Graduate School of Biomedical Science and Engineering, Hanyang University, Seoul, South Korea.
Department of Food Biotechnology, College of Medical and Life Science, Silla University, Seoul, South Korea.
Prog Mol Biol Transl Sci. 2018;159:157-176. doi: 10.1016/bs.pmbts.2018.05.001. Epub 2018 Jun 12.
Clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated system (Cas) is comprised of repetitive bases followed by short fragments of DNA from a previously invading organism that provide immunity to the most prokaryotic organisms. An RNA-dependent spacer is required for CRISPR/Cas9 to recognize the target DNA. Delivery of the CRISPR/Cas9-guide RNA (gRNA) complex to any cell results in modification of the target sequence. The CRISPR/Cas9-mediated genome editing technique is currently in the spotlight and has several research interests, including molecular medicine and agriculture. There are several factors that hinder the delivery of this complex, such as the large size of the plasmid or high dosage of the chemical agent. There are several methods available to deliver CRISPR/Cas9 and its components to the target cells. It includes viral, non-viral and physical methods to deliver plasmid or ribonucleoprotein (RNP) of CRISPR components. But in vivo CRISPR/Cas9 delivery remains challenging to the researchers due to insertional mutagenesis, targeted delivery, immunogenicity, and off-targets. However, studies suggesting that the CRISPR/Cas9-RNP delivery can overcome these hurdles. Here, we review the various methods for delivery of CRISPR/Cas9 and gRNA to several cell lines, highlighting the limitations of each approach, and suggest possible alternative methods.
成簇规律间隔短回文重复序列 (CRISPR)/CRISPR 相关系统 (Cas) 由重复碱基组成，后面跟着来自先前入侵生物体的短 DNA 片段，为大多数原核生物提供免疫力。CRISPR/Cas9 需要依赖 RNA 的间隔子来识别靶 DNA。将 CRISPR/Cas9 指导 RNA (gRNA) 复合物递送到任何细胞都会导致靶序列的修饰。CRISPR/Cas9 介导的基因组编辑技术目前备受关注，具有许多研究兴趣，包括分子医学和农业。有几个因素会阻碍该复合物的递送，例如质粒的大小或化学试剂的高剂量。有几种方法可将 CRISPR/Cas9 及其组件递送到靶细胞。它包括病毒、非病毒和物理方法来递送质粒或 CRISPR 组件的核糖核蛋白 (RNP)。但由于插入突变、靶向递送、免疫原性和脱靶效应，体内 CRISPR/Cas9 递送仍然对研究人员具有挑战性。然而，有研究表明，CRISPR/Cas9-RNP 递送可以克服这些障碍。在这里，我们综述了将 CRISPR/Cas9 和 gRNA 递送到几种细胞系的各种方法，强调了每种方法的局限性，并提出了可能的替代方法。
