UCSF

Since the discovery that CRISPR-Cas9 is an RNA-guided DNA-endonuclease and can perform programmable cutting, the genome engineering field has moved from making a simple double-stranded break towards performing a precise edit, where you change the identity of one or many nucleotides to another. However, making a precise repair requires a template for that repair, often made out of single-stranded DNA. In this dissertation, I will detail optimization of one such tool for intracellular DNA production, the bacterial retron, and its utilization as a template for precise repair in: eukaryotes (Chapter 2), bacteriophage (Chapter 3), and then high-throughput libraries for higher rates of precise editing in human cells (Chapter 4).