Adventures in PhuDland: Effing up

I knew that in the course of attempting to do some actual science I would fuck something up. Once again, ladies and gentlemen, Kristie does not disappoint. First mistake I made was to forget just how weird cholera bacteria are. They live in the environment, which is normally quite a bit cooler than body temperature (except for the last week or two), so you'd think that you could toss a Vibrio culture in the fridge and everything should be just fine. Not so. Vibrio cholerae is not only a genomic hoarder, with two chromosomes, but it's more thermopersnickety than a woman with a malfunctioning thyroid gland- it really doesn't like to be cold. So, I grew overnight Vibrio cultures for my fish infections last week and, not thinking, took them out of the incubator and popped them in the fridge while I finished up some other stuff and went to lunch. They were in the fridge for two hours at the most. When I got back, I took them out of the fridge and was spotted doing so by two of my fellow cholera cult (lab) mates:

"That wasn't Vibrio you just pulled out of the fridge, was it?"

"Uh...yeah." Uh oh...

"You know you might have just killed your cultures, right? Vibrio dies if you put it in the fridge."

Shit! "Oh, right. I totally forgot. Well, I guess we'll see if two hours is enough to kill them!"

Even though I felt like a complete moron, I did still have the presence of mind to plate a couple of dilutions of each of my cultures, just to make sure I really didn't kill them. When I infected my fish later that day, I had no idea if I was adding live cholera or just polluting their water with dead useless cells. Dumb, dumb, dumb. At the very least, I figured out how to set up the fish infections on my own - which tank to take fish from, to make sure that they hadn't been fed first, which water/beakers to put them in, how to mix up the tricaine solution to euthanize the fish, etc. Plus all the other stupid little details that get forgotten by people who've been doing this stuff for a while - like where they keep the ethanol, how do I clean up the fish shit when I'm done, where the dissecting kits are...it's getting a little bit easier every day. Oh, and it turns out that I hadn't killed my cholera after all! Sometimes I think a lot of research is figuring out just how much you can fuck things up without ruining the entire experiment. That sounds smart-ass-ish, but I think it's absolutely true. You need to know how much wiggle room you have in your protocols - then you try to figure out why, which sometimes leads you to new side projects. It has been strongly and repeatedly suggested to me (by a favorite cantankerous former professor of mine) that a good student should be working on at least three projects simultaneously (at the time of my scolding, I think I was only juggling two things. And rather lamely, at that).

Tomorrow, I'm going to attempt my first ELISA assay (ever!), looking for cholera toxin in last week's fish water. If it doesn't work, then there's a good possibility I fucked up precipitating protein from the fish water. I really have no idea what the hell I'm doing! I'm also still stuck in cloning hell, but so is everyone else in the lab. We think that the UV transilluminator is frying all of the DNA we try to gel purify - even though I limit the gel exposure to 15 seconds or less, plus we add guanosine to the gel to act as "sunscreen". We're supposed to be getting a blue LED transilluminator soon, I think. Maybe that will help.

I still need to get some work done tonight, but to hold you over until my next post, here are some crappy iPad pictures of the lab.