- Water (already prepared and antibiotic added):
- 24ml D2O YPD
- 16ml DI YPD
- 0.8g agar
- Stir and heat on hotplate
- Pour 15-20ml into petri dish and allow to cool
- seal with parafilm
Category Archives: D2O Adaptation 3
60% D2O Time Trial
Setup. I actually did this experiment yesterday, but time constraints prevented me from posting the live results (which is funny because that requires literally 5 minutes, oh well). Anyway here is some fun data:
40% D2O Yeast colonies comparison
Attached are some images taken of the colonies with my iPhone, my iPhone macro lens, and my iPhone microscope (respectively). Notice the difference in growth and colony size:
40% D2O yeast cell morphology
It is tough to determine differences between the 20% D2O cells and these, but they appear slightly more spherical and also a bit larger. Evidence of stalled cellular division is not yet apparent. Check out the cells:
40% D2O yeast colony morphology: Part 1
The colonies are not nearly as well developed as those grown on 20% D2O, but there is evidence of stress even at this stage. The colonies (1) are obviously smaller than the 20% D2O growth, and (2) have a little less spherical symmetry. I’ll continue to grow the yeast in the incubator and check the progress tomorrow (but once before I leave tonight).
20% D2O yeast colony morphology
The yeast grown on 20% D2O YPD exhibits growth not all that different from yeast grown on normal DI water plates. It’s tough to ascertain colony growth time because analysis was not performed in time intervals. Check out what a standard individual colony looks like:
Long Term Yeast Adaptation Update
After 2 weeks of yeast growth, I thought I’d re-share the long term adaptation spreadsheet. So far no bacterial contamination, but that’s thanks to antibiotics. Anyways, here is the spreadsheet again:
40% D2O Yeast Growth Time Trial 2 Live Results
New YPD made
I just ran out of my last batch of both DI and D2O YPD. This time I am making the protocol with 500ul of Ampicillin added (460ul added to D2O).
Yesterday I added amp to my 10ml test tube sample and it seems to have killed the bacterial infestation without impacting the yeast growth. Normally when there is bacterial growth, the yeast settles and the bacteria floats in solution. This morning the solution was clear and the yeast was nicely nestled at the bottom.
Adding Ampicillin to YPD
So I’ve noticed that no matter what, there is some bacterial contamination, and based on an experiment I did with the reconstituted yeast, it seems the contamination exists there. After doing some research, it seems I can add antibiotics to the YPD without affecting yeast growth and preventing bacterial growth. So today I’m doing just that. I’ll take a picture of tomorrow’s culture to reveal whether or not this method works against the bacteria, and if it impacts the yeast growth.
It turns out that I have ampicillin as a powder. So I added 20ml of water to it to bring it to a concentration of 10mg/ml. According to a protocol I found on my old stomping grounds (openwetware.org), I need to add 50ul to 10ml of YPD to get a final concentration of 50ug/ml.