Technology Development of High-resolution Single-cell Lineage Tracing for Cell Reprogramming Systems
ORCID
https://orcid.org/0000-0003-4826-3574
Date of Award
12-18-2024
Degree Name
Doctor of Philosophy (PhD)
Degree Type
Dissertation
Abstract
A critical element of cell identity is the cell that created the observed cell. This parent cell dictates many aspects of its identity, whether measured by function, phenotype, or specific assays. The cell’s lineage impacts the ability of the cell to differentiate into final cell types in development or successfully reprogram in direct cell reprogramming. Here, we evolve molecular barcoding techniques to increase the resolution of lineage recording at single-cell resolution. Using single-cell RNA-sequencing as the readout for this measurement, polyadenylated reporter gene transcripts with engineered 3’ untranslated regions were designed to initially label and then record lineage information. While reporter genes with nucleotide “barcode” libraries have become common, we developed an editable region adjacent to this barcode that is targeted with a novel AIDx-dCas12a base editor. This base editor creates single base transitions on the editable region, enabling information to be written over time. We validated this technology in cell lines to illustrate the resolution of the technology and applied it in cell reprogramming to exemplify how lineage information can contribute to our understanding of biology.
Language
English (en)
Chair and Committee
Samantha Morris
Committee Members
Thorold Theunissen
Recommended Citation
VanHorn, Sadie, "Technology Development of High-resolution Single-cell Lineage Tracing for Cell Reprogramming Systems" (2024). Arts & Sciences Electronic Theses and Dissertations. 3339.
https://openscholarship.wustl.edu/art_sci_etds/3339