Hannah Holland-Moritz is a graduate student working in Noah Fierer’s lab. She graduated from UC Davis in June 2014 with a major in Biochemistry and Molecular Biology and minor in Bioinformatics and most recently spend a gap year working in Jonathan Eisen’s lab on the microbiome of seagrasses. Interested in Evolution, Ecology, Bioinformatics and all things microbial, she plans to pursue a career in research.
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About a year ago I started an Intra-plant biogeography project. Limited in scope, this project’s primary aim was to determine how much variation there was in the microbial communities across a single plant in “high resolution.” The goal was to determine whether it mattered where our ZEN collaborators cut their samples from along the roots and leaves.
The general project was this: Cut a plant into about 50 strategically chosen pieces and look at the community variation across the surface.
One thing that always bothered me about these results were that they were for only one plant. I didn’t know if the cool patterns I was seeing were normal or a fluke. That’s where Biogeography 2 comes in, it’s a continuation of the first project but with more replicates (five, to be precise) all collected at the same time and from the same place. In the coming weeks I’ll be processing these samples and updating you about the progress.
This week’s update:
This week I finally was able to mutilate dissect the plants and now we can begin extracting DNA from the samples. Here are some pictures of plants prior to dissection.
For a plant that withstands daily tidal forces, seagrass are surprisingly delicate when taken out of water. When they dry out, they crumble so I try to section them as fast as possible to prevent drying.
Sample preparation includes painstakingly disentangling these roots from each other and from the shoots without breaking them. (About a 2 hour process per plant).
You might be wondering, “Whatever happened to that Zen Project stuff?”
ZEN (Zostera Experimental Network) is a global partnership of seagrass researchers studying Zostera marina. Over the summer and late last spring we sent 24 sampling kits to researchers across the world. Except for a few sites (international sample collection is hard) we’ve received nearly all of the kits with samples intact and we’re well on our way to finishing extracting DNA from each of them.
The first kit we received,
from North Carolina.
Here’s a map of all of the samples we’ve received so far (it will be updated in real time as we receive more samples).
We have also had a couple of experimental challenges that are worth noting:
Collection strategies: As with any carefully planned field experiment, things go wrong once you’re out in the field. It seems like some of our collection techniques are harder to carry out than we had originally though. The filtering process is not always clear and it’s often hard to fully submerge roots and leaves in the Zymo storage buffer we provided.
Extraction: It turns out that despite its great capacity for sample preservation, the Zymo buffer does not play nicely with our DNA extraction method. We’ve dealt with this by modifying the extraction protocol, but we still get lower yields per sample than we had hoped.
In spite of these challenges we’ve still been overwhelmingly successful:
Received kits: Nearly all of the kits are back and being extracted!
Wonderful Collaborators: Many of our partners have graciously offered to help collect extra seagrass species for other aspects of the project. (Thank you!)
Often when microbiologists begin to explore an unknown (or relatively unknown) environment, they begin by using classical microbiological techniques to try to characterize the communities of microbes living in that environment. These classical techniques are often referred to as “culture-based” because they are oriented towards the goal of trying to grow (or culture) microbes in the lab. Although culture-based techniques can be limiting (it’s nearly impossible to culture every single organism in any given environment), they are very useful for laying the foundation for the non-culture-based techniques the Seagrass Microbiome project will be using.
As a result, we in the Eisen lab have been playing around with some classical microbiology techniques alongside our non-culture-based explorations. One of the coolest techniques we’re using is Winogradsky columns. Winogradsky columns are essentially microbial terrariums. The basic recipe is as follows: take a clean tube, add a few essential chemicals, spike in some wild microbes, close the lid and let natural nutrient cycles take over. If you’ve done everything right, in a couple of weeks, you should start to see layers of microbes each living in a different mini niche within your mini ecosystem.
Two weeks ago we did just that. We’ve been following the progress of the tubes on the side as we pursue other non-culture-based projects. We’ve also been exploring different ways of making the columns to figure out the best way to get clean layers while still preserving the integrity of the column. Below are some pictures of our progress to date: