Genome-Wide Screening Approaches for Biochemical Reactions Independent of Cell Growth.

ABSTRACT

Genome-wide screening is a potent approach for comprehensively understanding the molecular mechanisms of biological phenomena. However, despite its widespread use in the past decades across various biological targets, its application to biochemical reactions with temporal and reversible biological outputs remains a formidable challenge. To uncover the molecular machinery underlying various biochemical reactions, we have recently developed the revival screening method, which combines flow cytometry-based cell sorting with library reconstruction from collected cells. Our refinements to the traditional genome-wide screening technique have proven successful in revealing the molecular machinery of biochemical reactions of interest. In this article, we elucidate the technical basis of revival screening, focusing on its application to CRISPR-Cas9 single guide RNA (sgRNA) library screening. Finally, we also discuss the future of genome-wide screening while describing recent achievements from in vitro and in vivo screening.