Ultra-high-throughput microbial community analysis on the Illumina HiSeq and MiSeq platforms
J. Gregory Caporaso,Christian L. Lauber,William A. Walters,Donna Berg-Lyons,James Huntley,Noah Fierer,Noah Fierer,Sarah M. Owens,Jason Betley,Louise Fraser,Markus J. Bauer,Niall Anthony Gormley,Jack A. Gilbert,Jack A. Gilbert,Geoff Smith,Rob Knight +15 more
TLDR
It is shown that the protocol developed for these instruments successfully recaptures known biological results, and additionally that biological conclusions are consistent across sequencing platforms (the HiSeq2000 versus the MiSeq) and across the sequenced regions of amplicons.Abstract:
DNA sequencing continues to decrease in cost with the Illumina HiSeq2000 generating up to 600 Gb of paired-end 100 base reads in a ten-day run. Here we present a protocol for community amplicon sequencing on the HiSeq2000 and MiSeq Illumina platforms, and apply that protocol to sequence 24 microbial communities from host-associated and free-living environments. A critical question as more sequencing platforms become available is whether biological conclusions derived on one platform are consistent with what would be derived on a different platform. We show that the protocol developed for these instruments successfully recaptures known biological results, and additionally that biological conclusions are consistent across sequencing platforms (the HiSeq2000 versus the MiSeq) and across the sequenced regions of amplicons.read more
Citations
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DADA2: High-resolution sample inference from Illumina amplicon data
Benjamin J. Callahan,Paul J. McMurdie,Michael J. Rosen,Andrew W. Han,Amy Jo A. Johnson,Susan Holmes +5 more
TL;DR: The open-source software package DADA2 for modeling and correcting Illumina-sequenced amplicon errors is presented, revealing a diversity of previously undetected Lactobacillus crispatus variants.
Journal ArticleDOI
Diet rapidly and reproducibly alters the human gut microbiome
Lawrence A. David,Corinne F. Maurice,Rachel N. Carmody,David B. Gootenberg,Julie E. Button,Benjamin E. Wolfe,Alisha V. Ling,A. Sloan Devlin,Yug Varma,Michael A. Fischbach,Sudha B. Biddinger,Rachel J. Dutton,Peter J. Turnbaugh +12 more
TL;DR: Increases in the abundance and activity of Bilophila wadsworthia on the animal-based diet support a link between dietary fat, bile acids and the outgrowth of microorganisms capable of triggering inflammatory bowel disease.
Journal ArticleDOI
Development of a dual-index sequencing strategy and curation pipeline for analyzing amplicon sequence data on the MiSeq Illumina sequencing platform.
TL;DR: This work presents an improved method for sequencing variable regions within the 16S rRNA gene using Illumina's MiSeq platform, which is currently capable of producing paired 250-nucleotide reads and demonstrates that it can provide data that are at least as good as that generated by the 454 platform while providing considerably higher sequencing coverage for a fraction of the cost.
Journal ArticleDOI
Evaluation of general 16S ribosomal RNA gene PCR primers for classical and next-generation sequencing-based diversity studies
Anna Klindworth,Elmar Pruesse,Timmy Schweer,Jörg Peplies,Christian Quast,Matthias Horn,Frank Oliver Glöckner +6 more
TL;DR: The results of this study may be used as a guideline for selecting primer pairs with the best overall coverage and phylum spectrum for specific applications, therefore reducing the bias in PCR-based microbial diversity studies.
Journal ArticleDOI
Quality-filtering vastly improves diversity estimates from Illumina amplicon sequencing.
Nicholas A. Bokulich,Sathish Subramanian,Jeremiah J. Faith,Dirk Gevers,Jeffrey I. Gordon,Rob Knight,Rob Knight,David A. Mills,J. Gregory Caporaso,J. Gregory Caporaso +9 more
TL;DR: It is demonstrated that high-quality read length and abundance are the primary factors differentiating correct from erroneous reads produced by Illumina GAIIx, HiSeq and MiSeq instruments.
References
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