![]() Once a targeted DSB has been made, HDR may reconstruct the cleaved DNA using an exogenous DNA template analog to the break site sequence. 8 After it was discovered that DSBs could raise the incidence of HDR by multiple orders of magnitude, targeted nucleases have been found as an alternative approach to increase the efficiency of HDR-mediated genetic alteration. ![]() Historically, homologous recombination (HR), in which undamaged homologous DNA fragments are used as templates, has been the approach to realize targeted gene addition, replacement, or inactivation however, the utility of HR is heavily limited due to its inefficiency in mammalian cells and model organisms. 5, 6 Nuclease-induced DNA DSBs can be repaired by one of the two major mechanisms that occur in almost all cell types and organisms: homology-directed repair (HDR) and nonhomologous end-joining (NHEJ), 7 resulting in targeted integration or gene disruptions, respectively (Fig. ![]() ![]() 3, 4 Targeted DNA alterations begin from the generation of nuclease-induced double-stranded breaks (DSBs), which leads to the stimulation of highly efficient recombination mechanisms of cellular DNA in mammalian cells. 2 Genome editing can be achieved in vitro or in vivo by delivering the editing machinery in situ, which powerfully adds, ablates and “corrects” genes as well as performs other highly targeted genomic modifications. 1 Based on engineered or bacterial nucleases, genome editing technologies have been developed at a rapid pace over the past 10 years and have begun to show extraordinary utility in various fields, ranging from basic research to applied biotechnology and biomedical research. In the 1970s, the development of genetic engineering (manipulation of DNA or RNA) established a novel frontier in genome editing. Over the last few years, the exuberant development of genome editing has revolutionized research on the human genome, which has enabled investigators to better understand the contribution of a single-gene product to a disease in an organism. Finally, we provide an overview of the clinical trials applying genome editing platforms for disease treatment and some of the challenges in the implementation of this technology. Here, we review recent advances of the three major genome editing technologies (ZFNs, TALENs, and CRISPR/Cas9) and discuss the applications of their derivative reagents as gene editing tools in various human diseases and potential future therapies, focusing on eukaryotic cells and animal models. Recent progress in developing programmable nucleases, such as zinc-finger nucleases (ZFNs), transcription activator-like effector nucleases (TALENs) and clustered regularly interspaced short palindromic repeat (CRISPR)–Cas-associated nucleases, has greatly expedited the progress of gene editing from concept to clinical practice. Genome editing has extended our ability to elucidate the contribution of genetics to disease by promoting the creation of more accurate cellular and animal models of pathological processes and has begun to show extraordinary potential in a variety of fields, ranging from basic research to applied biotechnology and biomedical research. Based on engineered or bacterial nucleases, the development of genome editing technologies has opened up the possibility of directly targeting and modifying genomic sequences in almost all eukaryotic cells.
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