biostudies-literatureUnknown5(2)Marinov GKHuman Frontier Science ProgramNational Institutes of HealthCarnegie Institution for ScienceNSF IOSChan Zuckerberg InitiativeRita Allen FoundationDinoflagellate genomes often are very large and difficult to assemble, which has until recently precluded their analysis with modern functional genomic tools. Here, we present a protocol for mapping three-dimensional (3D) genome organization in dinoflagellates and using it for scaffolding their genome assemblies. We describe steps for crosslinking, nuclear lysis, denaturation, restriction digest, ligation, and DNA shearing and purification. We then detail procedures sequencing library generation and computational analysis, including initial Hi-C read mapping and 3D-DNA scaffolding/assembly correction. For complete details on the use and execution of this protocol, please refer to Marinov et al.¹.STAR protocols102941https://www.ebi.ac.uk/biostudies/studies/S-EPMC10950746biostudies-literatureProtocol for mapping the three-dimensional organization of dinoflagellate genomes.PMC10950746Grossman ARGreenleaf WJMarinov GKKundaje AfalseProtocol for mapping the three-dimensional organization of dinoflagellate genomes.Dinoflagellate genomes often are very large and difficult to assemble, which has until recently precluded their analysis with modern functional genomic tools. Here, we present a protocol for mapping three-dimensional (3D) genome organization in dinoflagellates and using it for scaffolding their genome assemblies. We describe steps for crosslinking, nuclear lysis, denaturation, restriction digest, ligation, and DNA shearing and purification. We then detail procedures sequencing library generation and computational analysis, including initial Hi-C read mapping and 3D-DNA scaffolding/assembly correction. For complete details on the use and execution of this protocol, please refer to Marinov et al.¹.2024-01-01T00:00:00Z2024 Mar2025-04-25T18:54:45.16Z2025-04-06T07:47:26.908ZS-EPMC109507463848389810.1016/j.xpro.2024.102941