Completing bacterial genome assemblies with multiplex MinION sequencing

Illumina sequencing platforms have enabled widespread bacterial whole genome sequencing. While Illumina data is appropriate for many analyses, its short read length limits its ability to resolve genomic structure. This has major implications for tracking the spread of mobile genetic elements, includ...

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Veröffentlicht in:Microbial genomics Jg. 3; H. 10; S. e000132
Hauptverfasser: Wick, Ryan R., Judd, Louise M., Gorrie, Claire L., Holt, Kathryn E.
Format: Journal Article
Sprache:Englisch
Veröffentlicht: England Microbiology Society 01.10.2017
Schlagworte:
Gene Library
Genomics
Klebsiella pneumoniae - genetics
Methods Paper
Phylogeny
Whole Genome Sequencing
multiplex sequencing
oxford nanopore
long-read sequencing
Klebsiella pneumoniae
genome assembly
hybrid assembly
ISSN:2057-5858, 2057-5858
Online-Zugang:Volltext
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Zusammenfassung:Illumina sequencing platforms have enabled widespread bacterial whole genome sequencing. While Illumina data is appropriate for many analyses, its short read length limits its ability to resolve genomic structure. This has major implications for tracking the spread of mobile genetic elements, including those which carry antimicrobial resistance determinants. Fully resolving a bacterial genome requires long-read sequencing such as those generated by Oxford Nanopore Technologies (ONT) platforms. Here we describe our use of the ONT MinION to sequence 12 isolates of Klebsiella pneumoniae on a single flow cell. We assembled each genome using a combination of ONT reads and previously available Illumina reads, and little to no manual intervention was needed to achieve fully resolved assemblies using the Unicycler hybrid assembler. Assembling only ONT reads with Canu was less effective, resulting in fewer resolved genomes and higher error rates even following error correction with Nanopolish. We demonstrate that multiplexed ONT sequencing is a valuable tool for high-throughput bacterial genome finishing. Specifically, we advocate the use of Illumina sequencing as a first analysis step, followed by ONT reads as needed to resolve genomic structure.
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These authors contributed equally to this work.
All supporting data, code and protocols have been provided within the article or through supplementary data files. One supplementary table and four supplementary figures are available with the online Supplementary Material.
ISSN:2057-5858
2057-5858
DOI:10.1099/mgen.0.000132