Unlocking Short Read Sequencing for Metagenomics
Publication Date
July 28, 2010
Journal
PLOS ONE
Authors
Sébastien Rodrigue, Arne C. Materna, Sonia C. Timberlake, Matthew C. Blackburn, et al
Volume
5
Issue
7
Pages
e11840
DOI
https://dx.plos.org/10.1371/journal.pone.0011840
Publisher URL
http://journals.plos.org/plosone/article?id=10.1371%2Fjournal.pone.0011840
PubMed
http://www.ncbi.nlm.nih.gov/pubmed/20676378
PubMed Central
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2911387
Europe PMC
http://europepmc.org/abstract/MED/20676378
Web of Science
000280520100016
Scopus
77955603295
Mendeley
http://www.mendeley.com/research/unlocking-short-read-sequencing-metagenomics
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Scopus | Further Information

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  • {"files"=>["http://files.figshare.com/417657/Table_S1.pdf"], "pos_in_sequence"=>0, "stats"=>{"downloads"=>0, "page_views"=>10, "likes"=>0}, "users"=>["Matthew C. Blackburn", "Sébastien Rodrigue", "Rex R. Malmstrom", "Eric J. Alm", "Arne C. Materna", "Sallie W. Chisholm", "Sonia C. Timberlake"], "links"=>[], "tags"=>["sequencing", "metagenomics", "unlocking"], "title"=>"Unlocking Short Read Sequencing for Metagenomics", "figshare_url"=>"http://figshare.com/articles/Unlocking_Short_Read_Sequencing_for_Metagenomics/142457", "defined_type"=>3, "doi"=>["http://dx.doi.org/10.1371/journal.pone.0011840.s001"], "published_date"=>"2010-07-28 00:40:57", "article_id"=>142457, "categories"=>["Genetics", "Molecular Biology", "Biological Sciences", "Biochemistry", "Biotechnology"], "description"=>"<div><section><h3>Background</h3><p>Different high-throughput nucleic acid sequencing platforms are currently available but a trade-off currently exists between the cost and number of reads that can be generated versus the read length that can be achieved.</p></section><section><h3>Methodology/Principal Findings</h3><p>We describe an experimental and computational pipeline yielding millions of reads that can exceed 200 bp with quality scores approaching that of traditional Sanger sequencing. The method combines an automatable gel-less library construction step with paired-end sequencing on a short-read instrument. With appropriately sized library inserts, mate-pair sequences can overlap, and we describe the SHERA software package that joins them to form a longer composite read.</p></section><section><h3>Conclusions/Significance</h3><p>This strategy is broadly applicable to sequencing applications that benefit from low-cost high-throughput sequencing, but require longer read lengths. We demonstrate that our approach enables metagenomic analyses using the Illumina Genome Analyzer, with low error rates, and at a fraction of the cost of pyrosequencing.</p></section></div>"}
  • {"files"=>["http://files.figshare.com/838053/Figure_1.tif"], "pos_in_sequence"=>0, "stats"=>{"downloads"=>0, "page_views"=>7, "likes"=>0}, "users"=>["Matthew C. Blackburn", "Sébastien Rodrigue", "Rex R. Malmstrom", "Eric J. Alm", "Arne C. Materna", "Sallie W. Chisholm", "Sonia C. Timberlake"], "links"=>[], "tags"=>["dna", "fragments", "genomic", "sheared"], "title"=>"<p>Size-dependent isolation of DNA fragments from sheared genomic DNA via dSPRI.</p>", "figshare_url"=>"http://figshare.com/articles/_Size_dependent_isolation_of_DNA_fragments_from_sheared_genomic_DNA_via_dSPRI_/508414", "defined_type"=>1, "doi"=>["http://dx.doi.org/10.1371/journal.pone.0011840.g001"], "published_date"=>"2010-07-28 02:20:14", "article_id"=>508414, "categories"=>["Genetics", "Molecular Biology", "Biological Sciences", "Biochemistry", "Biotechnology"], "description"=>"<p>AMPure XP SPRI beads bind DNA fragments in a size dependent manner according to the concentration of salts and polyethylene glycol (PEG) in the reaction <a href=\"http://www.plosone.org/article/info:doi/10.1371/journal.pone.0011840#pone.0011840-Lennon1\" target=\"_blank\">[4]</a>–<a href=\"http://www.plosone.org/article/info:doi/10.1371/journal.pone.0011840#pone.0011840-Krizova1\" target=\"_blank\">[7]</a>, which can easily be changed by using different volume ratios of DNA to SPRI bead solutions. A two-step procedure is employed to isolate targeted DNA size fractions. Panels A to H present Bioanalyzer DNA-1000 assays showing the sheared genomic DNA used as starting material (black), the larger size DNA fragments discarded in separation 1 (red), and the size fraction purified and recovered after separation 2 (blue). Panel I is a table summarizing the conditions and results displayed in panels A to H. All Bioanalyzer DNA-1000 traces after separation 1 (panel J), and after separation 2 (panel K), are respectively displayed on a graph for the conditions presented in panels A to H. The conditions displayed in panel H were used to obtaine the Illumina composite reads discussed in the text. The wider DNA fragment size distribution from panel H allowed to better analyze the effects of shorter versus longer overlapping regions on consensus reads.</p>"}

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Relative Metric

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