CRISPR/Cas9 technology can be used to investigate how double-strand breaks (DSBs) occurring in constitutive heterochromatin are getting repaired. This technology can be used to induce specific breaks on mouse pericentromeric heterochromatin, by using a guide RNA specific for the major satellite repeats and co-expressing it with Cas9. Those clean DSBs can be visualized later by confocal microscopy. More specifically, immunofluorescence can be used to visualize the main factors of each DSB repair pathway and quantify their percentage and pattern of recruitment at the heterochromatic region.
Keywords: CRISPR/Cas9; DNA repair; DSB; Heterochromatin; Immunofluorescence; Major satellite repeats.