Abstract
The recent development of metagenomic assembly has revolutionized metagenomic data analysis, thanks to the improvement of sequencing techniques, more powerful computational infrastructure and the development of novel algorithms and methods. Using longer assembled contigs rather than raw reads improves the process of metagenomic binning and annotation significantly, ultimately resulting in a deeper understanding of the microbial dynamics of the metagenomic samples being analyzed. In this chapter, we demonstrate a typical metagenomic analysis pipeline including raw read quality evaluation and trimming, assembly and contig binning. Alternative tools that can be used for each step are also discussed.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
References
Pell J, Hintze A, Canino-Koning R et al (2012) Scaling metagenome sequence assembly with probabilistic de Bruijn graphs. Proc Natl Acad Sci U S A 109:13272–13277. https://doi.org/10.1073/pnas.1121464109
Sangwan N, Xia F, Gilbert JA (2016) Recovering complete and draft population genomes from metagenome datasets. Microbiome 4:8. https://doi.org/10.1186/s40168-016-0154-5
Kang DD, Froula J, Egan R, Wang Z (2014) MetaBAT: Metagenome binning based on abundance and tetranucleotide frequency. No. LBNL-7106E. Ernest Orlando Lawrence Berkeley National Laboratory, Berkeley, CA
Alneberg J, Bjarnason BS, de Bruijn I, et al (2014) Binning metagenomic contigs by coverage and composition. Nat Methods 11:1144–1146. doi: https://doi.org/10.1038/nmeth.3103
Sieber CMK, Probst AJ, Sharrar A et al (2017) Recovery of genomes from metagenomes via a dereplication, aggregation, and scoring strategy. bioRxiv:107789
Vollmers J, Wiegand S, Kaster AK (2017) Comparing and evaluating metagenome assembly tools from a microbiologist’s perspective—not only size matters! PLoS One 12:e0169662
Sczyrba A, Hofmann P, Belmann P et al (2017) Critical Assessment of Metagenome Interpretation—a benchmark of computational metagenomics software. bioRxiv:99127. https://doi.org/10.1101/099127
Awad S, Irber L, Brown CT (2017) Evaluating metagenome assembly on a simple defined community with many strain variants. bioRxiv:155358
Bolger AM, Lohse M, Usadel B (2014) Trimmomatic: a flexible trimmer for Illumina sequence data. Bioinformatics 30:2114–2120. https://doi.org/10.1093/bioinformatics/btu170
Li D, Liu C-M, Luo R et al (2015) MEGAHIT: an ultra-fast single-node solution for large and complex metagenomics assembly via succinct de Bruijn graph. Bioinformatics 31:1674–1676. https://doi.org/10.1093/bioinformatics/btv033
Li R, Zhu H, Ruan J et al (2010) De novo assembly of human genomes with massively parallel short read sequencing. Genome Res 20:265–272. https://doi.org/10.1101/gr.097261.109
Luo R, Liu B, Xie Y et al (2012) SOAPdenovo2: an empirically improved memory-efficient short-read de novo assembler. Gigascience 1:18. https://doi.org/10.1186/2047-217X-1-18
Mikheenko A, Saveliev V, Gurevich A (2016) MetaQUAST: evaluation of metagenome assemblies. Bioinformatics 32:1088–1090. https://doi.org/10.1093/bioinformatics/btv697
Kang DD, Froula J, Egan R, Wang Z (2015) MetaBAT, an efficient tool for accurately reconstructing single genomes from complex microbial communities. PeerJ 3:e1165. https://doi.org/10.7717/peerj.1165
Langmead B, Salzberg SL (2012) Fast gapped-read alignment with Bowtie 2. Nat Methods 9:357–359. https://doi.org/10.1038/nmeth.1923
Li H, Handsaker B, Wysoker A et al (2009) The sequence alignment/map format and SAMtools. Bioinformatics 25:2078–2079. https://doi.org/10.1093/bioinformatics/btp352
Parks DH, Imelfort M, Skennerton CT et al (2015) CheckM: assessing the quality of microbial genomes recovered from isolates, single cells, and metagenomes. Genome Res 25:1043–1055. https://doi.org/10.1101/GR.186072.114 gr.186072.114
Martin M (2011) Cutadapt removes adapter sequences from high-throughput sequencing reads. EMBnetJ 17:10. https://doi.org/10.14806/ej.17.1.200
Zhang Q, Awad S, Brown CT (2015) Crossing the streams: a framework for streaming analysis of short DNA sequencing reads. PeerJ Preprints. https://doi.org/10.7287/peerj.preprints.890v1
Crusoe MR, Alameldin HF, Awad S et al (2015) The khmer software package: enabling efficient nucleotide sequence analysis. F1000Res 4:900. https://doi.org/10.12688/f1000research.6924.1
Zhang Q, Pell J, Canino-Koning R et al (2014) These are not the k-mers you are looking for: efficient online k-mer counting using a probabilistic data structure. PLoS One 9:e101271. https://doi.org/10.1371/journal.pone.0101271
Ewels P, Magnusson M, Lundin S, Käller M (2016) MultiQC: summarize analysis results for multiple tools and samples in a single report. Bioinformatics 32:3047–3048. https://doi.org/10.1093/bioinformatics/btw354
Zerbino DR, Birney E (2008) Velvet: algorithms for de novo short read assembly using de Bruijn graphs. Genome Res 18:821–829. https://doi.org/10.1101/gr.074492.107
Bankevich A, Nurk S, Antipov D et al (2012) SPAdes: a new genome assembly algorithm and its applications to single-cell sequencing. J Comput Biol 19:455–477. https://doi.org/10.1089/cmb.2012.0021
Namiki T, Hachiya T, Tanaka H, Sakakibara Y (2012) MetaVelvet: an extension of Velvet assembler to de novo metagenome assembly from short sequence reads. Nucleic Acids Res 40:e155–e155. https://doi.org/10.1093/nar/gks678
Nurk S, Meleshko D, Korobeynikov A, Pevzner PA (2017) metaSPAdes: a new versatile metagenomic assembler. Genome Res 27:824–834. https://doi.org/10.1101/gr.213959.116
Peng Y, Leung HCM, Yiu SM, Chin FYL (2012) IDBA-UD: a de novo assembler for single-cell and metagenomic sequencing data with highly uneven depth. Bioinformatics 28:1420–1428. https://doi.org/10.1093/bioinformatics/bts174
Boisvert S, Raymond F, Godzaridis É et al (2012) Ray Meta: scalable de novo metagenome assembly and profiling. Genome Biol 13:R122. https://doi.org/10.1186/gb-2012-13-12-r122
Brown CT, Howe A, Zhang Q et al (2012) A reference-free algorithm for computational normalization of shotgun sequencing data. arXiv preprint arXiv 1203:4802
Howe AC, Jansson JK, Malfatti SA et al (2014) Tackling soil diversity with the assembly of large, complex metagenomes. Proc Natl Acad Sci U S A 111:4904–4909. https://doi.org/10.1073/pnas.1402564111
Wood DE, Salzberg SL (2014) Kraken: ultrafast metagenomic sequence classification using exact alignments. Genome Biol 15:R46. https://doi.org/10.1186/gb-2014-15-3-r46
Gregor I, Dröge J, Schirmer M et al (2016) PhyloPythiaS+: a self-training method for the rapid reconstruction of low-ranking taxonomic bins from metagenomes. PeerJ 4:e1603. https://doi.org/10.7717/peerj.1603
Dröge J, Gregor I, McHardy AC (2015) Taxator-tk: precise taxonomic assignment of metagenomes by fast approximation of evolutionary neighborhoods. Bioinformatics 31:817–824. https://doi.org/10.1093/bioinformatics/btu745
Huson DH, Auch AF, Qi J, Schuster SC (2007) MEGAN analysis of metagenomic data. Genome Res 17:377–386. https://doi.org/10.1101/gr.5969107
Markowitz VM, Chen IMA, Chu K et al (2013) IMG/M 4 version of the integrated metagenome comparative analysis system. Nucleic Acids Res 42(D1):D568–D573
Wilke A, Bischof J, Gerlach W et al (2015) The MG-RAST metagenomics database and portal in 2015. Nucleic Acids Res. https://doi.org/10.1093/nar/gkv1322
Wu Y, Simmons BA, Singer SW (2015) MaxBin 2.0: an automated binning algorithm to recover genomes from multiple metagenomic datasets. Bioinformatics:1–2. https://doi.org/10.1093/bioinformatics/btv638
Imelfort M, Parks D, Woodcroft BJ et al (2014) GroopM: an automated tool for the recovery of population genomes from related metagenomes. PeerJ 2:e603. https://doi.org/10.7717/peerj.603
Lin H-H, Liao Y-C (2016) Accurate binning of metagenomic contigs via automated clustering sequences using information of genomic signatures and marker genes. Sci Rep. https://doi.org/10.1038/srep24175
Li H, Durbin R (2009) Fast and accurate short read alignment with Burrows-Wheeler transform. Bioinformatics 25:1754–1760. https://doi.org/10.1093/bioinformatics/btp324
Li H (2013) Aligning sequence reads, clone sequences and assembly contigs with BWA-MEM arXiv Preprint arXiv:1303.3997
Author information
Authors and Affiliations
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2018 Springer Science+Business Media, LLC, part of Springer Nature
About this protocol
Cite this protocol
Zhang, Q. (2018). Metagenome Assembly and Contig Assignment. In: Beiko, R., Hsiao, W., Parkinson, J. (eds) Microbiome Analysis. Methods in Molecular Biology, vol 1849. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-8728-3_12
Download citation
DOI: https://doi.org/10.1007/978-1-4939-8728-3_12
Published:
Publisher Name: Humana Press, New York, NY
Print ISBN: 978-1-4939-8726-9
Online ISBN: 978-1-4939-8728-3
eBook Packages: Springer Protocols