Meeting My Match

            I think I’ve finally met my match with respect to doing lab work in my UROP. Over the summer, I was in a happy little bubble of being able to do everything nicely, as long as I wasn’t doing stupid things and messing up what reagents I was using. While I knew this wouldn’t last forever, I had hoped it would last slightly longer than it did. Alas, it was not meant to be. As it turns out, I’m really good at getting qualitative results. But if you ask me to get anything that needs to be analyzed numerically, it’s a whole different ball game.

            For the past few weeks, I’ve been running RT-PCR reactions in an attempt to analyze the genetic expression of certain genes in the cells that we’re testing. Let’s break this down a little. The way RT-PCR works is that you mix a bunch of reagents (which is a fancy term for chemicals that make reactions happen) together in a test tube, and then add some of each reagent mixture to a 96-well plate that looks like this:

Along with the reagent mixture, you add some cDNA, which is DNA that’s been transcribe from RNA. Basically, the way it normally works is that DNA is transcribed into RNA, which is then translated into proteins, which are used throughout your body for various functions. In this case, RNA is used to transcribe into a form of DNA called cDNA, or complementary DNA, so called because it’s made by putting together the bases that are complementary, or match with, the RNA strands. In general, this is useful because the genome contains a lot of DNA sequences that don’t actually code for anything, and aren’t all that useful. These sequences get removed, through a process called splicing, before the RNA is translated into protein. The theory is that since you’re making cDNA from RNA, the splicing will have already occurred, and only the useful DNA sequences will be put into the cDNA.

Whew. That was a lot of technical stuff... unfortunately, we’re not quite done with it yet. When you mix the cDNA with the reagents and load everything into the 96 well plate, you then stick it in a machine that modulates the temperature such that the cDNA replicates over and over again, creating the piece(s) of DNA indicated by the primer sequence that you put into the reagent mixture. These pieces of DNA are your product, and the computer program records how much of the product there is over time, and does a bunch of calculations with it. (Don’t worry, I’m not going to bore you with how the calculations work.)

Okay. Now we’re done with technical stuff. The fact of the matter is, this test requires a level of precision that qualitative tests don’t. I have my days where my precision is good, but it’s surprisingly difficult to stand for 2+ hours at a lab bench, doing nothing but pipetting reagents and samples together, and still be accurate at the end of the 2 hours when you’re trying to load everything into the plate. And since this is a computer program, it has levels of precision that I couldn’t even hope to achieve. Granted, this means that there’s some amount of flexibility in the test, but not a whole lot.

SO, I’ve basically spent the last few weeks trying to be computer-level accurate, which seems to be more or less impossible. But it can’t actually be impossible, because people run these RT-PCR tests all the time... so there’s got to be some way to do it well, right? It’s mostly just frustrating because things went well over the summer, more or less, and now I’ve found one test that I just can’t conquer. For whatever reason, and despite the fact that I’ve run the test multiple times over the course of the semester so far, it refuses to work. My grad student keeps telling me that it’s just a matter of practice though... so I’m going to keep going with this and hope that it eventually works. And not just for the sake of my sanity, but for the lab’s sake as well- the reaction plates and reagents aren’t cheap! :P (Not that I think my using a few more reaction plates is going to break the bank or anything... but you never know.)

In other news, I do actually occasionally have a life outside of schoolwork and UROPing, as evidenced by the Roadkill Buffet show I went to with a couple of friends on Friday night. Roadkill Buffet is our improv comedy group on campus, and they’re really, really funny. Improv comedy basically works by setting up a couple of scenarios, with audience input, and having the actors act out the scenes. Since they’ve never seen these scenarios before, they have to think on their feet and come up with possibilities while we, the audience, are watching. It’s not something that a lot of people can do, but when you can do it, it’s hilarious.

This week, I’m looking forward to seeing my family, who are flying out to Boston for my birthday. They’ll be here for about 2.5 days, which isn’t really enough time to see the town, but it’s probably enough to start. And they’ll be able to go out and look at Brandeis, which is one of the schools my sister is looking to apply to. So yay, I’ll get to see my family! :D Next weekend is going to be super crazy though, because my family will be here until Saturday, and then Sunday is Apple Bake, our annual cooking competition in the dorm. I’ve decided that I’m making three dishes (Apple Kuchen, Apple Cinnamon Roll Muffins, and Apple Cinnamon Pull Apart Bread). My suite’s kitchen is going to be a little bit insane on Sunday though... so we’ll see how we get through that. I figure if we can survive Apple Bake, we can probably survive just about anything. (The reason for the predicted insanity is that I think almost everyone in my suite is entering a dish, if not many dishes... so the kitchen will be crowded, to say the least.)

Anyway, I should go do homework, so that I can be prepared for the craziness that the end of this week will probably bring... I hope that you all have a good week!