Critical review of 16S rRNA gene sequencing workflow in microbiome studies: From primer selection to advanced data analysis.
16S rRNA gene
bioinformatics
microbiome
primer
sequencing
statistical analysis
Journal
Molecular oral microbiology
ISSN: 2041-1014
Titre abrégé: Mol Oral Microbiol
Pays: Denmark
ID NLM: 101524770
Informations de publication
Date de publication:
Oct 2023
Oct 2023
Historique:
revised:
01
09
2023
received:
26
04
2023
accepted:
14
09
2023
medline:
1
11
2023
pubmed:
7
10
2023
entrez:
7
10
2023
Statut:
ppublish
Résumé
The multi-batch reanalysis approach of jointly reevaluating gene/genome sequences from different works has gained particular relevance in the literature in recent years. The large amount of 16S ribosomal ribonucleic acid (rRNA) gene sequence data stored in public repositories and information in taxonomic databases of the same gene far exceeds that related to complete genomes. This review is intended to guide researchers new to studying microbiota, particularly the oral microbiota, using 16S rRNA gene sequencing and those who want to expand and update their knowledge to optimise their decision-making and improve their research results. First, we describe the advantages and disadvantages of using the 16S rRNA gene as a phylogenetic marker and the latest findings on the impact of primer pair selection on diversity and taxonomic assignment outcomes in oral microbiome studies. Strategies for primer selection based on these results are introduced. Second, we identified the key factors to consider in selecting the sequencing technology and platform. The process and particularities of the main steps for processing 16S rRNA gene-derived data are described in detail to enable researchers to choose the most appropriate bioinformatics pipeline and analysis methods based on the available evidence. We then produce an overview of the different types of advanced analyses, both the most widely used in the literature and the most recent approaches. Several indices, metrics and software for studying microbial communities are included, highlighting their advantages and disadvantages. Considering the principles of clinical metagenomics, we conclude that future research should focus on rigorous analytical approaches, such as developing predictive models to identify microbiome-based biomarkers to classify health and disease states. Finally, we address the batch effect concept and the microbiome-specific methods for accounting for or correcting them.
Substances chimiques
RNA, Ribosomal, 16S
0
Types de publication
Journal Article
Review
Langues
eng
Pagination
347-399Subventions
Organisme : Instituto de Salud Carlos III (ISCIII) and co-funded by the European Union
ID : PI21/00588
Informations de copyright
© 2023 The Authors. Molecular Oral Microbiology published by John Wiley & Sons Ltd.
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