CRISPR-Cas9-mediated loss-of-function screens

Abstract

The CRISPR-Cas9 system, which uses sgRNA for targeting and the nuclease Cas9 for cleavage, has emerged as a versatile and efficient tool for genome engineering; the system has overcome the limitations of previous technologies and implements various gene editing strategies. The large-scale loss-of-function (LOF) gene scanning technology based on the CRISPR-Cas9 system can be utilized to reveal associations between the genotype and phenotype by inducing efficient and scalable gene perturbations throughout the whole genome. This technology is playing a breakthrough role in the exploration of genes associated with tumors and viruses and its expansion together with RNA sequencing (RNA-seq) can be used to assess multiplexed gene interactions as well as more complicated phenotypes. Here, we introduce LOF genetic screens based on CRISPR knockout (CRISPR ko) and its applications in synthetic lethality and virus-host interactions. We highlight the recent progress on combining this technique with single cell RNA-seq. We also compare the advantages and pitfalls of CRISPR variants, discuss the future perspectives of gene therapy and raise important considerations regarding off-target effects.