Oxford Nanopore Develops Pore-C, Couples Chromatin Conformation Capture With Long Read Tech

By Bio-IT World Staff

November 14, 2019 | Last week Weill Cornell Medicine and Oxford Nanopore published a pre-print describing Pore-C, a new nanopore sequencing technique which couples chromatin conformation capture with Oxford Nanopore Technologies (ONT) long reads to directly sequence multi-way chromatin contacts without amplification.

The 3D genome is known to impact gene regulation, and we’ve made strides in understanding some of those interactions with chromosome conformation capture assays that use molecular readouts (e.g. PCR, sequencing) to map pairs of loci that spatially interact in the nucleus as chromatin folds. But pairwise interactions are likely to be insufficient. As the genome folds, many DNA loci may be involved in some gene expression. “Detection of such higher-order chromatin complexes may be necessary to reveal fundamental links between genome structure and function,” the authors write.

Coupling chromatin conformation capture with nanopore long reads means multiple contacts can be spanned in a single read, giving multi-way, higher order information. The authors claim that Pore-C is 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.

Pore-C

Researchers worked with Human B lymphocytes (GM12878) and HG002 cells, both from the Coriell Institute. Purified DNA was sequenced on ONT’s MinION, GridION or PromethION platforms. They developed a reproducible analytic pipeline for deriving multi-way chromatin contacts from Pore-C concatemers, and a workflow which uses the snakemake framework. The workflow involves aligning Pore-C reads to a reference genome with Pore-C specific parameters. Alignments are filtered and annotated. When each alignment along a Pore-C read is assigned to a single restriction fragment, a multi-way contact is constituted, and this multi-way contact can then be decomposed into pairwise contacts. The pairwise contacts are assigned to genome bins using conventional Hi-C workflows to generate a contact map.

Pore-C is consistent with gold-standard pairwise contact maps at the compartment, topologically-associated domain, and loop levels, and more efficient than existing multi-way methods. The long-range information encoded in Pore-C reads can be used to scaffold and correct genome assemblies and aid the reconstruction of complex rearrangements spanning multiple megabases and chromosomes.

As well as providing a more in-depth interrogation of the 3D genome and its impact on regulation of gene expression, it is possible to access interactions among repeat regions, and across structural variants. By omitting PCR, the technique also minimizes sequencing bias, enabling further insights in those regions with high or low GC-content.

Finally, as each nanopore long read contains more interactions compared to a short read, fewer sequencing reads are needed to build a complete picture.

“Our results establish Pore-C as the most simple and scalable assay for the genome-wide assessment of combinatorial chromatin interactions. We look forward to seeing how users of nanopore devices utilize this technique to gain new insights across a broad range of research applications,” said Sissel Juul, Director, Genomic Applications, Oxford Nanopore, in a press release.