1. Scalable Nanopore sequencing of human genomes provides a comprehensive view of haplotype-resolved variation and methylation
- Author
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Kolmogorov, Mikhail, Billingsley, Kimberley J, Mastoras, Mira, Meredith, Melissa, Monlong, Jean, Lorig-Roach, Ryan, Asri, Mobin, Alvarez Jerez, Pilar, Malik, Laksh, Dewan, Ramita, Reed, Xylena, Genner, Rylee M, Daida, Kensuke, Behera, Sairam, Shafin, Kishwar, Pesout, Trevor, Prabakaran, Jeshuwin, Carnevali, Paolo, Yang, Jianzhi, Rhie, Arang, Scholz, Sonja W, Traynor, Bryan J, Miga, Karen H, Jain, Miten, Timp, Winston, Phillippy, Adam M, Chaisson, Mark, Sedlazeck, Fritz J, Blauwendraat, Cornelis, and Paten, Benedict
- Subjects
Biological Sciences ,Bioinformatics and Computational Biology ,Genetics ,Neurosciences ,Human Genome ,Nanotechnology ,Bioengineering ,Neurodegenerative ,Brain Disorders ,Humans ,Sequence Analysis ,DNA ,Haplotypes ,Genome ,Human ,Nanopore Sequencing ,Methylation ,Pilot Projects ,High-Throughput Nucleotide Sequencing ,Technology ,Medical and Health Sciences ,Developmental Biology ,Biological sciences - Abstract
Long-read sequencing technologies substantially overcome the limitations of short-reads but have not been considered as a feasible replacement for population-scale projects, being a combination of too expensive, not scalable enough or too error-prone. Here we develop an efficient and scalable wet lab and computational protocol, Napu, for Oxford Nanopore Technologies long-read sequencing that seeks to address those limitations. We applied our protocol to cell lines and brain tissue samples as part of a pilot project for the National Institutes of Health Center for Alzheimer's and Related Dementias. Using a single PromethION flow cell, we can detect single nucleotide polymorphisms with F1-score comparable to Illumina short-read sequencing. Small indel calling remains difficult within homopolymers and tandem repeats, but achieves good concordance to Illumina indel calls elsewhere. Further, we can discover structural variants with F1-score on par with state-of-the-art de novo assembly methods. Our protocol phases small and structural variants at megabase scales and produces highly accurate, haplotype-specific methylation calls.
- Published
- 2023