Difference between revisions of "Team:Paris Bettencourt/Notebook/Idli and micro-organisms"

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<h2 class="date one">July 2nd to 9th</h2><br><br>
 
<h2 class="date one">July 2nd to 9th</h2><br><br>
  
I experiented different recipes to make idli come from India. I tried to do it with different rice and lentill (called dall in india). Finally, I chose to do  
+
I experienced different recipes to make Indian idli. I tried to do it with different rice and lentils (called dall in india). Finally, I chose to do  
 
<a href="https://2015.igem.org/Team:Paris_Bettencourt/Protocols/Idli_recipe">this recipe</a>
 
<a href="https://2015.igem.org/Team:Paris_Bettencourt/Protocols/Idli_recipe">this recipe</a>
 
with basmati rice and indian dall. <br><br>
 
with basmati rice and indian dall. <br><br>
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<IMG SRC= "https://static.igem.org/mediawiki/2015/5/52/Idli_water.JPG" width=20%>  
 
<IMG SRC= "https://static.igem.org/mediawiki/2015/5/52/Idli_water.JPG" width=20%>  
 
<br>
 
<br>
I did a media from an idli preparation. After fermentation of 18 hours, of ildi composed of 3 glasses of rice (75ml) and 1 glass of dall (25ml) before the soaking phase, I took water and butter mixed it together in 500ml bottle, and did an autoclave. I called this media "idli water", and it look like previously. I will just use the clear phase.<br><br>
+
I did a media from an idli preparation, using 3 glasses of rice (75ml) and 1 glass of dall (25ml) before the soaking phase. After 18 hours of fermentation, I took the water and butter of this idli, mixed it together in a 500ml bottle, and autoclaved it. This media was named "idli water", and it looked like previously. I will just use the clear phase.<br><br>
  
  
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<h2 class="date one">July 12th to 14th and 16th to 18th</h2><br><br>
 
<h2 class="date one">July 12th to 14th and 16th to 18th</h2><br><br>
  
I did grow ''Saccharomyce cerevisiae'' with mCherry and geneticin resistance genes in idli. I added 2 ml of a yeast growth solution (YPD) with an OD600 of 0.455 (around 1.2 x 10<sup>7</sup> cfu) after the soaking phase and the mixing and grinding of rice (3 glasses of rice ~ 75ml) and dall (1 glass of dall ~ 25ml) together.The first try was to see if there are the fermentation process will this laboratory strain. After the fermentation time, I had plated the µorganisms come from the butter phase (middle phase between water on the top, and grind matter in the bottom of the erlenmeyer) on YPD with geneticin. I observed a lot of part of crush rice or dall, and just few colonny, not red like expected with the mCherry gene. The second try was more successful. This time I diluated the butter 10 and 100 time before plating, always on YPD and geneticin. I observed few colonies for the plate with the 100th and around 30 with the 10th. <br><br>
+
I grew ''Saccharomyce cerevisiae'' with mCherry and geneticin resistance genes in idli. I added 2 ml of a yeast growth solution (YPD) with an OD600 of 0.455 (around 1.2 x 10<sup>7</sup> cfu) after the soaking phase and after mixing and grinding the rice (3 glasses of rice ~ 75ml) and dall (1 glass of dall ~ 25ml) together. The first try was to see if there is the fermentation process with these laboratory strains. After the fermentation time, I have plated the µorganisms coming from the butter phase (middle phase between water on the top, and grind matter in the bottom of the erlenmeyer) on YPD with geneticin. I observed a lot of particules of crush rice or dall, and just few colonies, not red like expected with the mCherry gene.<br>
 +
The second try was more successful. This time I diluted the butter 10 and 100 time before plating, still on YPD and geneticin. I could observed only few colonies for the plate with the 100th and around 30 with the 10th. <br><br>
  
  
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<h2 class="date one">July 20th</h2><br><br>
 
<h2 class="date one">July 20th</h2><br><br>
  
I did a culture on TECAN plate of few species that was ''Saccharomyces cerevisiae'' with mCherry on chromosom, with Lactobacillus plantarum NC8 and Lactococcus Lactis MG1363 on their respective media (YPD, MRS and M17) and on "idli water" and on water, with and without antibiotics (respectivly geneticin disulfate, and erythromycin for the 2 bacteria). <br>
+
I did a culture on a TECAN plate of few species: ''Saccharomyces cerevisiae'' with mCherry on chromosom, Lactobacillus plantarum NC8 and Lactococcus Lactis MG1363 on their respective media (YPD, MRS and M17), then on "idli water" and on water, with and without antibiotics (respectivly geneticin disulfate, and erythromycin for the two bacteria). <br>
I did a TECAN mesure after an over night growth of 18 hours, and we obtain the table followed. We can see that the mesure was nonsens.<br><br>
+
We did a TECAN measure after an overnight growth of 18 hours, and we obtained the table followed. We can see that the results are nonsens.<br><br>
  
  
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<h2 class="date one">August 10th</h2><br><br>
 
<h2 class="date one">August 10th</h2><br><br>
  
I did electrocompetent cell of ''Lactobacillus plantarum'' with <a href="https://2015.igem.org/Team:Paris_Bettencourt/Protocols/Electrocompetent_Cells_Preparation_for_Lactobacillus_Plantarum"> a particular protocol</a> <br>
+
We did electrocompetent cells of ''Lactobacillus plantarum'' with <a href="https://2015.igem.org/Team:Paris_Bettencourt/Protocols/Electrocompetent_Cells_Preparation_for_Lactobacillus_Plantarum"> a particular protocol</a> <br>
 
It worked very well.<br><br>
 
It worked very well.<br><br>
  
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I had done <a href="https://2015.igem.org/Team:Paris_Bettencourt/Protocols/Idli_receipe">idli protocol</a>, with 75 ml of rice, for 25 ml of dall. After grinding and mixing, I added 2ml of ''Saccharomyce cerevisiae'' (with mCherry gene and geneticin resistance) that contained 10<sup>8</sup> cells. I had plated at t0 and 2h, 4h, 6h, 19h30, 21h30 after t0 three different positons (middle top, lateral middle and bottom middle) at three different concentrations (10<sup>0</sup>, 10<sup>-2</sup> and 10<sup>-3</sup>). <br>
+
We did the <a href="https://2015.igem.org/Team:Paris_Bettencourt/Protocols/Idli_receipe">idli protocol</a>, with 75 ml of rice, for 25 ml of dall. After grinding and mixing, we added 2ml of ''Saccharomyce cerevisiae'' (with mCherry gene and geneticin resistance) that contained 10<sup>8</sup> cells. We plated at t0 and 2h, 4h, 6h, 19h30, 21h30 after t0 three different positons (middle top, lateral middle and bottom middle) at three different concentrations (10<sup>0</sup>, 10<sup>-2</sup> and 10<sup>-3</sup>). <br>
  
 
<IMG SRC= "https://static.igem.org/mediawiki/2015/2/26/Cfu_number_evolution.png" width=50%> <br>
 
<IMG SRC= "https://static.igem.org/mediawiki/2015/2/26/Cfu_number_evolution.png" width=50%> <br>
Like we can observe, the areas where I take sample for plating don't matter, and the yeast did't grow very well : just a doubling size of population during 22 hours of culture. <br><br>
+
We can see that the area from where we sample for plating doesn't matter, and the yeast didn't grow very well : just a doubling size of population in 22 hours of culture. <br><br>
  
 
<div class="column-left"align="center">
 
<div class="column-left"align="center">

Revision as of 15:35, 14 September 2015

July 2nd to 9th



I experienced different recipes to make Indian idli. I tried to do it with different rice and lentils (called dall in india). Finally, I chose to do this recipe with basmati rice and indian dall.


I did a media from an idli preparation, using 3 glasses of rice (75ml) and 1 glass of dall (25ml) before the soaking phase. After 18 hours of fermentation, I took the water and butter of this idli, mixed it together in a 500ml bottle, and autoclaved it. This media was named "idli water", and it looked like previously. I will just use the clear phase.

July 12th to 14th and 16th to 18th



I grew ''Saccharomyce cerevisiae'' with mCherry and geneticin resistance genes in idli. I added 2 ml of a yeast growth solution (YPD) with an OD600 of 0.455 (around 1.2 x 107 cfu) after the soaking phase and after mixing and grinding the rice (3 glasses of rice ~ 75ml) and dall (1 glass of dall ~ 25ml) together. The first try was to see if there is the fermentation process with these laboratory strains. After the fermentation time, I have plated the µorganisms coming from the butter phase (middle phase between water on the top, and grind matter in the bottom of the erlenmeyer) on YPD with geneticin. I observed a lot of particules of crush rice or dall, and just few colonies, not red like expected with the mCherry gene.
The second try was more successful. This time I diluted the butter 10 and 100 time before plating, still on YPD and geneticin. I could observed only few colonies for the plate with the 100th and around 30 with the 10th.

July 20th



I did a culture on a TECAN plate of few species: ''Saccharomyces cerevisiae'' with mCherry on chromosom, Lactobacillus plantarum NC8 and Lactococcus Lactis MG1363 on their respective media (YPD, MRS and M17), then on "idli water" and on water, with and without antibiotics (respectivly geneticin disulfate, and erythromycin for the two bacteria).
We did a TECAN measure after an overnight growth of 18 hours, and we obtained the table followed. We can see that the results are nonsens.

July 29th



Test of the titration protocol with spectrophotometer for the Vitamin A (protocol titration)
I tested this protocol and tried to calibrate it. I used, like food sample, 1g of rice and I did 8 bottles with 8 different concentrations of pure vitamin A from 10 -1 to 10 -8 mg.ml-1.

Results :
We can just observe concentrations higher than 10-3mg.ml-1.
dilution of the initial concentration Value of the spectrophotometer
10-1 1.625
10-2 0.078
10-3 0.034
10-4 0.008
10-5 0.002
10-6 0.005
10-7 -0.001
10-8 0.000

August 10th



We did electrocompetent cells of ''Lactobacillus plantarum'' with a particular protocol
It worked very well.

August 14th



We did the idli protocol, with 75 ml of rice, for 25 ml of dall. After grinding and mixing, we added 2ml of ''Saccharomyce cerevisiae'' (with mCherry gene and geneticin resistance) that contained 108 cells. We plated at t0 and 2h, 4h, 6h, 19h30, 21h30 after t0 three different positons (middle top, lateral middle and bottom middle) at three different concentrations (100, 10-2 and 10-3).

We can see that the area from where we sample for plating doesn't matter, and the yeast didn't grow very well : just a doubling size of population in 22 hours of culture.