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Analysis of Run-to-Run Variation of Bar-Coded Pyrosequencing for Evaluating Bacterial Community Shifts and Individual Taxa Dynamics

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  • Yuan Ge
  • Joshua P Schimel
  • Patricia A Holden

Abstract

Bar-coded pyrosequencing has been increasingly used due to its fine taxonomic resolution and high throughput. Yet, concerns arise regarding the reproducibility of bar-coded pyrosequencing. We evaluated the run-to-run variation of bar-coded pyrosequencing in detecting bacterial community shifts and taxa dynamics. Our results demonstrate that pyrosequencing is reproducible in evaluating community shifts within a run, but not between runs. Also, the reproducibility of pyrosequencing in detecting individual taxa increased as a function of taxa abundance. Based on our findings: (1) for studies with modest sequencing depth, it is doubtful that data from different pyrosequencing runs can be considered comparable; (2) if multiple pyrosequencing runs are needed to increase the sequencing depth, additional sequencing efforts should be applied to all samples, rather than to selected samples; (3) if pyrosequencing is used for estimating bacterial population dynamics, only the abundant taxa should be considered; (4) for less-abundant taxa, the sequencing depth should be increased to ensure an accurate evaluation of taxon variation trends across samples.

Suggested Citation

  • Yuan Ge & Joshua P Schimel & Patricia A Holden, 2014. "Analysis of Run-to-Run Variation of Bar-Coded Pyrosequencing for Evaluating Bacterial Community Shifts and Individual Taxa Dynamics," PLOS ONE, Public Library of Science, vol. 9(6), pages 1-6, June.
  • Handle: RePEc:plo:pone00:0099414
    DOI: 10.1371/journal.pone.0099414
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    References listed on IDEAS

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    1. James Robert White & Niranjan Nagarajan & Mihai Pop, 2009. "Statistical Methods for Detecting Differentially Abundant Features in Clinical Metagenomic Samples," PLOS Computational Biology, Public Library of Science, vol. 5(4), pages 1-11, April.
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