학술논문
Scalable Nanopore sequencing of human genomes provides a comprehensive view of haplotype-resolved variation and methylation
Document Type
Original Paper
Author
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; Paten, Benedict
Source
Nature Methods: Techniques for life scientists and chemists. 20(10):1483-1492
Subject
Language
English
ISSN
1548-7091
1548-7105
1548-7105
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.
This work introduces a wet lab and computational pipeline, Napu, for small variant calling and de novo assembly of Nanopore sequencing data, which leads to comparable performances to short-read sequencing and allows for large-scale long-read sequencing projects.
This work introduces a wet lab and computational pipeline, Napu, for small variant calling and de novo assembly of Nanopore sequencing data, which leads to comparable performances to short-read sequencing and allows for large-scale long-read sequencing projects.