Graphical Abstract.

Upon the generation of a DNA double strand break (DSB) operated by the Cas9 protein, the dominant negative variant of RNF168 (dnRNF168) recruits but is unable to activate 53BP1 thus creating a barrier to the endorsement of non-homologous end-joining (NHEJ) DNA repair. The simultaneous presence of CtIP facilitates the recruitment of factors essential for engaging the homology-directed repair (HDR) pathway, such as EXO1, hence promoting DNA resection and facilitating the precise repair of the Cas9-induced DSB.