Fig. 2

Precise genome-editing with site specific nucleases. ZFNs create double-strand breaks (DSBs) using the FokI restriction enzyme paired with specific zinc-finger DNA-binding domains. TALENs induce DSBs using the FokI restriction enzyme in conjunction with specific TALE DNA-binding domains. Cas9 nuclease, targeted by guide RNAs, creates DSBs using two distinct domains of nuclease. Genome-editing utilizing nucleases relies on two primary DNA repair pathways. The first pathway consists of end-joining mechanisms, which can be divided into classical nonhomologous end-joining (c-NHEJ), which can be used to produce targeted semi-random indels, and homology-independent target integration (HITI), which can be used to insert an exogenous sequence at a desired genomic target in the absence of homology arms. The second major repair mechanism is homology-directed repair (HDR), which primarily occurs in dividing cells, and can be used to create precise targeted edits via a single-strand or double-strand DNA donor template. ZFNs Zinc-finger nucleases, TALENs Transcription activator-like effector nucleases. This figure was produced using BioRender.com