Introduction

Bacterial diversity among environmental samples is commonly assessed with PCR-amplified 16S rRNA gene (16S) sequences. Perceived diversity, however, can be influenced by sample preparation, primer selection, and formation of chimeric 16S amplification products. Chimeras are hybrid products between multiple parent sequences that can be falsely interpreted as novel organisms, thus inflating apparent diversity. We developed a new chimera detection tool called Chimera Slayer (CS). CS detects chimeras with greater sensitivity than previous methods, performs well on short sequences such as those produced by the 454 Life Sciences (Roche) Genome Sequencer, and can scale to large data sets. By benchmarking CS performance against sequences derived from a controlled DNA mixture of known organisms and a simulated chimera set, we provide insights into the factors that affect chimera formation such as sequence abundance, the extent of similarity between 16S genes, and PCR conditions. Chimeras were found to reproducibly form among independent amplifications and contributed to false perceptions of sample diversity and the false identification of novel taxa, with less-abundant species exhibiting chimera rates exceeding 70%. Shotgun metagenomic sequences of our mock community appear to be devoid of 16S chimeras, supporting a role for shotgun metagenomics in validating novel organisms discovered in targeted sequence surveys.

Publications

  1. Chimeric 16S rRNA sequence formation and detection in Sanger and 454-pyrosequenced PCR amplicons.
    Cite this
    Haas BJ, Gevers D, Earl AM, Feldgarden M, Ward DV, Giannoukos G, Ciulla D, Tabbaa D, Highlander SK, Sodergren E, Methé B, DeSantis TZ, , Petrosino JF, Knight R, Birren BW, 2011-03-01 - Genome research

Credits

  1. Brian J Haas
    Developer

    Genome Sequencing and Analysis Program, The Broad Institute, United States of America

  2. Dirk Gevers
    Developer

  3. Ashlee M Earl
    Developer

  4. Mike Feldgarden
    Developer

  5. Doyle V Ward
    Developer

  6. Georgia Giannoukos
    Developer

  7. Dawn Ciulla
    Developer

  8. Diana Tabbaa
    Developer

  9. Sarah K Highlander
    Developer

  10. Erica Sodergren
    Developer

  11. Barbara Methé
    Developer

  12. Todd Z DeSantis
    Developer

  13. Joseph F Petrosino
    Developer

  14. Rob Knight
    Developer

  15. Bruce W Birren
    Investigator

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Summary
AccessionBT000768
Tool TypeApplication
Category
PlatformsLinux/Unix
TechnologiesC, Perl
User InterfaceTerminal Command Line
Download Count0
Submitted ByBruce W Birren