Oxford Nanopore sequencing, hybrid error correction, and de novo assembly of a eukaryotic genome

By Sara Goodwin, James Gurtowski, Scott Ethe-Sayers, Panchajanya Deshpande, Michael C. Schatz, W. Richard McCombie. bioRxiv, 2015

366 citations recorded by the source corpus.

Monitoring the progress of DNA molecules through a membrane pore has been postulated as a method for sequencing DNA for several decades. Recently, a nanopore-based sequencing instrument, the Oxford Nanopore MinION, has become available that we used for sequencing the S. cerevisiae genome. To make use of these data, we developed a novel open-source hybrid error correction algorithm Nanocorr (https://github.com/jgurtowski/nanocorr) specifically for Oxford Nanopore reads, as existing packages were incapable of assembling the long read lengths (5-50kbp) at such high error rate (between ∼5 and 40% error). With this new method we were able to perform a hybrid error correction of the nanopore reads using complementary MiSeq data and produce a de novo assembly that is highly contiguous and accurate: the contig N50 length is more than ten-times greater than an Illumina-only assembly (678kb versus 59.9kbp), and has greater than 99.88% consensus identity when compared to the reference. Furthermore, the assembly with the long nanopore reads presents a much more complete representation of the features of the genome and correctly assembles gene cassettes, rRNAs, transposable elements, and other genomic features that were almost entirely absent in the Illumina-only assembly. Reviewer link to data http://schatzlab.cshl.edu/data/nanocorr/

Subfields: Biomedical Engineering, Molecular Biology

View Paper: https://www.semanticscholar.org/paper/e18b489f02530ff144db28cb44537173e4296616

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Citations in the corpus

Contiguous and accurate de novo assembly of metazoan genomes with modest long read coverage (2015, neutral): “Long read sequencing technologies, such as those offered by PacBio, Oxford Nanopore (34) and Illumina TrueSeq (35) promise to improve the quality of de novo genome assemblies substantially.”
Mapping and phasing of structural variation in patient genomes using nanopore sequencing (2017, positive): “The MinION can sequence stretches of DNA of up to hundreds of kilobases in length, which already resulted in the sequencing of the genomes of several organisms 9,10 .”
HapCUT2: robust and accurate haplotype assembly for diverse sequencing technologies (2017, neutral): “ApartfromPacBio,OxfordNanoporesequencersarealsocapableof producinglongreads,albeitwithlowerthroughputthanPacBiose-quencers (Goodwin et al. 2015).”
Nanopore sequencing technology, bioinformatics and applications (2021, methodology): “Overall, method development for base calling went through four stages 32,44,58,67,68 : (1) base calling from the segmented current data by HMM at the early stage and by recurrent neural network in late 2016, (2) base calling from raw current data in 2017, (3) using a flip–flop model for identifying ”
Piercing the dark matter: bioinformatics of long-range sequencing and mapping (2018, methodology): “Hybrid error correction methods, such as PBcR 32 , Nanocorr 33 , Spades 34 and MaSuRCA 35 , use short-read data to error correct the long reads before assembly and are especially effective when a limited amount of long-read coverage is available (typically below ~30× coverage) 32,33 .”
From squiggle to basepair: computational approaches for improving nanopore sequencing read accuracy (2018, methodology): “When the MinION was first introduced, reads showed an accuracy of less than 60% [9, 10].”
Chromosome-scale shotgun assembly using an in vitro method for long-range linkage (2015, background): “Commercially available long-reads from Oxford Nanopore are promising but have even higher error rates and lower throughput (Goodwin et al. 2015).”