I’m not going to make many comments about these cells. It seems that in D2O, E. coli is more likely to say fused but it’s not as obvious as it is with yeast. I make no other observations.
E. coli cells in D2O
Tag Archives: data
Last Arabidopsis Update
The media is dwindling and the plants will die shortly. I’ll be restarting this experiment in larger environments with more media. Hopefully the plants can do better.
Yesterday Koch noticed that the roots in 10% D2O were much longer than those of the plants in DDW. Something interesting.
E. Coli Colony Morphology in D2O
Here are the results of yesterday’s setup. Here I’m comparing 4 samples:
- Wild type (WT) E. coli grown on DI LB agar
- WT E. coli grown on D2O LB agar
- D2O adapted E. coli grown on DI LB agar
- D2O adapted E. coli grown on D2O LB agar
All 4 samples were incubated for the same amount of time, and taken from starter cultures of similar absorbance. The absorbances of the starter cultures are as follows:
- Wild type (WT) E. coli grown in DI LB – 0.641
- WT E. coli grown in D2O LB – 0.325
- D2O adapted E. coli grown in DI LB – 0.489
- D2O adapted E. coli grown in D2O LB – 0.112
The D2O adapted E. coli took over the DI LB plate! I’ve re-inoculated those cells from that plate to get a picture that would show a typical colony and compare that morphology to the rest. I’ve also allowed the two D2O media samples to incubate for another 24 hours.
It should also be said that the D2O adapted colonies grown on D2O media look distressed compared to the WT colonies on DI media. But compare the two D2O adapted colonies and it’s tough to discern. Whatever mechanism gives the colonies a distressed appearance on D2O media, seems to be completely uninhibited on DI media. It’s tough to tell if the cells are distressed or just out of control.
Still these results seem pretty comparable to the last time I did these experiments (except using YPD). I’ll update again tomorrow, and insert these results into my dissertation.
Preliminary Results of YPD deterioration
These are the results of the experiment I stated a couple weeks ago. I have been tracking the deterioration (previously called aggregation, but I’m not entirely sure aggregation is the correct terminology) of YPD in both solvents. Today they looked pretty well degraded so I thought I’d share the results. Between the two, the DI YPD is more absorbent than the D2O YPD at nearly every wavelength measure (major uncertainty below 350nm).
I’m associating degradation with absorbance since the blank (which is also DI YPD) has an absorbance of zero at all the same frequencies.
D2O YPD also records 0 for absorbance at 600nm, which is the wavelength used for cell count studies, so there would be no interference from the solution. Whether or not the media is still usable by cells is undetermined.
I’m beginning a second experiment that would track the absorbance every few days via the same mechanism. If you recall, I began this experiment taking pictures and eventually moved toward using the nanodrop. This probe seems to do a good job so its continued use is reasonable.
Man I’ve been writing my dissertation for too long…
Arabidopsis Update (From March 6, 2013)
I took these images several days ago and again forgot to post them. Stupid dissertation…
Anywho, here they are. I’ll try and remember to update tomorrow when I take the images. Tomorrow will be the last day I update this experiment. I’ll be going into full dissertation mode and will be starting a new experiment when I return. I bought some larger test tubes (1in wide), which should give the plants all the room they need and should be large enough to provide more media to keep the plants alive for longer.
Arabidopsis update
This will probably be the last update. I’m going to (1) need to take a break until after I defend, and (2) need to start a new experiment because the plants are running low on media. I did just buy these awesome 1in diameter test tubes which should give the plants all the media and water they could need for a longer period of time. Anyways let’s go to the pictures:
YPD aggregation update
Remember this experiment?
Checking in on it, the YPD has already begun it’s aggregation in both cases, but it is more prevalent in the DI YPD case. Which should be expected. At this point I can see a concentration gradient in both solutions, but the full-on aggregation hasn’t happened yet. As is, it’s not worth photographing because there isn’t much difference.
But thinking back to here, I remembered we used a Dynamic Light Scattering Machine (the DynaPro Titan TC by Wyatt), which just uses scattered light to calculate particle size. In the aggregation experiments we didn’t care so much about specific particle size, just that we wanted to see the particles grow bigger.
If in fact the YPD is aggregating, the solution would become cloudier by eye. So since our Nanodrop measures absorbance, we could potentially see the same effect. The catch is that the DLS machine mentioned above measures at 830nm, and my nanodrop measures a broad spectrum from 250nm-700nm. In that range I’m not sure where it is most accurate except at 260nm, 280nm, and 600nm (which is where it measures DNA, proteins, and cell counts).
I’ll do a more thorough study on Monday, but here is the quick analysis:
orange – aggregated DI YPD
green – aggregated D2O YPD
Arabidopsis Update
Here is the update for my arabidopsis growth. It still seems to me that the plants in 10% D2O are doing better than the plants in DDW. I’ll have to make a huge purchase of D2O and DDW (still don’t understand why DDW is more expensive than D2O since one is a by product of the other) and try lower concentrations of deuterium (like 0-10% amounts).
Notice that the seeds in 99% D2O germinated (mostly likely due to D-exchange), but haven’t grown in 3 weeks.
I wanted to take images of the root growth, but my camera couldn’t see clearly enough in the agar to take decent pictures.
YPD aggregation study
Two summers ago Kenji Doering (an REU student that summer) and I did some protein aggregation studies comparing D2O and DI water. The results were pretty consistent in that proteins don’t aggregate under the same conditions in both samples. What that meant was the D2O is almost certainly better for longevity of chemicals.
A few months ago I noticed that my ypd stocks go noticeably bad after a couple of weeks. But they seemed to last longer in D2O, I just never quantified that. Well now I’m going to.
I have two cuvettes sitting on my bench. One is filled with 1ml of DI YPD and the other is 1ml of D2O YPD. They are sitting next to the benchtop cooler to supply a bit of heat to speed up the aggregation reaction. I will take daily pictures of the solutions to compare the aggregation times of each media. Here is the day=0 time point (on the left is the DI YPD sample, and the right is the 99% D2O sample):
Arabidopsis Update: From Feb 13, 2013 (6 days ago)
These pictures are a bit late, but better late than never. They are an update of the growth progress of arabidopsis in varying amounts of D2O. The group shot is inconsistent with the others because the sample captured for 0% D2O (DDW, deuterium depleted water) is different than the sample used in the individual 0% D2O image.
With that said, there are a couple morphological things I would like to point out. In my opinion the plants are growing better (bigger, faster, etc) in 10% D2O than they are in DDW. They also appear to be slightly more green. And finally there are little hairs on the leaves that seem to be more prominent than in the other samples.
Meanwhile, in 60% D2O the plants are very yellow in appearance. It is interesting that more plants sprouted in that sample, but they are significantly behind in development.
I’ll update tomorrow for the weekly update. And this time I’ll upload the images in real-time. Promise!