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Team:Valencia/Notebook/August
From 2009.igem.org
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August 3rd
- Yeasts haven't grown yet.
- iGEM meeting.
August 4th
Today we have had a yeast-team meeting. We suspect that our yeasts are not growing because they died during the trip between Barcelona and Valencia. We have decided to spread the yeasts we have left in YPD medium that had already been prepared, but we are skeptical about the results. If it doesn't work, Joaquin Arinyo and his teammates will send us more aequorin-transformed yeasts. We have also made 1 liter of newn solid SD lacking Leu medium.
August 5th
Today we have prepared the Coelenterazine stock dissolution. First we "put" N2 gas through a Methanol dissolution in order to substitute the O2, an inhibitor for the coelenterazine. After 5 or 10 minutes we took 200 microL and added them to 50 microg of Coelenterazine.
Then we have distribute them in eppendorfs of 5 microL each and kept them in the fridge at -20º.
For other side, we doubt if our yeast in the YPD medium have grown or not... We think we should wait until tomorrow... T-T... Like Cristina says, our yeasts are like Peter Pan: they don't want to grow up...
August 6th
Yeast have grown. YEaaaaaaaaaaaaahhhh!!! We have spread them into the SD solid medium lacking Leu that we did yesterday, we will wait until tomorrow for the results. Also we have prepared new SD liquid medium lacking Leu and once prepared we will also spread our super-yeasts in it. Tomorrow we will start with the protocol (we hope).
August 7th
Yeast have grown in SD medium lacking Leu (bothsolid and liquid) so we have started the protocol. The OD was low (0'18) but we have prepared an 1'8 OD dissolution of the yeasts by centrifugation and dilution with SD medium lacking Leu. Allright... well... we have mesured the "luminiscence" and we have seen nothing. We've made the experiment in an espectophotometer and in the electrophoresis gel photograph machine. Before the mesurements we checked the OD and it was 0'34 so very far from 1'8. Maybe that's the problem. We're not depressed anyway, on monday we will come back to demonstrate those yeasts what we can do. Valencia's iGEM way.
August 10th
Today, we've made a lot of things. Yeasts haven't grown after three days in SD lacking Leu liquid medium at 30º. We've found out that we were preparing the medium in the wrong way.
The right way is:
for 500ml
2'5 g (NH4)2SO4
1 g Yeast Synthetic Drop-Out Medium Supplement Without Leucine
10 g Glucose
7'5 g Agar (only for the solid medium)
We have also found out that the minimal volume for the spectophotometer is 400 microL so we will double all the quantities of the protocol.
August 11th
We repeated the protocol and we mesured in the espectrophotometer but we saw nothing. We think it's too few sensitive and we're going to ask for a fluorimeter. We also did the experiment in GelDoc but, again, we saw nothing.
