Learn ALL the things

If I were to characterize this week in one sentence, I’d say that I spent it learning as much as possible.  Luckily for me, most of the learning done was hands-on, rather than theoretical, so I got to spend a good deal of time in lab.  Unluckily for me, it seems that everytime I learn something new I completely forget about something else that I used to know.  I wish it were as easy to clean out my brain as it is to clean out my email inbox.  Unfortunately, that’s not the case, so I’ll have to continue stuffing as many things in my brain as possible, and hoping that the right things stick at the right time.  I’ll let you know how it goes... my fingers are crossed for positive results. :P

The first new thing I learned this week was actually a mix of new and old things.  This is a lot easier to explain with scientific jargon, but I’ve been reliably informed by my GRE prep book that I should avoid using jargon in order to not confuse my audience, so I’m going to try to use normal words... if it’s still super confusing, let me know and I’ll try to explain it a different way.

Okay, here goes: the basis of what I’ve been doing so far is designing a specific piece of circular DNA that’s mutated in a certain way, such that a specific DNA repair pathway will repair that mutation, but the other DNA repair pathways won’t.  Previously, the circular DNA that we’ve been using has been fairly resilient to things that disrupt the bonds between the DNA bases.  My project for the summer and for my thesis class in the fall uses a piece of circular DNA that is much less resilient, and can be easily disrupted by some of the chemicals that we would normally use to purify the DNA or to extract it from the gel.  As a result, we have to use a special kind of agarose gel that has a particularly low melting point, so that we can essentially just melt the gel and get the DNA back out of it.

I say that we’re essentially melting the gel to get the DNA back... I sincerely wish that it were that simple.  Low melting point agarose is extremely soft as compared to normal agarose (agarose is the type of gelatin-y substance that we use to ‘run out’ the DNA... basically it’s a porous material that the DNA moves through. Big DNA pieces run slower than small DNA pieces, so the differently sized pieces of DNA get separated out when you run the gel).  In this case, extremely soft means ‘liable to fall apart in your hands’ soft.  Which is something of a problem when you need to carry your gel around the lab in order to take pictures of it (to see if the bands are where they’re supposed to be) and cut out the bands (so that you can melt just the part of the gel that has the DNA band you want, not the whole gel).  Luckily we can more or less get around that problem by carrying the gel around on the tray that it sits in when you’re running it.  Unfortunately, cutting the band out of an extremely soft gel is also ridiculously difficult.  In the normal gels, it’s pretty easy to cut out only the part of the gel that you need, and to shave off fairly thin slices of agarose gel when you make a mistake in cutting.  In the low melting point gels, the tiny pieces of gel tend to stick together, making it incredibly difficult to fix your mistakes.  As a result, the process is a lot slower than it would be otherwise, and a lot more fine motor skills are required.  I don’t know about you, but I’m still working on perfecting my fine motor skills while wearing gloves and working at 4°C (39°F... not extremely cold, but not pleasant for long periods of time or while trying to do delicate work).  The point of this long diatribe on low melting point agarose is that it’s completely necessary, and not very complicated, but it’s a new thing that I have to get used to, and not all that fun. :P

After the DNA piece is cut out of the gel, the process to extract the DNA from low melting point agarose is also a lot longer than the process for normal agarose.  Where the normal steps take about an hour at most, the low melting point agarose steps take the better part of a day.  And when you include all of the steps necessary to actually make the DNA piece that we want in the first place, the entire process spans the better part of three days... which explains a bit of why I stayed in lab late for two days last week. It also gives me a new perspective on how costly science is.  I knew from last summer and various parts of last year that the monetary cost of science was extremely high- a miniscule amount of the antibody needed to look at the proteins I was trying to find last summer can run upwards of $400 (and that’s for 100 µL. For perspective, a single dose of Nyquil is about 30 mL, which is 3000 times as much as the amount of antibody you get for $400. Science is almost stupidly expensive.).  This summer has given me a new perspective on the man-hour costs of science.  Last week, I probably put in about 20 hours of work to get less than half of the DNA that we had put into the gel back out of it.  And honestly, that was a good result.  In the same way that some energy is always lost to do work on a system, some of the materials we’re working with are always lost to get to the point where we can use them.  And it’s both incredibly frustrating and fairly rewarding to see a final yield of less than 50%- on the one hand, so much of the work you put in to make the right DNA was just lost without a trace; on the other hand, 50% is pretty close to the upper limit of what it’s possible to recover, so at least I’m not doing worse than anyone else who’s following the same protocol.

All of this goes to say that low melting point gels are currently the extremely necessary but extremely annoying center of my scientific existence. :P

Anyway, moving on from gels, I also did other things in lab last week.  The main thing that I did other than playing with horrible, horrible gels, was learning to operate a flow cytometer.  This was a little bit terrifying, largely because flow cytometers play into the monetary section of how much science costs.  The list price isn’t available online (which probably tells you something about how much these things cost...) but the figure they gave us during orientation was somewhere in the multi-million dollar range.  Given that I have a small tendency toward breaking machinery when I touch it, I was rather nervous about running the flow cytometer.  Despite my nervousness, the training went pretty well, and I’m now a certified flow cytometer user at MIT.  UROP students normally aren’t allowed to run the flow cytometers (probably because they’re such expensive pieces of equipment), but since I’ll need it for my thesis class in the fall, they let it slide.  Hopefully at some point I’ll have some interesting data that I can share... right now all I have are dots that represent cells that were engineered to glow green when a laser was shined on them. Which isn’t all that exciting, but since this post is currently a long wall of text, I’ll include them below so that you have something mildly interesting to look at. :P

This is a typical plot showing the relative size of the cells. The axes (forward and side scatter) refer to how much of the laser light is bounced either to the side or forward rather than being stopped by the cell. This is a not-particularly-exact measurement of the size and shape of the cell. The cells inside the black pentagon are single cells that are probably alive still, and have a normal shape.

These are cells that have been engineered with a piece of DNA that causes them to make a protein that glows green when a laser shines on it. This time, the dots in the triangular section (labeled R2) are those that glowed green- the rest of the cells had some green fluorescence, but it was due to the auto-fluorescence of the cells themselves. (I know, cells have auto-fluorescence. It's weird, right?)

The last learning-related thing I did this week was studying for the GRE.  Like I’ve mentioned before, this is the entrance exam for grad school.  I can’t remember if I’ve said how hard it is though... I’ve done several practice sets, and I think I’ve come across more words that I don’t know than words that I do.  Since vocabulary plays a big role in the Verbal section of the GRE, we’ve been coming up with a vocab word of the day for most of the summer. Somehow, there’s still a mountain of words that I don’t know, though... I thought I had a decent vocabulary, but the GRE is reminding me that I’m at a tech school and haven’t seriously studied literature or English-based classes for three years.  Unfortunately, the GRE isn’t testing me on things that I’ve been focused on (biology and the like), but rather basic math and verbal skills.  So if anyone has a good way of remembering the difference between permutations and combinations (probability things... *cough*Kerri*cough*) or between insolent and indolent (vocab words that I just can’t keep straight at the moment) definitely let me know.  I could definitely use all the help I can get when it comes to this ridiculous test.

On a more fun note, three friends and I went out for Indian food on Saturday night.  I’m not a huge Indian food person, largely because it’s not really readily available in West Seattle, to the best of my knowledge, but I thoroughly enjoyed dinner.  Trying new things is always good, particularly when my friends who are well versed in Indian food are available to make decisions about what we should eat.  It was also just really nice to get out of the MIT/Sig Ep/grocery store bubble that I’d been in all week.  All of those places are great, for different reasons, and I love spending time in each of them.  But a break from routine in order to take a tiny adventure to a part of Boston I don’t usually get to see was exciting.  The delicious food was a definite benefit as well.  I definitely ate too much, as was evidenced by the fact that I was sound asleep a couple of hours prior to when I normally would be... struck by the proverbial ‘food coma’, as my friends and I call it.

Since this is now 20 minutes late and I’ve got to be up for work in the morning, I’m going to end it here. Next week I’ll hopefully be able to regale you with more stories of my successes in lab, and give you fewer boring explanations of why everything is difficult. :P