Rapid de novo assembly of the European eel genome from nanopore sequencing reads

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Abstract

We have sequenced the genome of the endangered European eel using the MinION by Oxford Nanopore, and assembled these data using a novel algorithm specifically designed for large eukaryotic genomes. For this 860 Mbp genome, the entire computational process takes two days on a single CPU. The resulting genome assembly significantly improves on a previous draft based on short reads only, both in terms of contiguity (N50 1.2 Mbp) and structural quality. This combination of affordable nanopore sequencing and light weight assembly promises to make high-quality genomic resources accessible for many non-model plants and animals.

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FLASH: fast length adjustment of short reads to improve genome assemblies.

T. Magoc, S. L. Salzberg (2013)

Next-generation sequencing technologies generate very large numbers of short reads. Even with very deep genome coverage, short read lengths cause problems in de novo assemblies. The use of paired-end libraries with a fragment size shorter than twice the read length provides an opportunity to generate much longer reads by overlapping and merging read pairs before assembling a genome. We present FLASH, a fast computational tool to extend the length of short reads by overlapping paired-end reads from fragment libraries that are sufficiently short. We tested the correctness of the tool on one million simulated read pairs, and we then applied it as a pre-processor for genome assemblies of Illumina reads from the bacterium Staphylococcus aureus and human chromosome 14. FLASH correctly extended and merged reads >99% of the time on simulated reads with an error rate of <1%. With adequately set parameters, FLASH correctly merged reads over 90% of the time even when the reads contained up to 5% errors. When FLASH was used to extend reads prior to assembly, the resulting assemblies had substantially greater N50 lengths for both contigs and scaffolds. The FLASH system is implemented in C and is freely available as open-source code at http://www.cbcb.umd.edu/software/flash. t.magoc@gmail.com.

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Comprehensive comparison of Pacific Biosciences and Oxford Nanopore Technologies and their applications to transcriptome analysis

Jason L Weirather, Mariateresa de Cesare, Yunhao Wang … (2017)

Background: Given the demonstrated utility of Third Generation Sequencing [Pacific Biosciences (PacBio) and Oxford Nanopore Technologies (ONT)] long reads in many studies, a comprehensive analysis and comparison of their data quality and applications is in high demand. Methods: Based on the transcriptome sequencing data from human embryonic stem cells, we analyzed multiple data features of PacBio and ONT, including error pattern, length, mappability and technical improvements over previous platforms. We also evaluated their application to transcriptome analyses, such as isoform identification and quantification and characterization of transcriptome complexity, by comparing the performance of size-selected PacBio, non-size-selected ONT and their corresponding Hybrid-Seq strategies (PacBio+Illumina and ONT+Illumina). Results: PacBio shows overall better data quality, while ONT provides a higher yield. As with data quality, PacBio performs marginally better than ONT in most aspects for both long reads only and Hybrid-Seq strategies in transcriptome analysis. In addition, Hybrid-Seq shows superior performance over long reads only in most transcriptome analyses. Conclusions: Both PacBio and ONT sequencing are suitable for full-length single-molecule transcriptome analysis. As this first use of ONT reads in a Hybrid-Seq analysis has shown, both PacBio and ONT can benefit from a combined Illumina strategy. The tools and analytical methods developed here provide a resource for future applications and evaluations of these rapidly-changing technologies.

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Reducing assembly complexity of microbial genomes with single-molecule sequencing

Sergey Koren, Gregory Harhay, Timothy Smith … (2013)

Background The short reads output by first- and second-generation DNA sequencing instruments cannot completely reconstruct microbial chromosomes. Therefore, most genomes have been left unfinished due to the significant resources required to manually close gaps in draft assemblies. Third-generation, single-molecule sequencing addresses this problem by greatly increasing sequencing read length, which simplifies the assembly problem. Results To measure the benefit of single-molecule sequencing on microbial genome assembly, we sequenced and assembled the genomes of six bacteria and analyzed the repeat complexity of 2,267 complete bacteria and archaea. Our results indicate that the majority of known bacterial and archaeal genomes can be assembled without gaps, at finished-grade quality, using a single PacBio RS sequencing library. These single-library assemblies are also more accurate than typical short-read assemblies and hybrid assemblies of short and long reads. Conclusions Automated assembly of long, single-molecule sequencing data reduces the cost of microbial finishing to $1,000 for most genomes, and future advances in this technology are expected to drive the cost lower. This is expected to increase the number of completed genomes, improve the quality of microbial genome databases, and enable high-fidelity, population-scale studies of pan-genomes and chromosomal organization.

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Author and article information

Contributors

Christiaan V. Henkel:

ORCID: http://orcid.org/0000-0001-9838-8215

c.v.henkel@biology.leidenuniv.nl

Journal

Journal ID (nlm-ta): Sci Rep

Journal ID (iso-abbrev): Sci Rep

Title: Scientific Reports

Publisher: Nature Publishing Group UK (London )

ISSN (Electronic): 2045-2322

Publication date (Electronic): 3 August 2017

Publication date PMC-release: 3 August 2017

Publication date Collection: 2017

Volume: 7

Electronic Location Identifier: 7213

Affiliations

[1 ]GRID grid.474238.e, , ZF-screens B.V., ; Leiden, The Netherlands

[2 ]ISNI 0000 0001 2312 1970, GRID grid.5132.5, Institute of Biology, , Leiden University, ; Leiden, The Netherlands

[3 ]ISNI 0000 0001 2112 9282, GRID grid.4444.0, Muséum National d’Histoire Naturelle, , Sorbonne Universités, Research Unit BOREA, Biology of Aquatic Organisms and Ecosystems, CNRS, IRD, UCN, UA, ; Paris, France

[4 ]Norwegian University of Life Sciences, Faculty of Veterinary Medicine, Department of Basic Science and Aquatic Medicine, Oslo, Norway

[5 ]DUPAN Foundation, Wageningen, The Netherlands

[6 ]ISNI 0000 0001 0791 5666, GRID grid.4818.5, Animal Breeding and Genomics Centre, , Wageningen Livestock Research, Wageningen University & Research, ; Wageningen, The Netherlands

[7 ]ISNI 0000 0001 2151 8122, GRID grid.5771.4, Institute of Zoology and Center for Molecular Biosciences, , University of Innsbruck, ; Innsbruck, Austria

[8 ]GRID grid.449761.9, , University of Applied Sciences Leiden, ; Leiden, The Netherlands

[9 ]Generade Centre of Expertise in Genomics, Leiden, The Netherlands

Author information

Finn-Arne Weltzien http://orcid.org/0000-0002-5111-1558

Christiaan V. Henkel http://orcid.org/0000-0001-9838-8215

Article

Publisher ID: 7650

DOI: 10.1038/s41598-017-07650-6

PMC ID: 5543108

PubMed ID: 28775309

SO-VID: d968b9c3-69e7-452f-b62f-fd73abdca106

License:

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

History

Date received : 16 March 2017

Date accepted : 28 June 2017

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Rapid de novo assembly of the European eel genome from nanopore sequencing reads

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Most cited references 33

FLASH: fast length adjustment of short reads to improve genome assemblies.

Comprehensive comparison of Pacific Biosciences and Oxford Nanopore Technologies and their applications to transcriptome analysis

Reducing assembly complexity of microbial genomes with single-molecule sequencing

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