Targeted In Situ Gene Correction of Dysfunctional APOE Alleles to Produce Atheroprotective Plasma ApoE3 Protein

Figure 5

Double-strand breaks (DSBs) stimulate homologous recombination (HR) repair. HR facilitates exchange of DNA sequence between donor and acceptor molecules, provided they share a certain amount of sequence similarity. Donor DNA molecules are designed to contain a central “repair” sequence, flanked by “homology” arms, whose sequence is identical to that of the acceptor (e.g., genomic) DNA. Through HR, the repair sequence replaces the entire sequence lying between the crossover points (marked with an X) on the acceptor. Gene targeting exploits HR in this way to make genetic changes to a cell’s DNA. Importantly, introduction of a DSB into the acceptor sequence, which can be achieved by adding an engineered nuclease with locus-specific cleavage (see Figure 6), markedly increases gene repair efficiency.