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.)

  • Pingback: Monday, 10/31/2011 « Alex Haddad's Notebook()

  • http://stevekochscience.blogspot.com Steve Koch

    Why no DDW?  I realize you’re saving time, but probably is worth doing, even though Crumley didn’t–it’s always possible the simple time to germination curve could change for arabidopsis in ddw

  • http://stevekochscience.blogspot.com Steve Koch

    I’m on the fence whether it’s OK to omit the control photos.  On the one hand, I understand it is time consuming and is just plain water and the camera can’t see anything you can’t.  On the other hand, I want as much information as possible.  Maybe a compromise would be to not embed the images for display (as you do with other photos), but at least link to them in case, for whatever reason, we want to see the pictures sometime in the future.

    • http://www.iheartanthony.com Anthony Salvagno

      This is in regards to both comments, but the way the system is setup I can only take pictures of 8 samples. I want to keep the pictures as repeatable as possible and my current method is optimized for this. I will do a DDW in D2O analysis in 10 days when this is done. As a result, the control also had to be sacrificed. And I thought about this too. I didn’t want to not take pictures of the control, but I literally don’t have room. So I will just have to observe and report findings until I figure out something better.

      • http://stevekochscience.blogspot.com Steve Koch

        You could snap “worse” photos of the control with your phone and upload to a public evernote notebook called “control” photos?

        • http://www.iheartanthony.com Anthony Salvagno

          Yea that’s a good idea which I will do.

  • http://stevekochscience.blogspot.com Steve Koch

    Also, for lack of a better place to post it, this is a good (long) link for you and Alex to look at: http://www.liveinternet.ru/users/1899851 It game up on my Google Scholar alert for citations of Lewis 1934 (I don’t know why it just came up, seems to be old article).  I didn’t read whole thing, but as far as I can tell it doesn’t talk about deuterium-depletion experiments, or mention that Lewis thought of it.  They do talk a lot about how deuterium does / might behave in cells.  A focus is on organisms adapted to high-deuterium content.  As you (or maybe Bill Hooker?) noted in the past, it would be interesting to investigate deuterium-adapted organisms when deuterium is taken away.  It seems obvious those organisms would be less fit in DDW.  This could just be due to a simple explanation that proteins have adapted to be more unstable.  But on the other hand, a surprising result may emerge that points to a fundamental process for utilizing / dealing with deuterium in cells.  Such as the root hair result, which still seems valid and repeatable, but we haven’t made much progress towards ruling out other causes.  So, a big hurdle remains in how to make sure the water we’re using is exactly what we think it is (and stays that way during experiments).

    • http://www.iheartanthony.com Anthony Salvagno

      It was Bill Hooker who suggested it.

      I think it would be interesting to do that as well. Alex told me she is an avid gardener as well. I think we could setup a hydroponic system to get an organism adjusted to D2O and then remove it. That would be awesome actually and I hope she is taking notes. Also I think we could do something similar with E coli and yeast. Grow them for a few generations in D2O and then put them in DDW and see what happens.

      We should talk about this in detail either in person, or I could create another comment thread that we could stick ideas in. What do you think?

      • http://stevekochscience.blogspot.com Steve Koch

        I think if pursuing this option (which is not a slam dunk), you should start with pre-existing strains that have been adapted.  That way you don’t have to learn to do it yourself.  I know there are D2O-sensitive yeast strains, but you want the opposite.  I’m guessing that D2O-resistant E. coli and yeast are available, but don’t know.  In either case, you need experience with “normal” ones first and also need to develop a good read-out for growth.  Plants are another story.  I don’t know if they’ve been adapted to D2O and are available.  But it would be a generational thing and thus much slower to do yourself than something that doubles every few hours.  You don’t have as much free time as Mendel.  As it stands, you have preliminary data with “normal” tobacco showing a phenotype to DDW.  Hopefully it will be similar with Arabidopsis.  If so, pursuing all angles of those would go a long, long way towards supporting the hypothesis.  Testing D2O-adapted organisms would be a supporting role (and fun!) but not as strong as if we see a phenotype in “normal” organisms.

        • http://www.iheartanthony.com Anthony Salvagno

          I know it would be a generational thing. From what I can tell arabidopsis grows fast so it may not be such a big deal, but I’ll definitely look into finding some D2O adjusted organisms. If you know where to find D2O sensitive stuff send it my way, that may be a good place to start looking.

  • http://www.facebook.com/people/Alexandria-Haddad/1266847552 Alexandria Haddad

    You could link to my page when you mention my name… Jeez.

    • http://www.iheartanthony.com Anthony Salvagno

      Well you’re going to get your own article today so I’m just building suspense. Jeez you’ve been in the lab two weeks and you’re already trying to abuse my fame for your own gain.

      • http://stevekochscience.blogspot.com Steve Koch

        Hurry up!  I’m dying to see her notebook (influenced by your fame, of course)!