Bao, Gang2020-08-182021-02-012020-082020-08-12August 202Li, Ang. "Development and Characterization of Viral-Based Gene Editing In Vivo." (2020) Diss., Rice University. <a href="https://hdl.handle.net/1911/109234">https://hdl.handle.net/1911/109234</a>.https://hdl.handle.net/1911/109234Adeno-Associated Viral (AAV) vectors packaging the CRISPR/Cas9 system (AAVCRISPR) can efficiently modify disease-relevant genes in somatic tissues with high efficiency. AAV vectors are a preferred delivery vehicle for tissue-directed gene therapy because of their ability to achieve sustained expression from largely non-integrating episomal genomes. However, for genome editing applications, permanent expression of non-human proteins such as the bacterially-derived Cas9 nuclease is undesirable. Recent studies indicate a high prevalence of neutralizing antibodies and T-cells specific to the commonly used Cas9 orthologs from Streptococcus pyogenes (SpCas9) and Staphylococcus aureus (SaCas9) in humans. Additionally, persistent expression of CRISPR/Cas9 has the potential to increase the chances of off-target cutting. There is a need for efficient genome editing in vivo, with controlled transient expression of CRISPR/Cas9. The topic of my thesis covers the development of a self-deleting AAVCRISPR system that introduces insertion and deletion mutations into AAV episomes, understanding the effects AAV-CRISPR editing in vivo in a Cas9 immunized mouse model, and the characterization of AAV integrations into the genome in the context of CRISPR-based gene editing.application/pdfengCopyright is held by the author, unless otherwise indicated. Permission to reuse, publish, or reproduce the work beyond the bounds of fair use or other exemptions to copyright law must be obtained from the copyright holder.Gene EditingAdeno-Associated VirusGene TherapyCRISPR/Cas9Thesis2020-08-18