Thoughts on Theses: Ashwin Balaji

Ashwin Balaji is a biophysics-biochemistry major. For his thesis, he is researching loop formation during DNA folding in sperm cells. His adviser is Associate Professor of Physics Ashley Carter.

Q: What does Ashley Carter’s lab study? A: The Carter lab looks at a few different things. Our main focus is understanding how DNA compacts into very small structures. In your cells DNA needs to fit into a very small space — it’s something on the order of a few microns, but DNA inside the human genome is about two meters long. That’s many, many thousands of times of compaction that need to happen. We’re … specifically looking at this compaction in sperm cells. In sperm cells the DNA is bound with this protein called protamine, which then leads to compaction of DNA into first loops of DNA … Then these loops condense further into toroids, these donut-like shapes of DNA, which allows all this genetic information to fit inside the cell. We know of intermediates that form in this process, the loops and the toroids that eventually result, but we don’t know how we go from a straight piece of DNA to a loop to a toroid. The two main questions in the lab right now are: how does a loop form, and then how does a toroid form from these loops? I’m working on the loop formation aspect, and my labmate, Hilary Bediako, also doing a thesis, is doing work on how the toroids form. We have some other projects in the lab. Mainly another thesis student, Peter Cho, is working on behavioral studies with zebrafish. He’s using programming and computer algorithms to monitor the zebrafish.

Q: What methods are you using to look at your question? A: That’s the part I find really exciting. Halfway through the summer we decided to take this new approach that Professor Carter found. [She] talked to someone at a conference about this new apparatus. It’s called a centrifuge force microscope … Actually the bulk of my thesis will probably be getting this apparatus built, because we’re kind of starting from scratch with it. The basis of it is: you can take the DNA and make tethers with them. You stick one end to the glass slide, so it’s anchored there. You can stick a small polystyrene bead to the other end of the DNA … You’re able to see it, so we can use a microscope to actually visualize it, and then take this tethered DNA and spin it in a centrifuge along with our microscope equipment. Because it’s spinning, the DNA starts to stretch out. What we’ll do is take these tethers, have them fold up with protamine. Then we’ll stretch them out to see how they unfold. That’s how we get information about the folding process.

Q: Have you found any results so far? A: Actually in terms of what I’ve just talked about, no, because we’re building the apparatus right now. I did do work over the summer which basically confirms how the DNA is folding. For that we don’t pull on it or anything. We just observe the DNA fold as we flow protamine in. We’ve kind of seen that the DNA folds in a multi-step process, and that loop formation, again, is multi-step. We see these clear intermediates. The tether shortens to one length, and then after a while shortens to another. That shows us that the shortest length is a looping form, but then there’s a state before this that another intermediate to loop formation.

Q: What does a day working your thesis look like? A: It’s mostly spent on the phone or on the computer ordering parts, talking with representatives and sale reps to get all the parts we need for this apparatus. That’s been kind of interesting to see that side of the science — getting all of our materials ready and here before we can start to build, which is something I feel like a lot of students may not get to experience because everything’s been set up. It’s been really unique starting a project from scratch … and building up from basically nothing until hopefully at the end of this year we have a working, functioning apparatus that gives us data and information and the tools to answer the questions we’re going after.

Q: What draws you to biophysics research? A: I’m really interested in the way things move, and the way they generate force in mechanical interactions. I thought these types of things didn’t happen at the molecular level, at the nanoscale. When I started realizing that there are … things that act very much like machines and motors and rotors that we see in the physical, macroscopic world at the microscopic level, that got me really excited to see how these things move and how biology has coded for such machines — and such logical, smart machines — at the nanoscale.

Q: Do you want to keep doing research in the future? A: Yes I do, actually. I’ll be applying to graduate schools at the end of this year. Hopefully something there pans out and I’ll be able to keep doing single molecule biophysics research down the line.

Q: What advice do you have for future thesis writers? A: I would say really don’t be afraid to take ownership of your project and, within the boundaries of safety and things like that, just take those few next steps, even though you may not have discussed them exactly thoroughly with an advisor. Start doing that kind of thing on your own. Of course, when you get the time, do go through it with your advisor, but I think you’ll learn a lot more when you start acting and thinking for yourself. Try things out, and then when you really get stuck, go to your advisor and see what the next steps might be.

Q: What is your research process when you run into an issue and you have to start thinking independently? A: … Really it’s just breaking down your problem or your apparatus into small discrete units … It requires you to intimately know what your apparatus is and how things work there … Then finetune and change individual parameters and see if anything changes. If something does, you know that’s the source of your problem. It’s really about thinking through what all goes into what you’re trying to do, and then isolating each one of those factors so you can see where the problem is.