Chapter 1 —— 12 —— adenoviral (AdV) and adeno-associated viral (AAV) vectors we postulated that a robust and precise GE system could be assembled by combining the former and latter platforms for delivering CRISPR/Cas9 reagents and donor DNA templates into human cells, respectively. Indeed, contrary to linear uncapped AAV genomes, terminally capped double-stranded AdV genomes are not prone to off-target DNA insertions making them suitable for the delivery of Cas9 enzyme constructs since it is especially important to prevent chromosomal integration of these constructs to minimize the buildup of off-target effects. In addition, also in contrast to AAV vectors, AdV vectors can accommodate full-length Cas9 constructs together with single or multiple gRNAs. Conversely, AAV genomes are proficient substrates for HDR possibly owing to the AAV DNA structure featuring secondary-structured inverted terminal repeats flanking singlestranded DNA. Besides allocating AdV and AAV systems for the delivery of, respectively, Cas9 constructs and distinct types of donor DNA templates, the role of distinct AAV donor DNA structures on the efficiency and accuracy of genome editing was investigated. Hence, in Chapter 4, we put forward and systematically evaluate different iterations of this dual viral vector system in HeLa cells, human mesenchymal stem cells (hMSCs) and skeletal muscle progenitor cells (myoblasts) in terms of their efficiencies, specificities, and accuracies. In this context, we extend earlier observations showing that 3rd generation, fully viral gene-deleted AdV vectors have a notoriously dampened cytotoxicity profile when compared to that of their 2nd generation counterparts. In Chapter 5, we build on the dual viral vector platform introduced in Chapter 4 by investigating its compatibility with a marker-free co-selection system for selecting gene-edited cells and, simultaneously, purging imprecisely edited cells via ouabain selection. The main sub-unit of the ubiquitously expressed sodium/potassium pump (Na+/K+ ATPase) is encoded by ATP1A1 whose ATPase product is responsible for many physiological
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