The Principles and Application of Gene Editing as a Tool in Research, Industry and Health Biotechnology

The discovery of zinc finger nucleases (ZNFs) and transcription activator-like effector nucleases (TALENs) has revolutionized the area of biological research, these engineered chimeric nucleases are protein based systems consisting of programmable sequence-specific DNA binding sites linked to a non specific DNA cleavage domain, capable of introducing double stranded breaks (DBS) or single strand nick (SSN) in organisms� genomic sites, thereby activating DNA damage response pathways homologous recombination (HR) or non-homologous end joining (NHEJ). These error prone DNA repair pathways, in the presence of donor sequences can introduce insertion/ deletion mutations (indels) to a targeted genomic site in various industrial or biomedically important cell types however, the recent development of clustered regularly interspaced short palindromic repeats (CRISPR) technology has redefined targeted genome editing owing to its simplicity and efficiency, adopted from bacterial and archaeal acquired immune system, CRISPR/Cas9 system is a complex of customizable single guide RNA fused to Cas9 nuclease which uses simple base pairing rules between the guide RNA and DNA at the target genomic site to mediate cleavage of complementary target-DNA sequences adjacent to protospacer adjacent motifs (PAM). Introduction of DBS or SSN by CRISPR/Cas9 system also induces the indigenous error-prone DNA repair pathways HR and NHEJ, thereby facilitating indels mutations or gene knockout, mutant Cas9 can mediate gene regulation. Although engineering of ZFNs has been challenging and the specificity of CRISPR/Cas9 across wide range of genomes has not been well established, the application of these genome editing tools promises a great future in research, industry and health biotechnology.