Retrieval of a million high-quality, full-length microbial 16S and 18S rRNA gene sequences without primer bias

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24 Citationer (Scopus)

Resumé

Small subunit ribosomal RNA (SSU rRNA) genes, 16S in bacteria and 18S in eukaryotes, have been the standard phylogenetic markers used to characterize microbial diversity and evolution for decades. However, the reference databases of full-length SSU rRNA gene sequences are skewed to well-studied ecosystems and subject to primer bias and chimerism, which results in an incomplete view of the diversity present in a sample. We combine poly(A)-tailing and reverse transcription of SSU rRNA molecules with synthetic long-read sequencing to generate high-quality, full-length SSU rRNA sequences, without primer bias, at high throughput. We apply our approach to samples from seven different ecosystems and obtain more than a million SSU rRNA sequences from all domains of life, with an estimated raw error rate of 0.17%. We observe a large proportion of novel diversity, including several deeply branching phylum-level lineages putatively related to the Asgard Archaea. Our approach will enable expansion of the SSU rRNA reference databases by orders of magnitude, and contribute to a comprehensive census of the tree of life.
OriginalsprogEngelsk
TidsskriftNature Biotechnology
Vol/bind36
Udgave nummer2
Sider (fra-til)190-195
Antal sider6
ISSN1087-0156
DOI
StatusUdgivet - 2018

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Ribosomal RNA
rRNA Genes
Genes
Ecosystem
Ecosystems
Small Ribosome Subunits
Chimerism
Poly A
Archaea
Nucleic Acid Databases
Censuses
Eukaryota
Reverse Transcription
Tailings
Transcription
Databases
Bacteria
Throughput
Molecules

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title = "Retrieval of a million high-quality, full-length microbial 16S and 18S rRNA gene sequences without primer bias",
abstract = "Small subunit ribosomal RNA (SSU rRNA) genes, 16S in bacteria and 18S in eukaryotes, have been the standard phylogenetic markers used to characterize microbial diversity and evolution for decades. However, the reference databases of full-length SSU rRNA gene sequences are skewed to well-studied ecosystems and subject to primer bias and chimerism, which results in an incomplete view of the diversity present in a sample. We combine poly(A)-tailing and reverse transcription of SSU rRNA molecules with synthetic long-read sequencing to generate high-quality, full-length SSU rRNA sequences, without primer bias, at high throughput. We apply our approach to samples from seven different ecosystems and obtain more than a million SSU rRNA sequences from all domains of life, with an estimated raw error rate of 0.17{\%}. We observe a large proportion of novel diversity, including several deeply branching phylum-level lineages putatively related to the Asgard Archaea. Our approach will enable expansion of the SSU rRNA reference databases by orders of magnitude, and contribute to a comprehensive census of the tree of life.",
author = "Karst, {S{\o}ren Michael} and Dueholm, {Morten Simonsen} and McIlroy, {Simon Jon} and Kirkegaard, {Rasmus Hansen} and Nielsen, {Per Halkj{\ae}r} and Mads Albertsen",
year = "2018",
doi = "10.1038/nbt.4045",
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TY - JOUR

T1 - Retrieval of a million high-quality, full-length microbial 16S and 18S rRNA gene sequences without primer bias

AU - Karst, Søren Michael

AU - Dueholm, Morten Simonsen

AU - McIlroy, Simon Jon

AU - Kirkegaard, Rasmus Hansen

AU - Nielsen, Per Halkjær

AU - Albertsen, Mads

PY - 2018

Y1 - 2018

N2 - Small subunit ribosomal RNA (SSU rRNA) genes, 16S in bacteria and 18S in eukaryotes, have been the standard phylogenetic markers used to characterize microbial diversity and evolution for decades. However, the reference databases of full-length SSU rRNA gene sequences are skewed to well-studied ecosystems and subject to primer bias and chimerism, which results in an incomplete view of the diversity present in a sample. We combine poly(A)-tailing and reverse transcription of SSU rRNA molecules with synthetic long-read sequencing to generate high-quality, full-length SSU rRNA sequences, without primer bias, at high throughput. We apply our approach to samples from seven different ecosystems and obtain more than a million SSU rRNA sequences from all domains of life, with an estimated raw error rate of 0.17%. We observe a large proportion of novel diversity, including several deeply branching phylum-level lineages putatively related to the Asgard Archaea. Our approach will enable expansion of the SSU rRNA reference databases by orders of magnitude, and contribute to a comprehensive census of the tree of life.

AB - Small subunit ribosomal RNA (SSU rRNA) genes, 16S in bacteria and 18S in eukaryotes, have been the standard phylogenetic markers used to characterize microbial diversity and evolution for decades. However, the reference databases of full-length SSU rRNA gene sequences are skewed to well-studied ecosystems and subject to primer bias and chimerism, which results in an incomplete view of the diversity present in a sample. We combine poly(A)-tailing and reverse transcription of SSU rRNA molecules with synthetic long-read sequencing to generate high-quality, full-length SSU rRNA sequences, without primer bias, at high throughput. We apply our approach to samples from seven different ecosystems and obtain more than a million SSU rRNA sequences from all domains of life, with an estimated raw error rate of 0.17%. We observe a large proportion of novel diversity, including several deeply branching phylum-level lineages putatively related to the Asgard Archaea. Our approach will enable expansion of the SSU rRNA reference databases by orders of magnitude, and contribute to a comprehensive census of the tree of life.

U2 - 10.1038/nbt.4045

DO - 10.1038/nbt.4045

M3 - Letter

VL - 36

SP - 190

EP - 195

JO - Nature Biotechnology

JF - Nature Biotechnology

SN - 1087-0156

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ER -