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Author Notes:

E-mail: kostas@ce.gatech.edu

We would like to thank Chad Haase and Ryan Weil for their assistance with sequencing and Rachel Poretsky for critically reading the manuscript.

The sponsors of this research had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.

Subjects:

Research Funding:

This research was supported, in part, by the U.S. Department of Energy (award DE-SC0004601).

The Emory Genome Center acknowledges the Georgia Research Alliance and the Atlanta Clinical and Translational Sciences Institute for funding for major equipment purchases.

T acknowledges the support of the Onassis Scholarship Foundation.

No additional external funding was received for this study.

Keywords:

  • Science & Technology
  • Multidisciplinary Sciences
  • Science & Technology - Other Topics
  • MULTIDISCIPLINARY SCIENCES
  • GENOME
  • GENERATION
  • ALIGNMENT
  • CATALOG
  • BLAST

Direct Comparisons of Illumina vs. Roche 454 Sequencing Technologies on the Same Microbial Community DNA Sample

Tools:

Journal Title:

PLoS ONE

Volume:

Volume 7, Number 2

Publisher:

, Pages e30087-e30087

Type of Work:

Article | Final Publisher PDF

Abstract:

Next-generation sequencing (NGS) is commonly used in metagenomic studies of complex microbial communities but whether or not different NGS platforms recover the same diversity from a sample and their assembled sequences are of comparable quality remain unclear. We compared the two most frequently used platforms, the Roche 454 FLX Titanium and the Illumina Genome Analyzer (GA) II, on the same DNA sample obtained from a complex freshwater planktonic community. Despite the substantial differences in read length and sequencing protocols, the platforms provided a comparable view of the community sampled. For instance, derived assemblies overlapped in ~90% of their total sequences and in situ abundances of genes and genotypes (estimated based on sequence coverage) correlated highly between the two platforms (R 2 & 0.9). Evaluation of base-call error, frameshift frequency, and contig length suggested that Illumina offered equivalent, if not better, assemblies than Roche 454. The results from metagenomic samples were further validated against DNA samples of eighteen isolate genomes, which showed a range of genome sizes and G+C% content. We also provide quantitative estimates of the errors in gene and contig sequences assembled from datasets characterized by different levels of complexity and G+C% content. For instance, we noted that homopolymer-associated, single-base errors affected ~1% of the protein sequences recovered in Illumina contigs of 10× coverage and 50% G+C; this frequency increased to ~3% when non-homopolymer errors were also considered. Collectively, our results should serve as a useful practical guide for choosing proper sampling strategies and data possessing protocols for future metagenomic studies.

Copyright information:

© 2012 Luo et al.

This is an Open Access work distributed under the terms of the Creative Commons Attribution 4.0 International License (https://creativecommons.org/licenses/by/4.0/).
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