Strategies for the improvement of genome editing in Arabidopsis thaliana

Increasing the efficiency\nof gene targeting (GT) as a genome editing tool in plants has been an important\ngoal in plant biotechnology. Improvements have been made using\nsequence-specific nucleases such as CRISPR/Cas9 to induce DNA double strand\nbreaks at target loci and activate repair via homologous recombination (HR). GT\ncan then be achieved by HR-mediated integration of an artificial repair\ntemplate, sharing homology with the target locus. Further improvements have\nbeen made with the in planta GT method, in which the repair template is\npre-inserted in the genome and can be excised by nucleases. Although these\nimprovements led to substantial increases in GT efficiency, GT is still not\nefficient enough to be feasible for crop biotechnology. This thesis describes\nstrategies to further improve GT efficiency in the model plant Arabidopsis\nthaliana. One of these strategies was to perform in planta GT in meiocytes,\ncells that already have a higher rate of HR. Another strategy was to find new\nArabidopsis mutants with increased GT frequencies and to identify genes\ninvolved in this phenotype. In the end, this may lead to a better understanding\nof the mechanisms underlying GT and these may be used to realize higher GT\nfrequencies in plants.