Fecha de publicación: -- Fuente: WIPO "tomato" The CRISPR-Cas9 system induces DNA double-strand breaks (DSBs) at targeted sites to activate mainly two competitive DNA repair pathways: homology-directed repair (HDR) which allows precise editing with a homologous DNA template, and non-conservative NHEJ/MMEJ pathways which connect two ends of the broken DNA and are often accompanied by random insertions and deletions (InDels). The relatively low efficiency of HDR compared to NHEJ/MMEJ has been a major bottleneck in achieving precise gene editing at desired loci. Here, the inventors describe the development of new efficient CRISPR-Cas9 tools, named ZIP- editors, that carry a donor repair template annealed to a modified gRNA allowing RNP stability. This specific heteroduplex gRNA-donor repair template is very easy to design. It does not require any specialized and/or expensive chemical modifications nor modified Cas9. The simplicity of using unmodified Cas9, donor repair template and easily-available modified gRNA allows its easy transferability to all laboratories. Importantly, it is versatile, allowing increased HDR for many targets, in cell lines and primary cells, with nuclease and unlocking HDR using safer single nickase.