</i> who product vitamin B2, <i>Propioni f.</i> who product vitamin B12 and <i>Lactobacillus plantarum</i> NC8 who should have obtained the vitamin B2 pathway that we engineered. | ||
+ | |||
+ | <div class="column-left"align="center"> | ||
+ | <IMG SRC= "https://static.igem.org/mediawiki/2015/d/df/Mean_Control_11sep.png" width=50%> | ||
+ | </div> | ||
+ | <div class="column-right"align="center"> | ||
+ | <IMG SRC= "https://static.igem.org/mediawiki/2015/9/9f/Mean_EcoliB2_11sep.png" width=50%> | ||
+ | </div><br> | ||
+ | <div class="column-left"align="center"> | ||
+ | <IMG SRC= "https://static.igem.org/mediawiki/2015/6/6b/Mean_Lactobacillus_11sep.png" width=50%> | ||
+ | </div> | ||
+ | <div class="column-right"align="center"> | ||
+ | <IMG SRC= "https://static.igem.org/mediawiki/2015/6/6c/Mean_Propionif_11sep.png" width=50%> | ||
+ | </div><br><br> | ||
+ | |||
+ | And particulary, the growth curve of the strains on idli water :<br> | ||
+ | <IMG SRC= "https://static.igem.org/mediawiki/2015/3/3a/Mean_Idliwater_11sep.png" width=50%> | ||
+ | |||
+ | <br><br> | ||
+ | With this two last experiments, we can say that the yeasts and the bacteria that we engineered, can grow in idli water, and with the data coming from the idli fermentation with our micro-organisms, it suggests that our micro-organisms grow well in idli in the higher phase of the idli. | ||
+ | <br> |
Latest revision as of 18:51, 20 November 2015