Microbial Community Profiling Protocol with Full-length 16S rRNA Sequences and Emu

Kristen D. Curry; Sirena Soriano; Michael G. Nute; Sonia Villapol; Alexander Dilthey; Todd J. Treangen

doi:10.1002/cpz1.978

Microbial Community Profiling Protocol with Full-length 16S rRNA Sequences and Emu

Kristen D. Curry, Sirena Soriano, Michael G. Nute, Sonia Villapol, Alexander Dilthey, Todd J. Treangen

Research output: Contribution to journal › Article › peer-review

Abstract

16S rRNA targeted amplicon sequencing is an established standard for elucidating microbial community composition. While high-throughput short-read sequencing can elicit only a portion of the 16S rRNA gene due to their limited read length, third generation sequencing can read the 16S rRNA gene in its entirety and thus provide more precise taxonomic classification. Here, we present a protocol for generating full-length 16S rRNA sequences with Oxford Nanopore Technologies (ONT) and a microbial community profile with Emu. We select Emu for analyzing ONT sequences as it leverages information from the entire community to overcome errors due to incomplete reference databases and hardware limitations to ultimately obtain species-level resolution. This pipeline provides a low-cost solution for characterizing microbiome composition by exploiting real-time, long-read ONT sequencing and tailored software for accurate characterization of microbial communities. © 2024 Wiley Periodicals LLC. Basic Protocol: Microbial community profiling with Emu Support Protocol 1: Full-length 16S rRNA microbial sequences with Oxford Nanopore Technologies sequencing platform Support Protocol 2: Building a custom reference database for Emu.

Original language	English (US)
Article number	e978
Pages (from-to)	e978
Journal	Current Protocols
Volume	4
Issue number	3
DOIs	https://doi.org/10.1002/cpz1.978
State	Published - Mar 2024

Keywords

16S rRNA
Oxford Nanopore Technologies
bioinformatics
community profile
long-read sequencing
microbiome
species
Dromaiidae/genetics
Bacteria/genetics
RNA, Ribosomal, 16S/genetics
Sequence Analysis, DNA/methods
Microbiota/genetics
Animals

ASJC Scopus subject areas

Neuroscience(all)
Biochemistry, Genetics and Molecular Biology(all)
Immunology and Microbiology(all)
Pharmacology, Toxicology and Pharmaceutics(all)
Health Informatics
Medical Laboratory Technology

Access to Document

10.1002/cpz1.978

Cite this

@article{0cf81a091bff47feb6826aad733b0557,

title = "Microbial Community Profiling Protocol with Full-length 16S rRNA Sequences and Emu",

abstract = "16S rRNA targeted amplicon sequencing is an established standard for elucidating microbial community composition. While high-throughput short-read sequencing can elicit only a portion of the 16S rRNA gene due to their limited read length, third generation sequencing can read the 16S rRNA gene in its entirety and thus provide more precise taxonomic classification. Here, we present a protocol for generating full-length 16S rRNA sequences with Oxford Nanopore Technologies (ONT) and a microbial community profile with Emu. We select Emu for analyzing ONT sequences as it leverages information from the entire community to overcome errors due to incomplete reference databases and hardware limitations to ultimately obtain species-level resolution. This pipeline provides a low-cost solution for characterizing microbiome composition by exploiting real-time, long-read ONT sequencing and tailored software for accurate characterization of microbial communities. {\textcopyright} 2024 Wiley Periodicals LLC. Basic Protocol: Microbial community profiling with Emu Support Protocol 1: Full-length 16S rRNA microbial sequences with Oxford Nanopore Technologies sequencing platform Support Protocol 2: Building a custom reference database for Emu.",

keywords = "16S rRNA, Oxford Nanopore Technologies, bioinformatics, community profile, long-read sequencing, microbiome, species, Dromaiidae/genetics, Bacteria/genetics, RNA, Ribosomal, 16S/genetics, Sequence Analysis, DNA/methods, Microbiota/genetics, Animals",

author = "Curry, {Kristen D.} and Sirena Soriano and Nute, {Michael G.} and Sonia Villapol and Alexander Dilthey and Treangen, {Todd J.}",

note = "{\textcopyright} 2024 Wiley Periodicals LLC.",

year = "2024",

month = mar,

doi = "10.1002/cpz1.978",

language = "English (US)",

volume = "4",

pages = "e978",

journal = "Current Protocols",

issn = "2691-1299",

publisher = "John Wiley & Sons Inc.",

number = "3",

}

TY - JOUR

T1 - Microbial Community Profiling Protocol with Full-length 16S rRNA Sequences and Emu

AU - Curry, Kristen D.

AU - Soriano, Sirena

AU - Nute, Michael G.

AU - Villapol, Sonia

AU - Dilthey, Alexander

AU - Treangen, Todd J.

PY - 2024/3

Y1 - 2024/3

N2 - 16S rRNA targeted amplicon sequencing is an established standard for elucidating microbial community composition. While high-throughput short-read sequencing can elicit only a portion of the 16S rRNA gene due to their limited read length, third generation sequencing can read the 16S rRNA gene in its entirety and thus provide more precise taxonomic classification. Here, we present a protocol for generating full-length 16S rRNA sequences with Oxford Nanopore Technologies (ONT) and a microbial community profile with Emu. We select Emu for analyzing ONT sequences as it leverages information from the entire community to overcome errors due to incomplete reference databases and hardware limitations to ultimately obtain species-level resolution. This pipeline provides a low-cost solution for characterizing microbiome composition by exploiting real-time, long-read ONT sequencing and tailored software for accurate characterization of microbial communities. © 2024 Wiley Periodicals LLC. Basic Protocol: Microbial community profiling with Emu Support Protocol 1: Full-length 16S rRNA microbial sequences with Oxford Nanopore Technologies sequencing platform Support Protocol 2: Building a custom reference database for Emu.

AB - 16S rRNA targeted amplicon sequencing is an established standard for elucidating microbial community composition. While high-throughput short-read sequencing can elicit only a portion of the 16S rRNA gene due to their limited read length, third generation sequencing can read the 16S rRNA gene in its entirety and thus provide more precise taxonomic classification. Here, we present a protocol for generating full-length 16S rRNA sequences with Oxford Nanopore Technologies (ONT) and a microbial community profile with Emu. We select Emu for analyzing ONT sequences as it leverages information from the entire community to overcome errors due to incomplete reference databases and hardware limitations to ultimately obtain species-level resolution. This pipeline provides a low-cost solution for characterizing microbiome composition by exploiting real-time, long-read ONT sequencing and tailored software for accurate characterization of microbial communities. © 2024 Wiley Periodicals LLC. Basic Protocol: Microbial community profiling with Emu Support Protocol 1: Full-length 16S rRNA microbial sequences with Oxford Nanopore Technologies sequencing platform Support Protocol 2: Building a custom reference database for Emu.

KW - 16S rRNA

KW - Oxford Nanopore Technologies

KW - bioinformatics

KW - community profile

KW - long-read sequencing

KW - microbiome

KW - species

KW - Dromaiidae/genetics

KW - Bacteria/genetics

KW - RNA, Ribosomal, 16S/genetics

KW - Sequence Analysis, DNA/methods

KW - Microbiota/genetics

KW - Animals

UR - http://www.scopus.com/inward/record.url?scp=85188322365&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=85188322365&partnerID=8YFLogxK

U2 - 10.1002/cpz1.978

DO - 10.1002/cpz1.978

M3 - Article

C2 - 38511467

AN - SCOPUS:85188322365

SN - 2691-1299

VL - 4

SP - e978

JO - Current Protocols

JF - Current Protocols

IS - 3

M1 - e978

ER -

Microbial Community Profiling Protocol with Full-length 16S rRNA Sequences and Emu

Abstract

Keywords

ASJC Scopus subject areas

Access to Document

Other files and links

Fingerprint

Cite this