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.

Committee Chair

Samantha Morris

Committee Members

Thorold Theunissen

Degree

Doctor of Philosophy (PhD)

Author's Department

Biology & Biomedical Sciences (Developmental, Regenerative, & Stem Cell Biology)

Author's School

Graduate School of Arts and Sciences

Document Type

Dissertation

Date of Award

12-18-2024

Language

English (en)

Author's ORCID

https://orcid.org/0000-0003-4826-3574

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