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. 2020 Jun 5;11:2861. doi: 10.1038/s41467-020-16641-7

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© The Author(s) 2020

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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Fig. 1 — a Genomic DNA sample is dephosphorylated and crRNA-guided target-specific double-stranded cuts are introduced through Cas9. b Phosphorylation-sites are exposed through the double-strand breaks; however, Cas9 remains bound to the PAM-distal side of the cut and blocks phosphorylation of the DNA on this side. DNA-ends are dA-tailed and adapters are ligated only to phosphorylated DNA-ends proximal to the PAM sequence. Sequencing direction is dictated by the adapters, towards the unknown sequence. c Targeted libraries are loaded and sequenced on one ONT flow cell (R9.4). NanoFG is run on the nanopore sequencing data, extracts fusion-spanning reads, detects fusion genes and provides exact fusion gene configuration, breakpoint-location, breakpoint-sequence and fusion-spanning primer sequences.