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Aguirre Rivera J, Mao G, Sabantsev A, Panfilov M, Hou Q, Lindell M, Chanez C, Ritort F, Jinek M, Deindl S
Science 385 (6711) 892-898 [2024-08-23; online 2024-08-22]
Single-molecule techniques are ideally poised to characterize complex dynamics but are typically limited to investigating a small number of different samples. However, a large sequence or chemical space often needs to be explored to derive a comprehensive understanding of complex biological processes. Here we describe multiplexed single-molecule characterization at the library scale (MUSCLE), a method that combines single-molecule fluorescence microscopy with next-generation sequencing to enable highly multiplexed observations of complex dynamics. We comprehensively profiled the sequence dependence of DNA hairpin properties and Cas9-induced target DNA unwinding-rewinding dynamics. The ability to explore a large sequence space for Cas9 allowed us to identify a number of target sequences with unexpected behaviors. We envision that MUSCLE will enable the mechanistic exploration of many fundamental biological processes.
NGI Short read [Technology development]
NGI Uppsala (SNP&SEQ Technology Platform) [Technology development]
National Genomics Infrastructure [Technology development]
PubMed 39172826
DOI 10.1126/science.adn5371
Crossref 10.1126/science.adn5371