Since my last post in May, the summer has come and all but gone. You might think I was larking about on summer vacation. You would be wrong, I’d be happy to write up in a tree house (not that I would actually take my summer vacation inside a tree house). No, the reason for silence was nothing less than disappointment. The world spat a fat glob on my enthusiastic face. And not a personal but a scientific disappointment (tho if you are regular reader of this blog you will know that for me the line between personal and scientific is thin and wavering).
In May, I blogged about the samples I was bringing to Brookhaven National Laboratory. From those sectioned maize roots, I was hoping to detect a signal from the oriented cellulose microfibrils. There was not a sausage. We fussed for a day and a half, with both thin and thick samples, mounted in various orientations with respect to the beam. Not a hint of any oriented structures. There were problems with beam alignment but evidently these were overcome and the results were still the same.
On the face of it, this should not be such a big deal. It was only the second time of trying, so as these things go, not a tremendous investment. Moreover, getting results from roots was a long shot anyway. I do think that the results Ingo Burgert’s group got from a single hypocotyl suggest that it is possible. But on the other hand they probably invested a lot more time to get everything exactly favorable. I should have been able to write about the various things we tried and how none of them succeeded.
But for some reason, I didn’t. Some nasty squishy psychological reason to be sure. Don’t worry, I am not going to go all psycho here. Just noting that this feeling of sadness or perhaps futility was strong and it oozed over me all summer, blocking blogging. There is also the fact that I did little in the way of lab work. It happened that an actual vacation and a trip to the International Botanical Congress were scheduled in a way that interrupted a productive lab work routine. I focused on papers and computer things. But I could have found something to recount.
Well, with the ending of summer, let’s say this disappointment is falling away like yellow leaves on a sugar maple. And I have started an experiment. This will probably fail too but I promise to write about it anyway. This one is in-house, no giant synchrotrons involved. But it does involve the red room I set up in the former Biology Department darkroom.
Way back on Jan 8th, my blog posted another entry in the segment wars. These episodes started way back in the first season of Lab Fab. I was trying to develop a system to study growth anisotropy in stem segments. In my original efforts, I grew the seedlings, and did the entire experiment, in total darkness. The scientific reasons were for reproducibility (no need for a super-sensitive light meter) and for fastest growth (light inhibits stem growth). But as I have confessed before, I took a certain macho pride in such exacting conditions. (If you are interested in how I worked in total darkness, read more here.)
That series of eight or so trials foundered on the segments, cut from the maize seedling stem, aka the mesocotyl, growing fast or slow in the same treatment. The variation was as intractable as inexplicable. On Jan 8th, I described testing a new explanation: dim red light instead of darkness. This was in response to a suggestion from Rainer Hertel, an outstanding plant physiologist, who believes that totally dark grown seedlings are in a metastable state. Nope, that was not it. Despite growing the seedlings and doing the experiment in dim red light, the segment growth was, as ever, ridiculously variable.
I thought of one more explanation, and the new experiment is designed to test this. Happily, I can continue in dim red light because while it didn’t solve the problem it didn’t aggravate it and working in light, even dim light, is a boon.
But first, some technical difficulties. For the experiment to test dim red light, I replicated the system that I had used to record segment growth while I was on sabbatical in Nottingham. Fortunately, I brought home the little washer dishes, the rack to hold them, and the camera to take their picture. This all made it relatively easy to rebuild the system here at UMass. My colleague Peter Hepler had a photographic darkroom in a corner of his lab that had long been idle thanks to the digital revolution. It even had an excellent light-tight revolving door. I bought some red LEDs and made a little experimental nest in there.
Alas, in Feb, just after that experiment, Hepler’s lab was decommissioned and he moved upstairs, with no darkroom. I was granted access to the Biology department’s old photographic darkroom, then just used for storage. During the summer, I recreated the segment measuring system up there. It doesn’t have a revolving door, but it does have two doors, and the light-tightness tho imperfect is reasonable. I taped over a few leaks. There is also reasonable temperature control and a bench smooth and wide, with generous space to cut and handle the segments as well as for the camera rack. All good.
This week, I made the last touches on things. I planted some maize seed (ok, the verb planted might make you think of soil but these seeds are “planted” on soaking wet spongy filter paper, inside a black plastic box, which is covered with plastic wrap). I discovered that the album feature of Micro-manager had stopped working. This is the thing that lets me save images without opening the computer lid (and hence dousing the plants in light from the screen) by pressing space-bar on a wireless keyboard. Even on the main keyboard, hitting space-bar caused nothing to happen. I had to assign a hotkey (the space-bar) to this function. I swear I never had to do this before. But it solved the problem. No updates on the computer, no changes at all. Truly strange. But whatever. Hope it is stable for the experiment this coming week.
I will test that new explanation this week, probably on Tuesday. And I promise to write about it this weekend, or anyway soon, revealing the nature of the explanation as well as whether it holds water, no matter how disappointing.