Yuki Noguchi, Risa Matsui, Jaeyeon Suh, Yu Dou, Jun Suzuki
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引用次数: 0
摘要
全基因组筛选是全面了解生物现象分子机制的有效方法。然而,尽管该方法在过去几十年中被广泛应用于各种生物靶标,但将其应用于具有时间性和可逆生物输出的生化反应仍是一项艰巨的挑战。为了揭示各种生化反应背后的分子机制,我们最近开发了复兴筛选方法,该方法将基于流式细胞仪的细胞分选与从收集的细胞中重建文库相结合。事实证明,我们对传统全基因组筛选技术的改进成功地揭示了相关生化反应的分子机制。在本文中,我们将阐明复兴筛选的技术基础,重点介绍其在 CRISPR-Cas9 单导 RNA (sgRNA) 文库和互补 DNA (cDNA) 文库筛选中的应用。最后,我们还讨论了全基因组筛选的未来,同时介绍了体外和体内筛选的最新成果。
Genome-Wide Screening Approaches for Biochemical Reactions Independent of Cell Growth
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 and complementary DNA (cDNA) library screening. Finally, we also discuss the future of genome-wide screening while describing recent achievements from in vitro and in vivo screening.
期刊介绍:
Since its inception in 2000, the Annual Review of Genomics and Human Genetics has been dedicated to showcasing significant developments in genomics as they pertain to human genetics and the human genome. The journal emphasizes genomic technology, genome structure and function, genetic modification, human variation and population genetics, human evolution, and various aspects of human genetic diseases, including individualized medicine.