Oxford Nanopore R10.4 long-read sequencing enables the generation of near-finished bacterial genomes from pure cultures and metagenomes without short-read or reference polishing

Sereika, Mantas; Kirkegaard, Rasmus Hansen; Karst, Søren Michael; Michaelsen, Thomas Yssing; Sørensen, Emil A.; Wollenberg, Rasmus Dam; Albertsen, Mads

doi:10.1038/s41592-022-01539-7

Cited by 239 publications

(291 citation statements)

References 58 publications

(63 reference statements)

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“…In particular, even higher-quality metagenomic assemblies are possible if HiFi reads from the Pacific Biosciences Sequel IIe system are available 46 . Also, the recent announcement of higher-quality reads from ONT could help improve assembly further and reduce costs 47 . Even as the sequencing landscape is constantly changing, the results from our study suggest that highquality population-specific metagenomic references are already feasible with a modest-sized cohort and limited sequencing resources.…”

Section: Discussionmentioning

confidence: 99%

Genome-centric analysis of short and long read metagenomes reveals uncharacterized microbiome diversity in Southeast Asians

et al. 2022

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Despite extensive efforts to address it, the vastness of uncharacterized ‘dark matter’ microbial genetic diversity can impact short-read sequencing based metagenomic studies. Population-specific biases in genomic reference databases can further compound this problem. Leveraging advances in hybrid assembly (using short and long reads) and Hi-C technologies in a cross-sectional survey, we deeply characterized 109 gut microbiomes from three ethnicities in Singapore to comprehensively reconstruct 4497 medium and high-quality metagenome assembled genomes, 1708 of which were missing in short-read only analysis and with >28× N50 improvement. Species-level clustering identified 70 (>10% of total) novel gut species out of 685, improved reference genomes for 363 species (53% of total), and discovered 3413 strains unique to these populations. Among the top 10 most abundant gut bacteria in our study, one of the species and >80% of strains were unrepresented in existing databases. Annotation of biosynthetic gene clusters (BGCs) uncovered more than 27,000 BGCs with a large fraction (36–88%) unrepresented in current databases, and with several unique clusters predicted to produce bacteriocins that could significantly alter microbiome community structure. These results reveal significant uncharacterized gut microbial diversity in Southeast Asian populations and highlight the utility of hybrid metagenomic references for bioprospecting and disease-focused studies.

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Section: Discussionmentioning

confidence: 99%

Genome-centric analysis of short and long read metagenomes reveals uncharacterized microbiome diversity in Southeast Asians

et al. 2022

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“…Higher basecall accuracy. Nanopore single-strand ('simplex') DNA basecall accuracy is 99% 11 . We believe that raising nanopore DRS accuracy to that level would revolutionize the field.…”

Section: Further Technical Improvements To Broaden Nanopore Drs Usementioning

confidence: 99%

Advances in nanopore direct RNA sequencing

et al. 2022

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“…Both E. coli and human genomes are sequenced using Oxford Nanopore Technologies (ONT) with R9-based chemistry [47]. This chemistry provides sequencing data with around 80-85% sequencing ac- 2 The E. coli dataset is available at: http://lab.loman.net/2016/07/30/ nanopore-r9-data-release/ curacy [142], which is slightly lower than the most recent chemistry (R10.4) that provides around 95-99% accuracy [143]. We include these less accurate datasets in our experiments to show the robustness and effectiveness of GenPIP in the presence of considerable sequencing inaccuracy.…”

Section: Evaluation Methodologymentioning

confidence: 99%

GenPIP: In-Memory Acceleration of Genome Analysis via Tight Integration of Basecalling and Read Mapping

Mao¹,

Alser²,

Sadrosadati³

et al. 2022

Preprint

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Nanopore sequencing is a widely-used high-throughput genome sequencing technology that can sequence long fragments of a genome into raw electrical signals at low cost. Nanopore sequencing requires two computationally-costly processing steps for accurate downstream genome analysis. The first step, basecalling, translates the raw electrical signals into nucleotide bases (i.e., A, C, G, T). The second step, read mapping, finds the correct location of a read in a reference genome. In existing genome analysis pipelines, basecalling and read mapping are executed separately. We observe in this work that such separate execution of the two most time-consuming steps inherently leads to (1) significant data movement and (2) redundant computations on the data, slowing down the genome analysis pipeline.This paper proposes GenPIP, an in-memory genome analysis accelerator that tightly integrates basecalling and read mapping. GenPIP improves the performance of the genome analysis pipeline with two key mechanisms: (1) in-memory fine-grained collaborative execution of the major genome analysis steps in parallel; (2) a new technique for early-rejection of low-quality and unmapped reads to timely stop the execution of genome analysis for such reads, reducing inefficient computation. Our experiments show that, for the execution of the genome analysis pipeline, GenPIP provides 41.6× (8.4×) speedup and 32.8× (20.8×) energy savings with negligible accuracy loss compared to the state-of-the-art software genome analysis tools executed on a state-of-the-art CPU (GPU). Compared to a design that combines state-of-the-art in-memory basecalling and read mapping accelerators, GenPIP provides 1.39× speedup and 1.37× energy savings.

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Oxford Nanopore R10.4 long-read sequencing enables the generation of near-finished bacterial genomes from pure cultures and metagenomes without short-read or reference polishing

Cited by 239 publications

References 58 publications

Genome-centric analysis of short and long read metagenomes reveals uncharacterized microbiome diversity in Southeast Asians

Genome-centric analysis of short and long read metagenomes reveals uncharacterized microbiome diversity in Southeast Asians

Advances in nanopore direct RNA sequencing

GenPIP: In-Memory Acceleration of Genome Analysis via Tight Integration of Basecalling and Read Mapping

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