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Tag Archives: ddw effects on life
Yeast in DI, DDW, 30%, 60%, and 99% D2O Trial 3: Setup
Yesterday I made starter cultures in D2O, DI water, and DDW. Today I’m doing the time trial experiment and here is the setup:
- Setup of five samples:
- 9ml of DI YPD + 1ml of DI starter culture
- 9ml of DDW YPD + 1ml of DDW starter culture
- 6ml of DDW YPD + 3ml of D2O YPD + 1ml of DDW starter culture (30% D2O sample)
- 3ml of DDW YPD + 6ml of D2O YPD + 1ml of DDW starter culture (60% D2O sample)
- 9ml of D2O YPD + 1ml of D2O starter culture
- Measuring:
- 400ul of each starter culture in semi-micro cuvettes – to measuring starting absorbance
- 400ul of each timed culture (the samples to be measured over time) in semi-micro cuvettes – to measure the time=0 value (should be close to 1/10 of the starting measurements)
- repeat B every hour
- Blanking:
- Before each hourly measurement you need to blank the nanodrop
- Blank the DI samples with DI YPD (no yeast added)
- Blank the DDW, 30% D2O, and 60% D2O samples with DDW YPD
- Blank the D2O samples with D2O YPD
Starter Cultures: D2O, DDW, and DI YPD
I just started three new starter cultures for another timed experiment tomorrow. Hopefully this time it goes better than last time.
Setup:
- 10ml of YPD broth in test tubes
- The water component of each test tube is different, one is DDW, one is DI water, and the other is 99% D20.
- The YPD was made last week and stored in the fridge.
Yeast hourly growth in DI, DDW, and 30%, 60%, 90%, and 99% D2O
Yeast growth in DI, DDW, and 30%, 60%, 90%, and 99% D2O. Anthony Salvagno. Figshare.
Retrieved 21:20, May 24, 2012 (GMT)
hdl.handle.net/10779/caa57855586213d79eea5576f8da23d0
Notes:
- I don’t think this is a reliable data set. Several of the samples actually read a lower absorbance after the first hour than they initially do. And then they all dip again at hour 4.
- I noticed a considerable amount of cell settling after hour 3 on the bottom of each test tube. I mixed prior to reading the absorbance values, but this is likely to skew the results. Next trial I will have to mix before reading every hour.
- The data between DI, 30%, 60%, and 99% D2O look consistent with Tuesday’s results, but it scares me that the DDW and 90% D2O are completely out of whack.
Yeast hourly growth in DI, DDW, and 30%, 60%, 90%, and 99% D2O: Setup
Yesterday I set up starter cultures for this experiment using D2O, DDW, and DI water as the basis for the liquid media. Today I’m running the time trial experiment again, but this time with a bit more of a spectrum in water amounts. Here is the setup:
- I am using 6 samples: DI water, DDW, 30% D2O mixed with DDW, 60% D2O mixed with DDW, 90% D2O mixed with DDW, and 99%D2O.
- For the mixes:
- add D2O, add DDW, add 1ml starter culture from DDW (1:10 dilution).
- Example: 30% D2O in DDW is 3ml D2O, 6ml DDW, and 1ml DDW starter culture
- For the other samples:
- DI: 9ml of DI YPD, 1ml of DI starter culture
- DDW: 9ml of DDW YPD, 1ml of DDW starter culture
- 99% D2O: 9ml of D2O YPD, 1ml of D2O starter culture
It should be noted that the 99% D2O starter culture was almost completely translucent. By this I mean it looked like there was no growth in the sample at all. Results that I’ll post later (when the experiment is complete) will show that this is indeed true, with an absorbance of 0.521 (compared to ~2.1 in both DI and DDW starter cultures, 4x the cell growth!).
I’ll be taking growth readings every hour via the nanodrop in micro cuvettes (til about hour 4) and then switching over to semi-micro cuvettes (because I’ll run out of micro cuvettes).
Love,
Ant
Starter Cultures in D2O, DDW, and DI Water
Based on the results from yesterday I’ll be carrying out the next phase of the experiment. I set up starter cultures to grow in DDW, DI water, and D2O so I can do hourly time points tomorrow starting early and hopefully getting about 7 hours of data (instead of 5 like yesterday).
I had grown yeast on solid media yesterday and got a bunch of colonies today, so I plucked three colonies and inoculated them in the liquid media stated above (YPD broth).
Yeast Growth Trial 1 Data
Here is the data courtesy of figshare:
S. cerevisiae growth in DI Water, DDW, 50% D2O, and 90% D2O. Anthony Salvagno. Figshare.
Retrieved 22:01, May 22, 2012 (GMT)
hdl.handle.net/10779/bfceb213ffb663b34149d97141755232
I would say this data is pretty exciting. Unlike the E. coli data (which was all over the place), there is a clear difference in growth rates between the D2O samples and the H2O samples. Also the growth between the DDW and DI (the H2O samples) were almost identical! That’s pretty solid.
A cool figure I forgot to share: E. Coli growth in Water
I made this figure for my rockethub proposal and forgot to post it here in my notebook. Look how pretty it is!
I made this in Adobe Illustrator and if anyone is interested I would be willing to host an online workshop to teach others how to use Illustrator for science. This particular image took almost no time (maybe 30 minutes), which is a marvel because I obsess over everything I do in Illustrator (which is why there isn’t anything tangibly Illustrator on this site).
Yeast Growth in D2O, DDW, and DI YPD: Setup
Yesterday I created a starter colony of yeast from the lyophilized yeast that I ordered from ATCC.org. Today I’m doing growth in different water types and checking the growth every hour just like the e. coli experiments. Here is my setup:
- Make YPD broth with DDW and D2O.
- the bottles of both come in 100g quantities which means there is 90-ish ml of D2O and 100-ish ml of DDW
- I had 91ml of D2O –> 4.55g of YPD powder
- 100ml of DDW –> 5g of YPD powder
- suspend powder in water types and filter via syringe. DO NOT AUTOCLAVE
- the reason is because of deuterium exchange. The powder is ultrapure and the water is pretty pure itself, so filtering should be just fine.
- To make diluted cultures:
- I have 4 samples: 1) DI water 2) DDW 3) D2O and 4) 50% D2O in 50% DI water
- Put 9ml of each water type into a test tube (except for the 50/50 mix which should be 5ml of d2o and 4ml of di water)
- Add 1ml of starter culture (which is made from DI water)
- put in incubator
- take time points every hour in semi-micro cuvettes using nanodrop. I fill them with 0.5ml of each culture.
Simple!
E. coli growth in different water types data
Via figshare:
E. Coli Growth in DI, DDW, D2O, 30% D2O, and 60% D2O. Anthony Salvagno. Figshare.
Retrieved 23:35, Apr 27, 2012 (GMT)
hdl.handle.net/10779/51fcd2f94fd7464449ee0f794642214c
I’ll post some interpretations here later…