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      Evaluation of the impact of Illumina error correction tools on de novo genome assembly

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          Abstract

          Background

          Recently, many standalone applications have been proposed to correct sequencing errors in Illumina data. The key idea is that downstream analysis tools such as de novo genome assemblers benefit from a reduced error rate in the input data. Surprisingly, a systematic validation of this assumption using state-of-the-art assembly methods is lacking, even for recently published methods.

          Results

          For twelve recent Illumina error correction tools (EC tools) we evaluated both their ability to correct sequencing errors and their ability to improve de novo genome assembly in terms of contig size and accuracy.

          Conclusions

          We confirm that most EC tools reduce the number of errors in sequencing data without introducing many new errors. However, we found that many EC tools suffer from poor performance in certain sequence contexts such as regions with low coverage or regions that contain short repeated or low-complexity sequences. Reads overlapping such regions are often ill-corrected in an inconsistent manner, leading to breakpoints in the resulting assemblies that are not present in assemblies obtained from uncorrected data. Resolving this systematic flaw in future EC tools could greatly improve the applicability of such tools.

          Electronic supplementary material

          The online version of this article (doi:10.1186/s12859-017-1784-8) contains supplementary material, which is available to authorized users.

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          Most cited references25

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          ART: a next-generation sequencing read simulator.

          ART is a set of simulation tools that generate synthetic next-generation sequencing reads. This functionality is essential for testing and benchmarking tools for next-generation sequencing data analysis including read alignment, de novo assembly and genetic variation discovery. ART generates simulated sequencing reads by emulating the sequencing process with built-in, technology-specific read error models and base quality value profiles parameterized empirically in large sequencing datasets. We currently support all three major commercial next-generation sequencing platforms: Roche's 454, Illumina's Solexa and Applied Biosystems' SOLiD. ART also allows the flexibility to use customized read error model parameters and quality profiles. Both source and binary software packages are available at http://www.niehs.nih.gov/research/resources/software/art.
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            Evaluation of genomic high-throughput sequencing data generated on Illumina HiSeq and Genome Analyzer systems

            Background The generation and analysis of high-throughput sequencing data are becoming a major component of many studies in molecular biology and medical research. Illumina's Genome Analyzer (GA) and HiSeq instruments are currently the most widely used sequencing devices. Here, we comprehensively evaluate properties of genomic HiSeq and GAIIx data derived from two plant genomes and one virus, with read lengths of 95 to 150 bases. Results We provide quantifications and evidence for GC bias, error rates, error sequence context, effects of quality filtering, and the reliability of quality values. By combining different filtering criteria we reduced error rates 7-fold at the expense of discarding 12.5% of alignable bases. While overall error rates are low in HiSeq data we observed regions of accumulated wrong base calls. Only 3% of all error positions accounted for 24.7% of all substitution errors. Analyzing the forward and reverse strands separately revealed error rates of up to 18.7%. Insertions and deletions occurred at very low rates on average but increased to up to 2% in homopolymers. A positive correlation between read coverage and GC content was found depending on the GC content range. Conclusions The errors and biases we report have implications for the use and the interpretation of Illumina sequencing data. GAIIx and HiSeq data sets show slightly different error profiles. Quality filtering is essential to minimize downstream analysis artifacts. Supporting previous recommendations, the strand-specificity provides a criterion to distinguish sequencing errors from low abundance polymorphisms.
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              Aggressive assembly of pyrosequencing reads with mates

              Motivation: DNA sequence reads from Sanger and pyrosequencing platforms differ in cost, accuracy, typical coverage, average read length and the variety of available paired-end protocols. Both read types can complement one another in a ‘hybrid’ approach to whole-genome shotgun sequencing projects, but assembly software must be modified to accommodate their different characteristics. This is true even of pyrosequencing mated and unmated read combinations. Without special modifications, assemblers tuned for homogeneous sequence data may perform poorly on hybrid data. Results: Celera Assembler was modified for combinations of ABI 3730 and 454 FLX reads. The revised pipeline called CABOG (Celera Assembler with the Best Overlap Graph) is robust to homopolymer run length uncertainty, high read coverage and heterogeneous read lengths. In tests on four genomes, it generated the longest contigs among all assemblers tested. It exploited the mate constraints provided by paired-end reads from either platform to build larger contigs and scaffolds, which were validated by comparison to a finished reference sequence. A low rate of contig mis-assembly was detected in some CABOG assemblies, but this was reduced in the presence of sufficient mate pair data. Availability: The software is freely available as open-source from http://wgs-assembler.sf.net under the GNU Public License. Contact: jmiller@jcvi.org Supplementary information: Supplementary data are available at Bioinformatics online.
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                Author and article information

                Contributors
                mahdi.heydari@ugent.be
                giles.miclotte@ugent.be
                piet.demeester@ugent.be
                yves.vandepeer@psb.vib-ugent.be
                jan.fostier@ugent.be
                Journal
                BMC Bioinformatics
                BMC Bioinformatics
                BMC Bioinformatics
                BioMed Central (London )
                1471-2105
                18 August 2017
                18 August 2017
                2017
                : 18
                : 374
                Affiliations
                [1 ]ISNI 0000 0001 2069 7798, GRID grid.5342.0, Department of Information Technology, , Ghent University-imec, ; IDLab, Ghent, B-9052 Belgium
                [2 ]Center for Plant Systems Biology, VIB, Ghent, B-9052 Belgium
                [3 ]ISNI 0000 0001 2069 7798, GRID grid.5342.0, Department of Plant Biotechnology and Bioinformatics, , Ghent University, ; Ghent, B-9052 Belgium
                [4 ]Bioinformatics Institute Ghent, Ghent, B-9052 Belgium
                [5 ]ISNI 0000 0001 2107 2298, GRID grid.49697.35, Department of Genetics, Genome Research Institute, , University of Pretoria, ; Pretoria, South Africa
                Author information
                http://orcid.org/0000-0002-9994-8269
                Article
                1784
                10.1186/s12859-017-1784-8
                5563063
                28821237
                a865136a-76e2-468a-a207-98c141292311
                © The Author(s) 2017

                Open Access This article is 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 you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver( http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated.

                History
                : 21 March 2017
                : 11 August 2017
                Funding
                Funded by: FWO-Vlaanderen
                Award ID: G0C3914N
                Award Recipient :
                Categories
                Research Article
                Custom metadata
                © The Author(s) 2017

                Bioinformatics & Computational biology
                next-generation sequencing,error correction,illumina,genome assembly

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