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Comparative evaluation of four bacteria-specific primer pairs for 16S rRNA gene surveys

Thijs, Sofie; De Beeck, Michiel Op LU ; Beckers, Bram; Truyens, Sascha; Stevens, Vincent; Van Hamme, Jonathan D.; Weyens, Nele and Vangronsveld, Jaco (2017) In Frontiers in Microbiology 8(MAR).
Abstract

Bacterial taxonomic community analyses using PCR-amplification of the 16S rRNA gene and high-throughput sequencing has become a cornerstone in microbiology research. To reliably detect the members, or operational taxonomic units (OTUs), that make up bacterial communities, taxonomic surveys rely on the use of the most informative PCR primers to amplify the broad range of phylotypes present in up-to-date reference databases. However, primers specific for the domain Bacteria were often developed some time ago against database versions that are now out of date. Here we evaluated the performance of four bacterial primers for characterizing complex microbial communities in explosives contaminated and non-contaminated forest soil and by in... (More)

Bacterial taxonomic community analyses using PCR-amplification of the 16S rRNA gene and high-throughput sequencing has become a cornerstone in microbiology research. To reliably detect the members, or operational taxonomic units (OTUs), that make up bacterial communities, taxonomic surveys rely on the use of the most informative PCR primers to amplify the broad range of phylotypes present in up-to-date reference databases. However, primers specific for the domain Bacteria were often developed some time ago against database versions that are now out of date. Here we evaluated the performance of four bacterial primers for characterizing complex microbial communities in explosives contaminated and non-contaminated forest soil and by in silico evaluation against the current SILVA123 database. Primer pair 341f/785r produced the highest number of bacterial OTUs, phylogenetic richness, Shannon diversity, low non-specificity and most reproducible results, followed by 967f/1391r and 799f/1193r. Primer pair 68f/518r showed overall low coverage and a bias toward Alphaproteobacteria. In silico, primer pair 341f/785r showed the highest coverage of the domain Bacteria (96.1%) with no obvious bias toward the majority of bacterial species. This suggests the high utility of primer pair 341f/785r for soil and plant-associated bacterial microbiome studies.

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organization
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Contribution to journal
publication status
published
subject
keywords
16S rRNA gene sequence primers, Explosives contamination, Microbial communities, Pyrosequencing, Soil
in
Frontiers in Microbiology
volume
8
issue
MAR
publisher
Frontiers
external identifiers
  • scopus:85016570043
  • wos:000397611700001
ISSN
1664-302X
DOI
10.3389/fmicb.2017.00494
language
English
LU publication?
yes
id
334bc2c1-2086-440a-901a-b1e792285dbb
date added to LUP
2017-05-04 07:02:15
date last changed
2017-09-18 13:33:58
@article{334bc2c1-2086-440a-901a-b1e792285dbb,
  abstract     = {<p>Bacterial taxonomic community analyses using PCR-amplification of the 16S rRNA gene and high-throughput sequencing has become a cornerstone in microbiology research. To reliably detect the members, or operational taxonomic units (OTUs), that make up bacterial communities, taxonomic surveys rely on the use of the most informative PCR primers to amplify the broad range of phylotypes present in up-to-date reference databases. However, primers specific for the domain Bacteria were often developed some time ago against database versions that are now out of date. Here we evaluated the performance of four bacterial primers for characterizing complex microbial communities in explosives contaminated and non-contaminated forest soil and by in silico evaluation against the current SILVA123 database. Primer pair 341f/785r produced the highest number of bacterial OTUs, phylogenetic richness, Shannon diversity, low non-specificity and most reproducible results, followed by 967f/1391r and 799f/1193r. Primer pair 68f/518r showed overall low coverage and a bias toward Alphaproteobacteria. In silico, primer pair 341f/785r showed the highest coverage of the domain Bacteria (96.1%) with no obvious bias toward the majority of bacterial species. This suggests the high utility of primer pair 341f/785r for soil and plant-associated bacterial microbiome studies.</p>},
  articleno    = {494},
  author       = {Thijs, Sofie and De Beeck, Michiel Op and Beckers, Bram and Truyens, Sascha and Stevens, Vincent and Van Hamme, Jonathan D. and Weyens, Nele and Vangronsveld, Jaco},
  issn         = {1664-302X},
  keyword      = {16S rRNA gene sequence primers,Explosives contamination,Microbial communities,Pyrosequencing,Soil},
  language     = {eng},
  month        = {03},
  number       = {MAR},
  publisher    = {Frontiers},
  series       = {Frontiers in Microbiology},
  title        = {Comparative evaluation of four bacteria-specific primer pairs for 16S rRNA gene surveys},
  url          = {http://dx.doi.org/10.3389/fmicb.2017.00494},
  volume       = {8},
  year         = {2017},
}