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dc.contributor.authorVos, M
dc.contributor.authorQuince, C
dc.contributor.authorPijl, AS
dc.contributor.authorde Hollander, M
dc.contributor.authorKowalchuk, GA
dc.date.accessioned2017-02-21T10:45:52Z
dc.date.issued2012
dc.description.abstractBACKGROUND: The 16S rRNA gene is the gold standard in molecular surveys of bacterial and archaeal diversity, but it has the disadvantages that it is often multiple-copy, has little resolution below the species level and cannot be readily interpreted in an evolutionary framework. We compared the 16S rRNA marker with the single-copy, protein-coding rpoB marker by amplifying and sequencing both from a single soil sample. Because the higher genetic resolution of the rpoB gene prohibits its use as a universal marker, we employed consensus-degenerate primers targeting the Proteobacteria. METHODOLOGY/PRINCIPAL FINDINGS: Pyrosequencing can be problematic because of the poor resolution of homopolymer runs. As these erroneous runs disrupt the reading frame of protein-coding sequences, removal of sequences containing nonsense mutations was found to be a valuable filter in addition to flowgram-based denoising. Although both markers gave similar estimates of total diversity, the rpoB marker revealed more species, requiring an order of magnitude fewer reads to obtain 90% of the true diversity. The application of population genetic methods was demonstrated on a particularly abundant sequence cluster. CONCLUSIONS/SIGNIFICANCE: The rpoB marker can be a complement to the 16S rRNA marker for high throughput microbial diversity studies focusing on specific taxonomic groups. Additional error filtering is possible and tests for recombination or selection can be employed.en_GB
dc.description.sponsorshipThis work was funded by a Netherlands Organisation for Scientific Research (NWO) Vici grant to Dr. Kowalchuk (http://www.nwo.nl/nwohome.nsf/ pages/sppd_5r2qe7_eng). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.en_GB
dc.identifier.citationVol. 7, e30600en_GB
dc.identifier.doi10.1371/journal.pone.0030600
dc.identifier.otherPONE-D-11-18381
dc.identifier.urihttp://hdl.handle.net/10871/25982
dc.language.isoenen_GB
dc.publisherPublic Library of Scienceen_GB
dc.relation.urlhttps://www.ncbi.nlm.nih.gov/pubmed/22355318en_GB
dc.rights© 2012 Vos et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.en_GB
dc.subjectDNA, Bacterialen_GB
dc.subjectGenetic Markersen_GB
dc.subjectGenetic Variationen_GB
dc.subjectPhylogenyen_GB
dc.subjectProteobacteriaen_GB
dc.subjectRNA Polymerase IIen_GB
dc.subjectRNA, Ribosomal, 16Sen_GB
dc.subjectSequence Analysis, DNAen_GB
dc.titleA comparison of rpoB and 16S rRNA as markers in pyrosequencing studies of bacterial diversityen_GB
dc.typeArticleen_GB
dc.date.available2017-02-21T10:45:52Z
dc.identifier.issn1932-6203
exeter.place-of-publicationUnited Statesen_GB
dc.descriptionThis is the final version of the article. Available from the publisher via the DOI in this record.en_GB
dc.identifier.journalPLoS Oneen_GB
dc.identifier.pmcidPMC3280256
dc.identifier.pmid22355318


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