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      LongQC: A Quality Control Tool for Third Generation Sequencing Long Read Data

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          Abstract

          We propose LongQC as an easy and automated quality control tool for genomic datasets generated by third generation sequencing (TGS) technologies such as Oxford Nanopore technologies (ONT) and SMRT sequencing from Pacific Bioscience (PacBio). Key statistics were optimized for long read data, and LongQC covers all major TGS platforms. LongQC processes and visualizes those statistics automatically and quickly.

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          NanoSim: nanopore sequence read simulator based on statistical characterization

          Chen Yang, Justin Chu, René L. Warren … (2017)
          Abstract Background: The MinION sequencing instrument from Oxford Nanopore Technologies (ONT) produces long read lengths from single-molecule sequencing – valuable features for detailed genome characterization. To realize the potential of this platform, a number of groups are developing bioinformatics tools tuned for the unique characteristics of its data. We note that these development efforts would benefit from a simulator software, the output of which could be used to benchmark analysis tools. Results: Here, we introduce NanoSim, a fast and scalable read simulator that captures the technology-specific features of ONT data and allows for adjustments upon improvement of nanopore sequencing technology. The first step of NanoSim is read characterization, which provides a comprehensive alignment-based analysis and generates a set of read profiles serving as the input to the next step, the simulation stage. The simulation stage uses the model built in the previous step to produce in silico reads for a given reference genome. NanoSim is written in Python and R. The source files and manual are available at the Genome Sciences Centre website: http://www.bcgsc.ca/platform/bioinfo/software/nanosim. Conclusion: In this work, we model the base-calling errors of ONT reads to inform the simulation of sequences with similar characteristics. We showcase the performance of NanoSim on publicly available datasets generated using the R7 and R7.3 chemistries and different sequencing kits and compare the resulting synthetic reads to those of other long-sequence simulators and experimental ONT reads. We expect NanoSim to have an enabling role in the field and benefit the development of scalable next-generation sequencing technologies for the long nanopore reads, including genome assembly, mutation detection, and even metagenomic analysis software.
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            poRe: an R package for the visualization and analysis of nanopore sequencing data

            Mick Watson, Marian Thomson, Judith Risse … (2014)
            Motivation: The Oxford Nanopore MinION device represents a unique sequencing technology. As a mobile sequencing device powered by the USB port of a laptop, the MinION has huge potential applications. To enable these applications, the bioinformatics community will need to design and build a suite of tools specifically for MinION data. Results: Here we present poRe, a package for R that enables users to manipulate, organize, summarize and visualize MinION nanopore sequencing data. As a package for R, poRe has been tested on Windows, Linux and MacOSX. Crucially, the Windows version allows users to analyse MinION data on the Windows laptop attached to the device. Availability and implementation: poRe is released as a package for R at http://sourceforge.net/projects/rpore/. A tutorial and further information are available at https://sourceforge.net/p/rpore/wiki/Home/ Contact: mick.watson@roslin.ed.ac.uk Supplementary information: Supplementary data are available at Bioinformatics online.
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              CarrierSeq: a sequence analysis workflow for low-input nanopore sequencing

              Angel Mojarro, Julie Hachey, Gary Ruvkun … (2018)
              Background Long-read nanopore sequencing technology is of particular significance for taxonomic identification at or below the species level. For many environmental samples, the total extractable DNA is far below the current input requirements of nanopore sequencing, preventing “sample to sequence” metagenomics from low-biomass or recalcitrant samples. Results Here we address this problem by employing carrier sequencing, a method to sequence low-input DNA by preparing the target DNA with a genomic carrier to achieve ideal library preparation and sequencing stoichiometry without amplification. We then use CarrierSeq, a sequence analysis workflow to identify the low-input target reads from the genomic carrier. We tested CarrierSeq experimentally by sequencing from a combination of 0.2 ng Bacillus subtilis ATCC 6633 DNA in a background of 1000 ng Enterobacteria phage λ DNA. After filtering of carrier, low quality, and low complexity reads, we detected target reads (B. subtilis), contamination reads, and “high quality noise reads” (HQNRs) not mapping to the carrier, target or known lab contaminants. These reads appear to be artifacts of the nanopore sequencing process as they are associated with specific channels (pores). Conclusion By treating sequencing as a Poisson arrival process, we implement a statistical test to reject data from channels dominated by HQNRs while retaining low-input target reads.
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                Author and article information

                Journal
                Journal ID (nlm-ta): G3 (Bethesda)
                Journal ID (iso-abbrev): Genetics
                Journal ID (hwp): G3: Genes, Genomes, Genetics
                Journal ID (pmc): G3: Genes, Genomes, Genetics
                Journal ID (publisher-id): G3: Genes, Genomes, Genetics
                Title: G3: Genes|Genomes|Genetics
                Publisher: Genetics Society of America
                ISSN (Electronic): 2160-1836
                Publication date (Electronic): 10 February 2020
                Publication date Collection: April 2020
                Volume: 10
                Issue: 4
                Pages: 1193-1196
                Affiliations
                [1]King Abdullah University of Science and Technology (KAUST), Core Labs, Thuwal, Makkah, Saudi Arabia, 23955-6900
                Author notes
                [2]

                Present address: Oxford Nanopore Technologies, Shanghai, 200000, China

                Author information
                http://orcid.org/0000-0002-0882-4392
                http://orcid.org/0000-0001-8109-6021
                Article
                Publisher ID: GGG_400864
                DOI: 10.1534/g3.119.400864
                PMC ID: 7144081
                PubMed ID: 32041730
                SO-VID: 857eada4-2c12-4c20-90cc-41631e741148
                Copyright © Copyright © 2020 Fukasawa et al.
                License:

                This is an open-access article distributed under the terms of the Creative Commons Attribution 4.0 International License ( http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

                History
                Date received : 26 October 2019
                Date accepted : 01 February 2020
                Page count
                Figures: 2, Tables: 0, Equations: 0, References: 11, Pages: 4
                Categories
                Subject: Software and Data Resources

                ScienceOpen disciplines: Genetics
                Keywords: quality control,long read,third generation sequencers,pacbio,oxford nanopore
                Data availability:
                ScienceOpen disciplines: Genetics
                Keywords: quality control, long read, third generation sequencers, pacbio, oxford nanopore

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