Nanopore sequencing of DNA concatemers reveals higher-order features of chromatin structure

Abstract

Higher-order chromatin structure arises from the combinatorial physical interactions of many genomic loci. To investigate this aspect of genome architecture we developed Pore-C, which couples chromatin conformation capture with Oxford Nanopore Technologies (ONT) long reads to directly sequence multi-way chromatin contacts without amplification. In GM12878, we demonstrate that the pairwise interaction patterns implicit in Pore-C multi-way contacts are consistent with gold standard Hi-C pairwise contact maps at the compartment, TAD, and loop scales. In addition, Pore-C also detects higher-order chromatin structure at 18.5-fold higher efficiency and greater fidelity than SPRITE, a previously published higher-order chromatin profiling technology. We demonstrate Pore-C's ability to detect and visualize multi-locus hubs associated with histone locus bodies and active / inactive nuclear compartments in GM12878. In the breast cancer cell line HCC1954, Pore-C contacts enable the reconstruction of complex and aneuploid rearranged alleles spanning multiple megabases and chromosomes. Finally, we apply Pore-C to generate a chromosome scale de novo assembly of the HG002 genome. Our results establish Pore-C as the most simple and scalable assay for the genome-wide assessment of combinatorial chromatin interactions, with additional applications for cancer rearrangement reconstruction and de novo genome assembly. Chromatin structure | Structural variation | Long read sequencing | De novo genome assembly | cancer genomicsCorrespondence: mski@mskilab.org

A complex CTCF binding code defines TAD boundary structure and function

“…Existing multi-contact approaches either lack the possibility for capture or can capture only a single site of interest (viewpoint) [58][59][60][61][62] , which limits their capacity for detailed analysis of CTCF peak clustering at selected TAD boundaries. To target multiple viewpoints within a single experiment, we therefore developed Nano-C (Fig 2a,b).…”

Sequencing and Chromosome-Scale Assembly of Plant Genomes, Brassica rapa as a Use Case

“…To determine from which part of the genome the Nanopore reads originates, an in silico digestion of the reference genome is performed, and each segment of the read is assigned to a subsection of the reference. Although relatively young [ 20 ], the Pore-C technology seems to be interesting and competitive with traditional Hi-C for the scaffolding of complex genomes [ 21 ].…”

Deciphering Multi-way Interactions in the Human Genome

“…From our Pore-C experiments using adult human dermal fibroblasts, and additional publicly available Pore-C data from B lymphocytes, we constructed hypergraphs as multiple resolutions (read level, 100 kb, 1 Mb, and 25 Mb) [5]. We first analyzed individual chromosomes at 100 kb resolution, and decomposed the multi-way contacts into their pairwise counterparts to identify topologically associated domains (TADs) using established methods (Materials and Methods, [9,10,11]).…”

“…Recently, an extension of Hi-C enabled preservation and sequencing of long reads (e.g. Pore-C) [5], which can be used to unambiguously identify sets of contacts among multiple loci. These data will undoubtedly clarify higher order structures in the genome, but new frameworks are required for analysis and representation of the multidimensional data captured in this assay.…”