I removed the seeds from the fridge and decided to keep them in a styrofoam cuvette case. This should keep light and temperature pretty stable in the lab. I then put tape over like groups of seeds so I can easily remove the cuvettes and set them up for picture taking.
Believe it or not, that little trick saves me a load of time.
Wow, that sucked. I had to crop, rename, and label 42 pictures for this notebook. Stupid open science movement. Why do I strive to be a good scientist? Why can’t I just let myself be lazy and allow myself to be a bad scientist? Who cares about accurate data and organization?
I do unfortunately.
Anyways here are the Day 0 pictures. Luckily I don’t have to do this every day, that would suck. From now on I may label the water types as follows:
RODI – Easypure RoDI water from the lab
CHTM – DI water from the CHTM plant
SMB – Sigma Molecular Biology DI water
STC – Sigma Tissue Culture DI water
DDW – deuterium depleted water
DDDW – 1% D2O in DDW
Since the last time I did this, I have learned a ton about proper exposure with a camera. And I actually know what aperture, ISO, and shutter speed are for. I’ll do a post that explains all that but in the mean time here are the settings I used for these pictures and those in the future:
Aperture – Set at F11
Shutter Speed – Set at 1/13″ (I think, it’s a Nikon camera so it just says 13)
ISO – Set at 1600
I’m sure these settings are saved with the images so if you download the pics you’ll see the camera settings in photo software.
After 4 trials and talking with Steve, there is no way to ensure that the results we are getting have anything to do with deuterium content. We can’t tell at all that our DI water is just not the same kind of “pure” that the DDW is. So I bought deionized water from Sigma (same people who make our DDW and D2O) to compare with our water and to compare to the DDW results to determine if the root hairs grow based on water purity or if it has something to do with deuterium content.
Here is the setup of the experiment:
I am using 4 different “types” of deionized water. We have a Easypure RoDI from Thermo (see Experiment Product Page at top) that I get DI water from. CHTM also has their own deionized water filtration system and I’m using that water in this experiment as well. I also purchased two different kinds of pure water from Sigma, one is molecular biology grade water and the other is double purified water for tissue cell culture. I honestly don’t understand the differences, but this chart says there are some.
I also used pure DDW for one sample and a 1% D2O mixture with DDW for another (because why should I exclude the DDW results from this study?)
I am keeping this experiment to just the tobacco seeds since the arabidopsis results are perplexing and not as obvious. So I am using both kinds of tobacco seeds (Havana and Virginia Gold #1, see Experiment Product Page).
I chose to do four samples per water type per seed type. With 5-7 seeds per sample. That means for each water type there are 8 samples, 4 for each type of seed.
Today’s protocol was really easy. I used my macro cuvettes (see Experiment Product Page) and set up 4 cuvettes for each water type giving me a total of 24 cuvettes. I did this so I could set up samples for one seed type at a time, and because I only had enough racks for this many cuvettes. I also prepared the D2O/DDW mixture (29.7mL of DDW, 0.3mL of D2O).
I poured 5-7 seeds in each macro cuvette until all the setup cuvettes had seeds in them. Then I added 3mL of water to the samples (again making 4 samples per water type). Then I sealed the cuvettes with PE caps.
I repeated this setup for the next seed type (24 cuvettes, 5-7 seeds per sample, 4 samples per water type, with 3mL of water per sample) and sealed and labeled all cuvettes.
I placed the cuvettes in the 4C fridge to synchronize the growth among all the samples. Tomorrow I’ll remove the seeds and take Day 1 pictures.