Team:Paris Bettencourt/Notebook/Idli and micro-organisms

July 2nd to 9th



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


We 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, W 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. We will just use the clear phase.

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



We grew ''Saccharomyces cerevisiae'' with mCherry and geneticin resistance genes in idli. We 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, We 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. We 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. We could observed only few colonies for the plate with the 100th and around 30 with the 10th.

July 20th



We 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)
We tested this protocol and tried to calibrate it. We used, like food sample, 1g of rice and We 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 ''Saccharomyces 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.




August 19th and 20th



We started an idli 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.
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 Phytic acid assay kit on the samples.

The analysis of the phyticacid with the kit said us that ther are no phytic acid in the samples. (cf. Phytase Notebook)

August 20th



We did again the experiment where we added 108 cells for the fermentation step of the idli recipe. We used S. cerevisiae BY4743 mCherry, Lactobacillus plantarum NC8, and Lactococcus lactis MG1363. With a plate counter, we obtained this numbers of colonies in function of the dilution for the plating.
From the media in the top of the idli :
MediaDilutions Number of colonies of Yeast
YPD+gen 10-1 79
YPD+gen 10-2 32
YPD+gen 10-3 10
MRS 10-1 1
MRS 10-2 0
MRS 10-3 0
LB 10-1 0
LB 10-2 0
LB 10-3 1
M17 10-1 0
M17 10-2 0
M17 10-3 0

MediaDilutions Number of colonies of L. Plantarum/L. Lactis
YPD+gen 10-1 0
YPD+gen 10-2 0
YPD+gen 10-3 0
MRS 10-1 465
MRS 10-2 99
MRS 10-3 6
LB 10-1 96
LB 10-2 51
LB 10-3 15
M17 10-1 123
M17 10-2 3
M17 10-3 7

From the batter phase of the idli:
MediaDilutions Number of colonies of Yeast
YPD+gen 10-1 217
YPD+gen 10-2 150
YPD+gen 10-3 19
MRS 10-1 6
MRS 10-2 1
MRS 10-3 1
LB 10-1 2
LB 10-2 0
LB 10-3 0
M17 10-1 0
M17 10-2 0
M17 10-3 0

MediaDilutions Number of colonies of L. Plantarum/L. Lactis
YPD+gen 10-1 0
YPD+gen 10-2 0
YPD+gen 10-3 0
MRS 10-1 396
MRS 10-2 51
MRS 10-3 6
LB 10-1 125
LB 10-2 13
LB 10-3 8
M17 10-1 503
M17 10-2 111
M17 10-3 29

August 27th