MicroPIPE: validating an end-to-end workflow for high-quality complete bacterial genome construction

BMC Genomics

Valentine Murigneux¹  Minh-Duy Phan²  Nguyen Thi Khanh Nhu²  Mark A. Schembri²  Scott A. Beatson³  Brian M. Forde⁴  David L. Paterson⁴  Patrick N. A. Harris⁵  Adam D. Irwin⁶  David M. Whiley⁶  Leah W. Roberts⁷ 

[1] QCIF Facility for Advanced Bioinformatics, Institute for Molecular Bioscience, The University of Queensland, Brisbane, Queensland, Australia;School of Chemistry and Molecular Biosciences, The University of Queensland, Brisbane, Queensland, Australia;School of Chemistry and Molecular Biosciences, The University of Queensland, Brisbane, Queensland, Australia;Australian Centre for Ecogenomics, The University of Queensland, Brisbane, Queensland, Australia;University of Queensland Centre for Clinical Research, Brisbane, Queensland, Australia;University of Queensland Centre for Clinical Research, Brisbane, Queensland, Australia;Central Microbiology, Pathology Queensland, Royal Brisbane & Women’s Hospital, Brisbane, Queensland, Australia;University of Queensland Centre for Clinical Research, Brisbane, Queensland, Australia;Queensland Children’s Hospital, Brisbane, Queensland, Australia;University of Queensland Centre for Clinical Research, Brisbane, Queensland, Australia;Queensland Children’s Hospital, Brisbane, Queensland, Australia;European Bioinformatics Institute, European Molecular Biology Laboratory (EMBL), Hinxton, Cambridge, UK;

关键词: Nanopore;    ONT;    Pipeline;    Sequence;    Bacteria;    Assembly;    Polishing;   

DOI  :  10.1186/s12864-021-07767-z

来源: Springer

【 摘 要 】

BackgroundOxford Nanopore Technology (ONT) long-read sequencing has become a popular platform for microbial researchers due to the accessibility and affordability of its devices. However, easy and automated construction of high-quality bacterial genomes using nanopore reads remains challenging. Here we aimed to create a reproducible end-to-end bacterial genome assembly pipeline using ONT in combination with Illumina sequencing.ResultsWe evaluated the performance of several popular tools used during genome reconstruction, including base-calling, filtering, assembly, and polishing. We also assessed overall genome accuracy using ONT both natively and with Illumina. All steps were validated using the high-quality complete reference genome for the Escherichia coli sequence type (ST)131 strain EC958. Software chosen at each stage were incorporated into our final pipeline, MicroPIPE.Further validation of MicroPIPE was carried out using 11 additional ST131 E. coli isolates, which demonstrated that complete circularised chromosomes and plasmids could be achieved without manual intervention. Twelve publicly available Gram-negative and Gram-positive bacterial genomes (with available raw ONT data and matched complete genomes) were also assembled using MicroPIPE. We found that revised basecalling and updated assembly of the majority of these genomes resulted in improved accuracy compared to the current publicly available complete genomes.ConclusionsMicroPIPE is built in modules using Singularity container images and the bioinformatics workflow manager Nextflow, allowing changes and adjustments to be made in response to future tool development. Overall, MicroPIPE provides an easy-access, end-to-end solution for attaining high-quality bacterial genomes. MicroPIPE is available at https://github.com/BeatsonLab-MicrobialGenomics/micropipe.

【 授权许可】

CC BY