Change search
CiteExportLink to record
Permanent link

Direct link
Cite
Citation style
  • apa
  • ieee
  • modern-language-association-8th-edition
  • vancouver
  • Other style
More styles
Language
  • de-DE
  • en-GB
  • en-US
  • fi-FI
  • nn-NO
  • nn-NB
  • sv-SE
  • Other locale
More languages
Output format
  • html
  • text
  • asciidoc
  • rtf
Confident phylogenetic identification of uncultured prokaryotes through long read amplicon sequencing of the 16S-ITS-23S rRNA operon
Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular Evolution. Uppsala University, Science for Life Laboratory, SciLifeLab.ORCID iD: 0000-0001-7651-5252
Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular Evolution. Uppsala University, Science for Life Laboratory, SciLifeLab.
Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular Evolution. Uppsala University, Science for Life Laboratory, SciLifeLab.ORCID iD: 0000-0002-4453-4173
Uppsala University, Science for Life Laboratory, SciLifeLab. Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular Evolution.
Show others and affiliations
2019 (English)In: Environmental Microbiology, ISSN 1462-2912, E-ISSN 1462-2920, Vol. 21, no 7, p. 2485-2498Article in journal (Refereed) Published
Abstract [en]

Amplicon sequencing of the 16S rRNA gene is the predominant method to quantify microbial compositions and to discover novel lineages. However, traditional short amplicons often do not contain enough information to confidently resolve their phylogeny. Here we present a cost-effective protocol that amplifies a large part of the rRNA operon and sequences the amplicons with PacBio technology. We tested our method on a mock community and developed a read-curation pipeline that reduces the overall read error rate to 0.18%. Applying our method on four environmental samples, we captured near full-length rRNA operon amplicons from a large diversity of prokaryotes. The method operated at moderately high-throughput (22286-37,850 raw ccs reads) and generated a large amount of putative novel archaeal 23S rRNA gene sequences compared to the archaeal SILVA database. These long amplicons allowed for higher resolution during taxonomic classification by means of long (similar to 1000 bp) 16S rRNA gene fragments and for substantially more confident phylogenies by means of combined near full-length 16S and 23S rRNA gene sequences, compared to shorter traditional amplicons (250 bp of the 16S rRNA gene). We recommend our method to those who wish to cost-effectively and confidently estimate the phylogenetic diversity of prokaryotes in environmental samples at high throughput.

Place, publisher, year, edition, pages
John Wiley & Sons, 2019. Vol. 21, no 7, p. 2485-2498
National Category
Microbiology
Identifiers
URN: urn:nbn:se:uu:diva-390901DOI: 10.1111/1462-2920.14636ISI: 000474294900020PubMedID: 31012228OAI: oai:DiVA.org:uu-390901DiVA, id: diva2:1343923
Funder
EU, European Research Council, 310039-PUZZLE_CELLSwedish Foundation for Strategic Research , SSF-FFL5Swedish Research Council, 2015-04959Available from: 2019-08-19 Created: 2019-08-19 Last updated: 2019-08-19Bibliographically approved

Open Access in DiVA

fulltext(529 kB)39 downloads
File information
File name FULLTEXT01.pdfFile size 529 kBChecksum SHA-512
b8637964ab075e5bd8cd5c56e3fcf1e8586d6c006f49a2696b72b735506970b0d6bd8536aac070144ed60b8d54255bddc6956be6ad05b3471df6e6c3d25acfd7
Type fulltextMimetype application/pdf

Other links

Publisher's full textPubMed

Search in DiVA

By author/editor
Martijn, JoranLind, Anders E.Schön, Max E.Juzokaite, LinaBunikis, IgnasPettersson, Olga V.Ettema, Thijs J. G.
By organisation
Molecular EvolutionScience for Life Laboratory, SciLifeLab
In the same journal
Environmental Microbiology
Microbiology

Search outside of DiVA

GoogleGoogle Scholar
Total: 39 downloads
The number of downloads is the sum of all downloads of full texts. It may include eg previous versions that are now no longer available

doi
pubmed
urn-nbn

Altmetric score

doi
pubmed
urn-nbn
Total: 39 hits
CiteExportLink to record
Permanent link

Direct link
Cite
Citation style
  • apa
  • ieee
  • modern-language-association-8th-edition
  • vancouver
  • Other style
More styles
Language
  • de-DE
  • en-GB
  • en-US
  • fi-FI
  • nn-NO
  • nn-NB
  • sv-SE
  • Other locale
More languages
Output format
  • html
  • text
  • asciidoc
  • rtf