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Gigascience. 2019 May 1;8(5). pii: giz043. doi: 10.1093/gigascience/giz043.

Ultra-deep, long-read nanopore sequencing of mock microbial community standards.

Author information

1
Institute of Microbiology and Infection, School of Biosciences, University of Birmingham, Edgbaston, B15 2TT, UK.
2
Zymo Research Corporation, 17062 Murphy Ave., Irvine, CA 92614, USA.

Abstract

BACKGROUND:

Long sequencing reads are information-rich: aiding de novo assembly and reference mapping, and consequently have great potential for the study of microbial communities. However, the best approaches for analysis of long-read metagenomic data are unknown. Additionally, rigorous evaluation of bioinformatics tools is hindered by a lack of long-read data from validated samples with known composition.

FINDINGS:

We sequenced 2 commercially available mock communities containing 10 microbial species (ZymoBIOMICS Microbial Community Standards) with Oxford Nanopore GridION and PromethION. Both communities and the 10 individual species isolates were also sequenced with Illumina technology. We generated 14 and 16 gigabase pairs from 2 GridION flowcells and 150 and 153 gigabase pairs from 2 PromethION flowcells for the evenly distributed and log-distributed communities, respectively. Read length N50 ranged between 5.3 and 5.4 kilobase pairs over the 4 sequencing runs. Basecalls and corresponding signal data are made available (4.2 TB in total). Alignment to Illumina-sequenced isolates demonstrated the expected microbial species at anticipated abundances, with the limit of detection for the lowest abundance species below 50 cells (GridION). De novo assembly of metagenomes recovered long contiguous sequences without the need for pre-processing techniques such as binning.

CONCLUSIONS:

We present ultra-deep, long-read nanopore datasets from a well-defined mock community. These datasets will be useful for those developing bioinformatics methods for long-read metagenomics and for the validation and comparison of current laboratory and software pipelines.

KEYWORDS:

de novo assembly; Illumina; benchmark; bioinformatics; metagenomics; mock community; nanopore; real-time sequencing; single-molecule sequencing

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