Author Archives: Nadeska Montalvan

All good things come to an end… or do they?

Now that BSURF is coming to an end, it’s time to reflect on these past 8 weeks. This experience has been an incredible one. I expected to learn a lot, but ended up learning much more than I could have ever imagined. The funny thing though, is that the more I learn, the less I feel I know. That’s the beauty of science: there’s always so much you don’t know. Once you answer a question, 10 more arise.

I came into the lab very nervous and afraid to mess things up. I came in afraid to ask questions and not confident in my abilities at all. Now, I ask tons of questions on the daily. Now, I am confident in my knowledge of the assays I have learned to run. Now, I know that mess-ups are expected, and even encouraged – how else are we going to learn what works and what doesn’t?

Being in my lab this summer has definitely solidified my interest in research. I always knew I wanted to try research, and now that I have, I definitely see my future self doing research I wasn’t sure if graduate school was something I wanted to do, but this experience has definitely changed my mind.

Overall, I’ve really enjoyed research at the Hargrove lab this summer, and am incredibly grateful for the opportunity. I feel sad to leave the lab, but I feel better knowing that is only a temporary end. I’m excited to continue research here in the fall!

High-Throughput Screening Against RNA Triple Helices: Insights Into Selectivity and Small Molecule Development

Nadeska Montalvan

Mentors: Martina Zafferani1, Amanda E. Hargrove1,2

1Department of Chemistry
2Department of Biochemistry

Out of the total RNA transcribed in cells, only 1.5% is translated into proteins. About 70% of the rest of the human genome is transcribed into non-coding RNA (ncRNA). While the roles of most ncRNAs remain unknown, several ncRNAs have been found to be overexpressed in various cancers which makes them attractive therapeutic targets. Small molecules have been successfully developed to bind to RNAs. However, it remains unclear whether small molecules selectively bind to different RNA sequences with similar structural motifs. Thus, we aimed to screen a large in-house small molecule library against three RNA sequences with triple helix structures, namely MALAT1, NEAT1 and PAN. We hypothesized that most molecules that would bind to one RNA would bind to the other two, indicating low selectivity. We used high-throughput screening to determine which small molecules bound to each RNA. Surprisingly, the screening revealed several small molecules that selectively bind to each of the three triple helices. Additionally, findings yielded a 100-fold higher hit rate than the average rates of high-throughput screenings against general RNA targets. Current efforts are focused on evaluating the newly identified selective small molecules against their respective RNA target to determine any possible unique properties of selective binders.

Cam’s Drug Delivery

This past week in BSURF, we had chalk talks all week. What this meant was that every single BSURFer stood up in front of the rest of us and explained what they are researching this summer while drawing helpful diagrams on a board. As nerve-wracking as it was for everyone, I really enjoyed learning about what everyone was doing this summer.

One chalk talk in particular that stood out to me was Cam’s. Cam’s talk was titled “ELP in Drug Delivery.” I’ve always been very interested in how drugs work in the body, and would love to take classes about these mechanisms in the future. Cam explained how an issue that exists with many drugs in today’s medicinal world is that they require a large dosage because they do not stay in the body for very long. Her research involves trying to create a system that will address this issue to make drug therapy more effective.

She’s doing this through the use of ELP’s or elastin-like polypeptides. ELPs are large and can change solubility, which means they can last longer in the body. She explained how at higher temperatures, ELPs are insoluble, and at lower temperatures they are soluble. This characteristic makes them an interesting target for research because scientists have the potential to modify conditions in a way that will allow ELPs to change solubility in such a way that the drug will last in the body. So, her research involves obtaining, purifying, and using these ELPs to attach proteins to them to hopefully increase the amount of time that these proteins can thus last in the body.

Overall, I think this is such an interesting topic of study and I really enjoyed Cam’s chalk talk! I can’t wait to hear more about how her research is progressing by the end of the summer!

A Day in My Life – Lab Edition

Every morning this summer I take the bus and walk to French Family Science Center and report to my lab, Hargrove lab. I usually walk into the office before the lab itself, where I drop off my backpack in my designated workspace (I have my own one!) and greet all the other members of my lab. I then always meet with my mentor, and I show her any data I analyzed the night before and we have a discussion about where we should proceed next in our experimentation. Now that it is week 4, these discussions are very interesting and my understanding has improved so much that my mentor really values my input!

Next, I proceed to take out all the materials I need for the assays I am going to run. We keep everything in freezers to help preservation, so I always have to brace myself as my hands reach into -20 degrees Celsius freezers and even -80 degrees!

Once I take everything out and let it thaw, the chemistry begins! I usually begin by taking my RNA samples (which vary depending on what assay I am running that day) and nanodrop them. Nanodropping gives me a value of purity for the RNA (RNA purity changes on the daily basis!) which I use to make calculations on how much of each reagent I need to make the solutions I will need that day.

Once I have made all my calculations, I prepare the solutions. This can take a while, as I often need to heat up and ice the RNA solutions. Once the solutions are all made, its time to pipette!

As I am testing whether small molecules bind to specific RNA targets or not, there are usually 2 things that must be added to wells in a microplate for every assay. First, the small molecules are added. I sometimes do this manually, and other times my mentor uses a robot in the medical center to do this. Next, the RNA solutions are added. I always do this manually, and have become faster as time goes on. No matter how fast I get though, this is quite the process. Sometimes I pipette into over 2,000 wells in a day!

Once everything has been added to the microplates, I spin the plates in a centrifuge and leave them sitting for a while to let the reaction play out so that it reaches equilibrium.

Finally, I read the plates on a machine that detects fluorescence. The plates are read one by one, and I always save the data on a flash drive so that I can analyze it later.

Example of a microplate that I prepared.

Me in the lab

Life Missions: Accomplished – A Talk With My PI

I had the pleasure of meeting and interviewing my PI, Dr. Amanda Hargrove, last week. I remember walking into her office very nervous, as I had not been able to meet her yet, but quickly felt comfortable and found Dr. Hargrove to be easy to talk to.

Dr. Hargrove went to college at Trinity University in Texas. She grew up in Houston and wanted to stay near family for college, which I found to be a very endearing detail. She originally wanted to be a doctor, but had a research experience over one summer that changed her mind. She described how she felt that everyone has gifts, and she thought the best way to use hers to help society was be to become a researcher instead of a doctor. I really liked this particular ideology, because I also believe that every person has their own gift. The shift from doctor to scientist also resonated with me because I used to want to be a doctor all throughout middle and high school, but then fell in love with chemistry in my later high school years.

Dr. Hargrove then decided to go to graduate school at the University of Texas, where she continued her research and loved TAing. She then continued on to the California Institute of Technology for her post-doc in chemical biology. Finally, she got a faculty position at Duke where she has been since. Her goal at Duke was to build a lab with undergraduates and graduates students, where everyone feels supported. Based on my experience at her lab so far, I would say Dr. Hargrove has done a great job of accomplishing this goal.

Another goal Dr. Hargrove had was to be a good mother to her children, which she feels she has also accomplished. She described how she chose priorities throughout her life so that not only her research would thrive, but her family life as well. I really like and appreciate that Dr. Hargrove shared this with me, as I also find family to be one of my priorities in life. Hearing Dr. Hargrove share how she was able to accomplish both goals professionally and personally was really inspiring, as I definitely want that for my life in the future as well.

When asked what she didn’t like about science, Dr. Hargrove said she feels that science tends to focus on papers and grants instead of training good scientists or including all good ideas. She also doesn’t like how the people who get grants tend to be the people who have already received grants before, and it’s very hard for those that don’t start of well initially. I thought this was a very interesting answer, and unfortunately, this is true of lots of aspects of society.

On a lighter note, Dr. Hargrove said what she likes about science is discovering things no one knew before or even thought were possible. I share this opinion, as that is also my favorite part of science: the mysteries we have yet to solve. Her favorite part of the job is training students, and her most fun part of the job is helping students realize they discovered something new.

A funny story Dr. Hargrove shared with me was how one time she was working in a lab and she left behind a compound without putting it where it belonged. One of her labmates moved the compound and hid it from her to teach her a lesson about leaving things around. She was very upset when she got back to the lab and couldn’t find it. The funny thing is that when she finally got her compound back, it turns out she hadn’t even made the right compound she needed in the first place and she went through all that frustration for nothing!

All in all, I really enjoyed speaking with Dr. Hargrove. I feel as though I got to know her more on a personal level, and am very excited to keep working with her in the lab.

lncRNA – An unknown world

We all know the basics about RNA and DNA. However, did you know that out of all the RNA that gets transcribed, only 1.5% gets translated into proteins? These types of RNA are referred to as non-coding RNA, or ncRNA. The lab I’m working at this summer, Hargrove Lab, is researching long non-coding RNA, or lncRNA, which is defined as ncRNA that is over 200 nucleotides in length.

LncRNA is a new, unknown, exciting world in the scientific research community. There is not much known about their functions, mechanisms, etc. I find this to be fascinating, as the research I will be doing this summer will help find out more about this hidden, unknown world.

My research project entails screening a large small molecule library (with thousands of small molecules in it) with certain target lncRNA tertiary and quaternary structures. The targets we are investigating are lncRNA structures that are known to have a disease-causing effect in the body. The goal of our research is to find small molecules that effectively bind to the targets in order to be able to manipulate/work with these lncRNA structures.

This can get really tricky though. In addition to the fact that we are looking into so many small molecules, we want to find out not only if they bind, but a plethora of other details as well. Can we find a small molecule that only binds to one specific target and no others (Is it selective)? Does the small molecule bind the same way at different pH levels? Does the small molecule bind better in one concentration than another?

It is a large task at hand, but a fascinating one too!

Pipette who? Never met her.

The level of excitement I felt when I opened and read the email that let me know I would be participating in my first ever research experience this summer was immeasurable. That moment was definitely one of the top ones in terms of top jumping-up-and-down moments in my life. However, once the excitement settled in, I had to face the overwhelming fear that came with knowing I’d be participating in research: what exactly is research?

Sure, I’ve read some research papers and plenty of newspaper articles about research. I’ve always enjoyed reading new scientific findings and I always find myself fascinated with the fact that someone or some people were able to run experiments and studies to reach such incredible findings. After all, that is why I love science so much: there’s always so much to learn, so much left to discover about the world. Thus, I want to be part of that, I want to find more pieces of the puzzle, so to speak.

The question is: how do I do that exactly? Well, stay tuned! This summer, I expect to answer that very question. I hope to gain the hands-on skills necessary for working in a laboratory. By this, I mean I hope to learn everything from being a master of speed with pipetting to using the extremely impressive machines in my lab that I have yet to learn the name of (but plan on doing so soon!). I also hope to know about what the daily life of a researcher looks like; everything from preparing experiments, to sharing data with others, to making mistakes and learning from them.