Chestuts #3–Late season and current thoughts

Long days of solitary work tend to encourage wandering thoughts. Plenty of them are ridiculous—“hey, maybe if I manage to selectively kill only the mosquitoes that land on me, I’ll be able to select for a new strain that won’t feed on people!”—but some are more serious. One of these more serious thoughts came to me on a particularly hot day when I was feeling drained and unguarded. I knew that the goal of the chestnut reintroduction program is to create a hybrid blight-resistant tree that will be capable of establishing itself and spreading in the environment with a minimum of human assistance. When you stop to think about it, as I did then, this is very close to the definition of an invasive species. If the reintroduction effort were to succeed, the shock to the ecosystem from the return of the species could potentially have cascading effects, as is so often the case with any change in an environment. And even if the new chestnut is able to readjust to its former niche without incident, that 6% foreign DNA in the average individual produced by the Foundation might be just enough to significantly alter the species’ ecological behavior and cause unpredictable disruptions to the habitat. These are just conjectures, of course. They have not been tested and cannot yet be tested. But they are necessary cautions to bear in mind as work proceeds.

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Ion Chromatography

In order to describe the dissolved matter in agricultural runoff, we analyze the filtrate in several ways.  The first of these is ion chromatography for anions.  An ion chromatograph (IC) functions by transporting sample in an eluent through a column.  The column is packed with material that attracts the anions in the solution.  Since each ion has a different level of affinity for the column material, each anion species exits the column at a different time after injection.  The instrument measures the conductivity of the liquid exiting the column over time.  On a graph, this translates into peaks which correspond to different ions.  Peak areas are calibrated using known standards at a range of concentrations.  The anions calibrated for are: fluoride, chloride, nitrite, bicarbonate, sulfate, nitrate, and phosphate.  The standard mixed from ultra pure water and the salts of the relevant anions.

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Chestnuts #2–Midseason

After two days of measuring seedlings, two major trends in survival became apparent. First, almost all of the seedlings planted in the forested plots were dead. I also noticed that they tended to die from the top down, with new stems regenerating from near the base of the original stem. Second, and much to my surprise, the fenced-in plots (protected from deer browsing) had actually suffered far higher rates of mortality than the unprotected plots.

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Filtering Procedure

Now that we have samples, the next step is to filter them.  Because organic matter can break down rapidly, runoff samples need to be processed as quickly as possible.  We filter about 150 mL of each sample through 0.7 µm glass fiber filters.  The filters are pre-combusted in a furnace to destroy any organic contaminants.  The filtration apparatus is thoroughly rinsed between each sample.  Ultra pure (Milli-Q) water is passed through the filter paper before any sample.  The filtration is expedited by a vacuum pump (Büchner-style).

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Thermoregulatory Tracing Part III: ISH

In situ Hybridization is the final step of the experiment, but it is also the hardest step.  Although there is a basic outline for ISH protocols, each protocol is customized for the specific probe.  This customization is largely a matter of trial and error, which can be extremely frustrating and time-consuming.  This issue is further compounded by the fact that ISH protocols require three full days to run before any results are seen, making it very possible to make mistakes and thus waste a lot of time and effort.

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