Hello everyone. My name is Isabelle Massaro and like Antolette, I am fortunate to be one of the local California State University, San Marcos (CSUSM) students accepted into the NFS REU Research Program this summer. I am a senior here at CSUSM (with only 1 more semester left!) and am majoring in Molecular and Cellular Biology with a minor in Chemistry. As I near the end of my bachelor's degree, I plan to continue my education to pursue my passion in plant biology research. Since I joined Dr. Escobar's molecular plant biology lab, I have gained hands-on experience working with plants. My project in the Escobar lab focuses on working with the model plant, Arabidopsis thaliana, to help characterize class III glutaredoxins (GRXs) and identify their role in plant growth and development. GRXs are small oxidoreductase enzymes that reduce disulfide bonds in target proteins. The interesting thing about class III GRXs, is that they are exclusively found in land plants, but most remain functionally uncharacterized. To help identify the biological function of these genes, we have created different transgenic plant lines that either overproduce class III glutaredoxins, or have knockout mutations in class III glutaredoxin genes. For instance, this summer I have been growing two knockout lines that have five GRX genes (AtGRXS3/4/5/7/8) inactivated and two overexpression lines that constitutively overexpresses AtGRXS8. We previously found that this cluster of class III GRXs are significantly upregulated by nitrogen in the soil, specifically in the form of nitrate. This is important because nitrogen is essential for plant growth and development and without nitrogen, plants can not produce DNA, RNA, proteins, or chlorophyll. This summer, my main focus has been on extracting RNA from hydroponically grown transgenic plants with the end goal of sequencing the whole transcriptome using Next-Generation Sequencing (NGS). This will help identify how our GRXs influence patterns of gene expression in plants provided with nitrate as a nitrogen source. The start of the summer involved lots of tissue grinding, RNA extractions, gels for quality control, and cDNA synthesis. This week was especially exciting because we were finally able to start performing Real-Time PCR runs on all of the samples. From these results, we can identify differences in gene expression between our transgenic lines and the wildtype for some specific nitrate transporter genes in roots and shoot growth genes. These results will also help us identify the best samples to send for RNA sequencing. In addition to the molecular work that I have been doing, we have also been growing lots of plants on soil for phenotyping. This allows us to better understand how GRXs affect overall plant growth. For instance, our overexpression plants have significantly smaller shoots compared to the wildtype, which is why we hypothesize that these GRXs genes are negative regulators for shoot growth. As for phenotyping, I have been tracking flower production, weighing total shoot biomass, and imaging pavement cells. And within the next couple of weeks, we plan to complete total nitrate and protein content analyses to better understand how efficiently our transgenic plants store and utilize nitrogen. Overall this allows us to help characterize these class III GRXs and their role in plant growth and development. At the end of the week, we were finally able to go tide pooling! Low tide this summer has been a little tricky to plan around, since it was always very early in the morning. That did not deter us though, and a group of us got up at 4:00 AM to venture out to the coast. We ending up going to Sunset Cliffs National Park and it was well worth waking up so early. This was probably one of the best tide pooling experiences I have had. We found so many neat creatures that I have never seen before. Immediately when we got there, we found an octopus hiding within the algae, which was a great start to our early tide pool adventure. After that, we also found an eel, a lobster, a beautiful red Hopkins’ rose nudibranch, a delicate brittle star, and a ginormous sea hare. And there were so many crabs, shrimps, and little isopods scurrying everywhere. I am very grateful to be a part of such an incredible cohort this year. This summer has definitely been an amazing experience and I want to thank Dr. Sethuraman and Dr. Read for running an excellent program during these challenging times. Good luck with everyone's research project as we come to the home stretch.
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Watch this space for weekly updates!Every week, one of our CSUSM NSF REU students will post their blurb, summarizing their week, and chronicling our program. AuthorWrite something about yourself. No need to be fancy, just an overview. Archives
August 2023
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