Category Archives: RC4

Repeating Crumley et al (1950)

Koch asked me in a comment to replicate an experiment done in 1950 by Helen A Crumley et al demonstrating Tobacco seed growth in deuterium oxide (D2O). The experiment is rather simple (and the figure from the paper is shown below) as Crumley placed 100 seeds in differing amounts of D2O (double distilled water, 33%, 66%, and 99.8% D2O) and analyzed the growth.

Influence of ordinary water, 33%, 66%, and 99% deuterium oxide on tobacco seed germination. Counts made at daily intervals for 39 days. (From Crumley, Fig 3)

So here I am planning the experiment. I will change some things from their experiment. First they placed the seeds on wet cloths (paper towels?), and I will submerge the seeds in the water amounts they used. They also used a variety of plant species (tobacco, clover, radish, Kentucky bluegrass), where I will just use tobacco seeds (but I will try two different species). Finally they talk about their results in terms of percent germination, but it isn’t clear from the paper if they mean number of plants that have exhibited germination, or if they are referring to some amount of growth exhibited by each plant. I will look for both possibilities and report the results as I find them.

In a preliminary experiment I will submerge the plants in water in petri dishes and seal it with parafilm. I will be looking into a more airtight solution as time goes on. I also won’t do 100 seeds but probably on the order of 33 seeds per sample. And in the future I will look into figuring out a way to measure the seed growth.

Original Crumley paper can be found here.

Typical Specimen for the Repeating Crumley Experiment

typical examples, of no germination, beginning germination, obvious germination, etc

Bill Hooker suggested (and rightly so!) that I document what I consider to be typical specimens. I found, today that the zoom feature on the webcam I’m using is quite sufficient for getting close enough to demonstrate this and so I snapped a picture and tried to document. Powerpoint messed me up a little bit, but this is good enough.

The orange box is highlighting a seed that is showing no signs of germination. From my studies I’ve found that the the seed coat becomes slightly transparent just before radicle (the pre root) penetration and you can see the precursor to puncture. The seed coat usually gets lighter too and in this case is really dark in color.

The pink box (hehe) is what I would count as germination when I’m counting the seeds. I see the tip of the radicle penetrating the seed coat and so germination is officially underway.

In the very near dead center of the image is a seed that has what I referred to in an earlier post as an extended radicle. Basically I meant that the radicle was pushed through and the root is now forming. The root will continue to grow until the seed coat has come off, which over to the right it has (there is a green leaf that is cropped on the edge of the image).

The end.

Repeating Crumley 4: The Setup

I’m going to keep this post short and sweet since the setup is nearly identical to the setup for RC3. That can be found here (nice and big link for visibility). Here are the big changes to the setup:

  • I used Viriginia Gold #1 tobacco seeds because I forgot I used the totally awesome Cuban cigar seeds in the last batch. Bad Anthony…
  • I also used Columbia arabidopsis seeds.
  • I did not count 30 seeds per sample. This time I poured seeds and counted until the number of seeds per sample was over 30. This way I can try and enhance the number of seeds in the middle of the sample. This was very helpful with the arabidopsis setup, because the seeds are much smaller than the tobacco seeds and have more attraction/repulsion from the analyslide and weigh paper. As I found out the hard way, something like 60% of the seeds would end up in the sample container and the rest would disappear into oblivion.
  • I omitted the DDW samples and will not take any pictures of the control, but will note if anything extraordinary happens in this sample. I figure it is really boring to see pictures of water everyday. I omitted the DDW samples so I can make room for the arabidopsis samples (because I can only fit 8 samples right now).
  • There are 4 samples per organism. DI, 33% D2O, 66% D2O, and 99.9% D2O, just as Crumley intended.

Originally, I had Alex (this person again? Who is he/she?) help me with the seed counting on Monday. I realized that I was out of D2O and ordered some. I didn’t get the D2O until yesterday and the presorted seeds had been left out over 24 hours. Since seeds may start sprouting or doing things in light and air, I trashed those seeds and started anew.

I assure you that I used D2O this time. Let’s repeat Crumley! (Day 1 pics to come later today.)

RC4: Day 1

RC4: Day 1 (control)

image

RC4: Day 2

Arabidopsis has already sprouted. In the future I may have to do a separate trial where I track growth more frequently.

RC4: Day 3