Kwon Jiwoong, Park Yeji, Ha Taekjip
Program in Cellular and Molecular Medicine, Boston Children's Hospital, Boston, MA, USA.
Department of Pediatrics, Harvard Medical School, Boston, MA, USA.
Exp Mol Med. 2025 Jul;57(7):1400-1408. doi: 10.1038/s12276-025-01434-z. Epub 2025 Jul 31.
CRISPR-based imaging technologies have emerged as powerful tools for visualizing specific genomic loci, providing groundbreaking insights into chromatin structure and dynamics. Here, in this Review, we discuss the development and recent advances in these techniques, highlighting key strategies such as signal amplification, background reduction, multiplexing and enhanced genomic resolution. By engineering Cas proteins and guide RNAs, and incorporating peptide and aptamer tags, researchers have remarkably improved the sensitivity, specificity and resolution of CRISPR-based imaging, enabling the detection of nonrepetitive genomic regions and single-nucleotide polymorphisms. Recent studies have further pushed the boundaries of CRISPR-based imaging with the introduction of degron-mediated fluorogenic labeling and light-controllable background reduction. Despite remaining challenges, such as the bulkiness of signal amplification systems, limitations in guide RNA design and the effects of fixation on chromatin-protein interactions, CRISPR-based imaging holds great promise for advancing our understanding of chromatin dynamics, genomic interactions and their roles in various biological processes.
基于CRISPR的成像技术已成为可视化特定基因组位点的强大工具，为染色质结构和动力学提供了开创性的见解。在本综述中，我们讨论了这些技术的发展和最新进展，重点介绍了信号放大、背景降低、多重分析和提高基因组分辨率等关键策略。通过对Cas蛋白和引导RNA进行工程改造，并结合肽和适体标签，研究人员显著提高了基于CRISPR成像的灵敏度、特异性和分辨率，能够检测非重复基因组区域和单核苷酸多态性。最近的研究通过引入降解子介导的荧光标记和光控背景降低，进一步拓展了基于CRISPR成像的边界。尽管仍存在挑战，如信号放大系统的庞大、引导RNA设计的局限性以及固定对染色质-蛋白质相互作用的影响，但基于CRISPR的成像在推进我们对染色质动力学、基因组相互作用及其在各种生物过程中的作用的理解方面具有巨大潜力。
