Trials and Tribulations of undergrad projects

So at Reed we do independent projects in biology classes. They're usually kind of fun, and it's kind of a unique part of the curriculum here. However they get annoying for the following reasons:

1) we have 6 weeks to do them, so we are discouraged from doing time consuming or complicated techniques (because you just can't)
2) We aren't allowed to do something expensive. If we need to buy something expensive (like, say, a $300 antibody), we better damn have a reason for purchasing it, like someone else in the department will use it later. It can't just be your pet antibody for your pet protein. We used to have the resources to write a grant for expensive materials for independent projects, but one of our big teaching grants has run its course, so the department is tighter on cash than it has been for a while.

This makes sense; we're a small liberal arts school with limited resources. Our science program is solid, but as a small liberal arts school we don't have a lot of fancy schmancy equipment nor do we have a large budget. The opportunity to do independent projects is great for gaining lab experience, and a great opportunity, but at the same time, more of the money is spent on the faculty's research and people's undergrad theses. As it should be.

Depending on the class there is more or less guidance. Animal Physiology, Bio 381, happens to be a junior/senior class. Thus, there is little guidance at all and we are encouraged to think like scientists and figure out our own problems based on issues we are interested in. Good, great, I'm liking that. But it's hard because...shit son, it's hard enough to find an experiment that hasn't been done before, period. Add that on to a no more than 30 dollar budget and a 4-6 week time scale, and you have really slim pickings.

So we searched and searched and found this interesting pro-apoptotic signalling molecule, ceramide. It's kind of neat because it's a structural membrane lipid as well as being a pro-apoptotic signalling molecule, which is unusual. An experiment in the past showed elevated amounts of ceramide in the prescence of ionizing radiation (well in this case gamma radiation), in similar fashion to the way it was elevated in the prescence of TNF alpha (tumor necrosis factor) and chemotherapeutic drugs. Well, Reed has a nuclear reactor, kind of one of those odd little anomolous things--I'm actually a licensed operator, it's nearly completely run by undergrads--so it seemed like a good resource to take advantage of. Why not repeat the experiment looking for ceramide levels, but look for it with a greater variety of types of ionizing radiation (neutrons, gammas, and betas) in the reactor? Well cool.

Except then the issue of how to quantify the ceramide comes up. There's a kit that uses a radioactive phosphorous and a kinase, but that kit is expensive. No dice. Using a HPLC is probably the easiest method, but to my knowledge, both the HPLC in Arch's phyisology lab and the HPLC in the organic/biochem reasearch labs are not up and running. Okay. Oh here's a method, you can 1) extract the lipids out of the cells 2) run some silica gel chromotography (we found an elution plan in the lit) 3) functionalize it with benzyl chloride 4) do absorbance studies. Oh wait, forget that silica gel chromotography is a pain in the butt, that would take a shit ton of cells, we'd need to check to see if we actually purified it with the GC-MS/NMR, and...well....that's not exactly happening in 6 weeks on top of coursework. Okay, nix that.

Well, it turns out that a molecule in that pathway, spingomyelin (well, it's actually made from spingosine, but it's in the same pathway) turns into ceramide. So it would just be easier to take a look at upregulation or downregulation of the spingomylenase. We found a paper (which I don't have on hand and am too lazy to look up) about how TNF alpha and chemotherapeutic drugs lead to an upregulation of singomylenase, just as they lead to elevated levels of ceramide. BUT, no experiment was done with ionizing radiation. So, bingo. Primers are cheap. These are even published, so we don't have to worry about our primers not working (even if primer 3 input has done me well in life for designing primers for arbitrary genes, it's always nice to be assured that they have worked in the past).

Okay, problem. We asked if we could culture some fibroblasts to do the experiment. And got a "no, it's too complicated to do in 6 weeks". But I mean, if we take arbitrary dead tissue, that's confounding the experiment. Ischemia causes apoptosis and we're studying apoptosis! Not cool. I mean I see what he's saying, but damn. He suggested looking into using, say, sheep's blood. That shit is cheap. He also said using qPCR was unnecessary. Well, everyone uses qPCR now. Band brightness is a shitty way of determining upregulation vs. downregulation. I mean, I know that the qPCR machine is in heavy use by people doing their theses and the cytogreen dye is expensive, but it's really hard to make any sort of real interpretation of anything by band brightness and no one really does it anymore.

So here we are. We tried to come up with a cheap, fisable experiment and got shut down. All due to culturing a few fibroblasts. Maybe we can still do this experiment with sheep's blood? Or maybe the cell bio prof has a few extra cell lines floating around her incubators?

In a similar fashion, anyone who wants to do a Western Blot pretty much, well, can't unless they are studying a protein that other people in the department are studying.

This shit is hard.