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Ultralong Oxford Nanopore reads enable the development of a reference-grade perennial ryegrass genome assembly


Frei, Daniel; Veekman, Elisabeth; Grogg, Daniel; Stoffel-Studer, Ingrid; Morishima, Aki; Shimizu-Inatsugi, Rie; Yates, Steven; Shimizu, Kentaro K; Frey, Jürg E; Studer, Bruno; Copetti, Dario (2021). Ultralong Oxford Nanopore reads enable the development of a reference-grade perennial ryegrass genome assembly. Genome Biology and Evolution, 13(8):evab159.

Abstract

Despite the progress made in DNA sequencing over the last decade, reconstructing telomere-to-telomere genome assemblies of large and repeat-rich eukaryotic genomes is still difficult. More accurate basecalls or longer reads could address this issue, but no current sequencing platform can provide both simultaneously. Perennial ryegrass (Lolium perenne L.) is an example of an important species for which the lack of a reference genome assembly hindered a swift adoption of genomics-based methods into breeding programs. To fill this gap, we optimized the Oxford Nanopore Technologies sequencing protocol, obtaining sequencing reads with a N50 of 62 kb—an unprecedented value for a plant sample. The assembly of such reads produced a highly complete (2.3 of 2.7 Gb), correct (QV 45), and contiguous (contig N50 and N90 11.74 and 3.34 Mb, respectively) genome assembly. We show how read length was key in determining the assembly contiguity. Sequence annotation revealed the dominance of transposable elements and repeated sequences (81.6% of the assembly) and identified 38,868 protein coding genes. Almost 90% of the bases could be anchored to seven pseudomolecules, providing the first high quality haploid reference assembly for perennial ryegrass. This protocol will enable producing longer Oxford Nanopore Technology reads for more plant samples and ushering forage grasses into modern genomics-assisted breeding programs.

Abstract

Despite the progress made in DNA sequencing over the last decade, reconstructing telomere-to-telomere genome assemblies of large and repeat-rich eukaryotic genomes is still difficult. More accurate basecalls or longer reads could address this issue, but no current sequencing platform can provide both simultaneously. Perennial ryegrass (Lolium perenne L.) is an example of an important species for which the lack of a reference genome assembly hindered a swift adoption of genomics-based methods into breeding programs. To fill this gap, we optimized the Oxford Nanopore Technologies sequencing protocol, obtaining sequencing reads with a N50 of 62 kb—an unprecedented value for a plant sample. The assembly of such reads produced a highly complete (2.3 of 2.7 Gb), correct (QV 45), and contiguous (contig N50 and N90 11.74 and 3.34 Mb, respectively) genome assembly. We show how read length was key in determining the assembly contiguity. Sequence annotation revealed the dominance of transposable elements and repeated sequences (81.6% of the assembly) and identified 38,868 protein coding genes. Almost 90% of the bases could be anchored to seven pseudomolecules, providing the first high quality haploid reference assembly for perennial ryegrass. This protocol will enable producing longer Oxford Nanopore Technology reads for more plant samples and ushering forage grasses into modern genomics-assisted breeding programs.

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Item Type:Journal Article, refereed, original work
Communities & Collections:07 Faculty of Science > Institute of Evolutionary Biology and Environmental Studies
08 Research Priority Programs > Evolution in Action: From Genomes to Ecosystems
Dewey Decimal Classification:570 Life sciences; biology
590 Animals (Zoology)
Scopus Subject Areas:Life Sciences > Ecology, Evolution, Behavior and Systematics
Life Sciences > Genetics
Uncontrolled Keywords:Genetics, Ecology, Evolution, Behavior and Systematics
Language:English
Date:3 August 2021
Deposited On:14 Jul 2021 15:28
Last Modified:25 Feb 2024 02:40
Publisher:Oxford University Press
ISSN:1759-6653
OA Status:Gold
Free access at:Publisher DOI. An embargo period may apply.
Publisher DOI:https://doi.org/10.1093/gbe/evab159
Project Information:
  • : FunderJST CREST
  • : Grant IDJPMJCR16O3
  • : Project Title
  • : FunderSNF
  • : Grant ID31003A_182318
  • : Project Title
  • : FunderURPP Evolution in Action
  • : Grant ID
  • : Project Title
  • : Project Websitehttps://www.evolution.uzh.ch/en.html
  • Content: Accepted Version
  • Language: English
  • Content: Published Version
  • Language: English
  • Licence: Creative Commons: Attribution 4.0 International (CC BY 4.0)