Potential pitfalls of <scp>CRISPR</scp>/Cas9‐mediated genome editing

<jats:p>Recently, a novel technique named the clustered regularly interspaced short palindromic repeat (<jats:styled-content style="fixed-case">CRISPR</jats:styled-content>)/<jats:styled-content style="fixed-case">CRISPR</jats:styled-content>‐associated protein (Cas)9 system has been rapidly developed. This genome editing tool has improved our ability tremendously with respect to exploring the pathogenesis of diseases and correcting disease mutations, as well as phenotypes. With a short guide <jats:styled-content style="fixed-case">RNA</jats:styled-content>, Cas9 can be precisely directed to target sites, and functions as an endonuclease to efficiently produce breaks in <jats:styled-content style="fixed-case">DNA</jats:styled-content> double strands. Over the past 30 years, <jats:styled-content style="fixed-case">CRISPR</jats:styled-content> has evolved from the ‘curious sequences of unknown biological function’ into a promising genome editing tool. As a result of the incessant development in the <jats:styled-content style="fixed-case">CRISPR</jats:styled-content>/Cas9 system, Cas9 co‐expressed with custom guide <jats:styled-content style="fixed-case">RNA</jats:styled-content>s has been successfully used in a variety of cells and organisms. This genome editing technology can also be applied to synthetic biology, functional genomic screening, transcriptional modulation and gene therapy. However, although <jats:styled-content style="fixed-case">CRISPR</jats:styled-content>/Cas9 has a broad range of action in science, there are several aspects that affect its efficiency and specificity, including Cas9 activity, target site selection and short guide <jats:styled-content style="fixed-case">RNA</jats:styled-content> design, delivery methods, off‐target effects and the incidence of homology‐directed repair. In the present review, we highlight the factors that affect the utilization of <jats:styled-content style="fixed-case">CRISPR</jats:styled-content>/Cas9, as well as possible strategies for handling any problems. Addressing these issues will allow us to take better advantage of this technique. In addition, we also review the history and rapid development of the <jats:styled-content style="fixed-case">CRISPR</jats:styled-content>/Cas system from the time of its initial discovery in 2012.</jats:p>