Just a thought…

It’s been a while since I updated…

However, I did make quite an interesting observation this morning in lab. Terrific broth cultures, after subjecting them to denaturing conditions with GnHCl, are not pleasant to work with. After spinning down my cultures and getting ready to incubate them for purification, I proceeded to add 20% bleach to the tubes to make it easier to clean. Instead, the solution turned a bright orange upon addition of the bleach and it foams and smells really bad.

I let out a squeal because it was nasty. I can’t shake the smell or feel of it right now. Good thing I was wearing gloves because bleach and bacteria are no good combination ever.

Lesson? Lab safety! You never know what two chemicals can do when placed together.

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Save those samples!

I let out the biggest sigh of relief when I saw those three letters in that 1.5 mL eppendorf tube sitting in my box in the -20C freezer.

It has been a couple months since last posting due to the fact that a lot has been going on in my life personally and academically. I have just gotten a chance to sit down, today, with my PI to discuss the project that I am currently working on and all the results that I have gotten.

A time crunch is an understatement for the situation that my project is in. I have been working on this since February, and since the middle of March, my project has been going in circles. Finally, after somewhat conclusive NMR data and yet again unexpected SEC-MALS data, we (my PI and I) have come to the conclusion that quite possibly, something as minor as a His-tag is affecting my entire protein sample.

It would have helped to realize this sooner, but despite all the failures that I have seen in the past couple of months, it is good to know that all my experiments and tests have not been in vain. Though I am going back to step one by transforming my plasmid with my target sequence into E. coli for overexpression, at least I know that what I am about to work on will give me a shot at producing some results at the very least.

My first year at Northwestern is coming to an end and my time in lab has been nothing short of amazing. I have pushed myself to so many limits and cannot have asked for a better lab to join. The people I work with are so incredibly talented and though it often times makes me realize how little I have accomplished in my life and how much of a failure I seem compared against my fellow labmates, I use that to drive me to do what I need to do.

I am motivated and determined to be able to push my project forward. It really is satisfying to see results coming out, to be able to interpret them on your own, and then discuss with your PI in a two-way conversation regarding what the next steps are.

This year has been such a whirlwind but I can say that I am so happy and grateful for the grant that I have been given to allow me to continue my research here during the summer months.

For now, this is it. Bacteria cells await my care. Just know, saving samples is crucial when doing research! Save those samples!

“It looks beautiful”

The first thought that came to my mind when I woke the computer from its sleep state was just one thought. It looks beautiful

I never understood why some people are attracted to research, especially when dealing with molecules such as proteins and DNA that cannot be visualized by the naked eye. It looks like a bunch of liquid, really, as you are going through the various steps of purification. Not to mention, in order to get to this ‘liquid’ phase, you need to go through growing bacteria and harvesting them and getting them — the bacteria, that is — to over express your protein.

And if it doesn’t work, you need to start from step one.

There is a lot of modification that happens in lab. Tweak the concentration of compound A. Add a little more of compound B. Let it incubate longer. Did you wake up your bacteria from the -4C freezer?

Not to mention, you never really know what the proteins are doing in this ‘liquid’ as you are working with it.

But despite all these challenges and the set backs that I have faced in lab, I am still amazed at the way various biochemistry techniques, such as SEC (size exclusion chromatography) can yield such clear, definitive results. If the protein is there, it will elute at the fraction corresponding roughly to its molecular weight. If it isn’t there, or if the protein decided it wants to aggregate and clump together, then the chromatogram will show just that.

There is such a satisfaction with seeing a pure peak, a clear, indicative peak that gives insight into what the protein in this ‘liquid’ might just be doing.

Its trivial, I know. I never understood it, from the outside looking in, reading journal publications on how researchers have this moment of “ah-hah”, how they see beauty in results. Now I get it. I have seen it from the other side. It truly is beautiful when you see the results you hope for, knowing that something you have done is right.

Back at the bench: in the dugout and in the lab

There is a strange about standing over the pH meter, watching the numbers rise — click, click, they sound. Or so I imagine. One drop too much of NaOH and there goes the beautiful click, click; it is a swoosh, an adrenaline rush.

Time to add HCl. Bring those numbers back down. But one too many drops — even half a drop — and there goes the delicate balance between hydrogen ions and hydroxide ions

There is precision to the work that I do for my independent study course. I am to spend whatever time I wish to in lab, to do my project at my own leisure. Sure, there are ‘deadlines’ I need to meet — have you purified yet? Did you run SEC? How about that protein gel? Do you have enough pellets in the -80C freezer ready to go on any given notice?

Yes. Yes. Yes. I think. No. I actually don’t have my act together. Or I didn’t. Because for the past 10 days or so, since the quarter started, I told myself that I need to focus on me, that I need to start reaching back to the old me — the one that loved to run and workout to stay sane, the girl that found joy in burying herself in her studies.

But that hasn’t worked. Back at the bench since my last class ended, following a day since my 7AM 4-hour emergency room shift, I find myself sitting at my lab desk, staying late into the night, doing one too many tasks. Run the agarose gel to see just exactly why your mutagenesis isn’t working. Incubate the EDTA with your resin in the cold room for 10 minutes before you elute. Collect your fractions from the SEC run and store them so you can run a gel on them later.

There is a list of tasks running through my head. It has been 15 hours since I was last sleeping. I tried to avoid coffee all day but half a cup down and I am going strong.

Fueled not by fear, but by passion. Back at the lab bench, it brings me back to the days when I had a seat on the bench. In the third base dugout, where despite convention of the home team getting the first base dugout, our team was, sun behind us and not in our eyes. Being back at the bench is just like being on the bench, up against the fence, cheering on the team. It is just like grabbing my battered Mizuno glove off the yellow bench, running out as I slip my sunglasses over my eyes, and looking back at the field to gather the sign and position myself, predicting where the ball will go. Because isn’t that what I am doing now? Making predictions and then running an experiment to see if what I think is reality? Because isn’t positioning yourself just slightly on the third base foul line indicating that an inside pitch is being called by the catcher and that the righty at bat will pull it down the line?

At the end of the day, I can tell myself I need this time for this and this time for that. But really, it doesn’t matter what my ”excuses” are, because this is where I feel at home. There is a feeling of serenity and calm that overcomes me as I methodically go through each step. I have my protocol memorized down to how many grams of each chemical I need. Step by step, walking a fine line.

Just like I knew the hitting signals and the fielding cues, I know my protocol. I know that an outside pitch requires me to go with it and poke it just over the first basemen’s head so I can beat the right fielder’s throw to the bag. Just like that. It is a science. Softball and research. Where I am me.

Back to the Grind: 2013

There are certain things that I feel as if I will never be able to wrap my head around. Nor will I ever come to fully understand the implications of many of the things that have been happening in my life recently.

Are there reasons why I am so fortunate for being able to come across this opportunity to research in a wonderful lab? What is the purpose of my deciding to push through with a quarter filled with endless hours performing lab research, understanding my project and its implications, as well as trying to keep up with very time consuming, content filled courses?

I guess this isn’t the place or time to be thinking about all of this. I can say though, that after a week back at school in 2013, I am very overwhelmed and wondering what it is I can cut out from my life in order to really, fully grasp everything this quarter has to offer, as well as push myself and do the best that I know I can do.

Overall, I am looking forward to tackling the project that I have been given by my PI. I feel like it is a great starting point for me to really come to understand how to study eukaryotic transcription at the molecular level, something that is truly fascinating and intriguing. Aside from doing basic preparations I have been reading up on the literature written about the proteins I am working on. There is so much unknown out there that it is amazing how I get to be a part of this discovery process. We may think science has offered us all the answers but to this day, there is still so much unknown, so much uncertainty that only through research and discovery will we be albe to come one step, one baby step, closer to understanding how our body works on the molecular level.