Date of Award
Doctor of Philosophy (PhD)
Technological innovation drives scientific discovery, unlocks new avenues of research, and allows us to ask questions in ways that were previously unavailable. With each technological advance, our ability to perturb and explore biological systems has grown in ways previously unimagined. The theme of my thesis is the development of new technologies in biology. To this end, I have worked on three technologies that contribute to the areas of protein sequencing, RNA barcoding for trans-splicing and single-cell applications, and a new method for transcriptional knockdown.
In my first project, digital analysis of proteins by end sequencing (DAPES), we set out to develop a highly parallel, wide dynamic range protein sequencing platform where peptides are fixed on a low background PEG-maleimide surface, probed by t-RNA synthetase probes, and visualized via single-molecule TIRF microscopy. In my second project, internal RNA identifier sequencing, I developed a way to use trans-splicing to barcode endogenous RNA without the need for cell separation or tissue dissociation. While the original goal of this project was to use IRIS to barcode transcripts for single-cell RNA sequencing experiments, an unexpected use of this technology came from the realization that it could also be used as a screen for exonic splice regulatory motifs and RNA trans-splicing molecules that are used for transient RNA therapeutics. My final project was the development of hnRNPA1 recruiting oligonucleotides (AROs), short RNA sequences capable of knocking down target transcript by recruiting the RNA binding protein hnRNPA1. Together, these projects represent significant advances in biotechnology that allow for new ways to probe biological processes and aid in the development and screening of therapeutic compounds.
Chair and Committee
Melendez, Justin Alexander, "Developments in Proteomics, Trans-Splicing Technology And Endogenous Transcript Manipulation" (2021). Arts & Sciences Electronic Theses and Dissertations. 2614.