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A Direct PCR Approach to Accelerate Analyses of Human-Associated Microbial Communities

Since the composition of the human microbiome is highly variable both within and between individuals, researchers are increasingly reliant on high-throughput molecular approaches to identify linkages between the composition of these communities and human health. While new sequencing technologies hav...

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Detalles Bibliográficos
Autores principales: Flores, Gilberto E., Henley, Jessica B., Fierer, Noah
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Public Library of Science 2012
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3433448/
https://www.ncbi.nlm.nih.gov/pubmed/22962617
http://dx.doi.org/10.1371/journal.pone.0044563
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author Flores, Gilberto E.
Henley, Jessica B.
Fierer, Noah
author_facet Flores, Gilberto E.
Henley, Jessica B.
Fierer, Noah
author_sort Flores, Gilberto E.
collection PubMed
description Since the composition of the human microbiome is highly variable both within and between individuals, researchers are increasingly reliant on high-throughput molecular approaches to identify linkages between the composition of these communities and human health. While new sequencing technologies have made it increasingly feasible to analyze large numbers of human-associated samples, the extraction of DNA from samples often remains a bottleneck in the process. Here we tested a direct PCR approach using the Extract-N-Amp Plant PCR Kit to accelerate the 16S rRNA gene-based analyses of human-associated bacterial communities, directly comparing this method to a more commonly-used approach whereby DNA is first extracted and purified from samples using a series of steps prior to PCR amplification. We used both approaches on replicate samples collected from each of five body habitats (tongue surface, feces, forehead skin, underarm skin, and forearm skin) from four individuals. With the exception of the tongue samples, there were few significant differences in the estimates of taxon richness or phylogenetic diversity obtained using the two approaches. Perhaps more importantly, there were no significant differences between the methods in their ability resolve body habitat differences or inter-individual differences in bacterial community composition and the estimates of the relative abundances of individual taxa were nearly identical with the two methods. Overall, the two methods gave very similar results and the direct PCR approach is clearly advantageous for many studies exploring the diversity and composition of human-associated bacterial communities given that large numbers of samples can be processed far more quickly and efficiently.
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spelling pubmed-34334482012-09-07 A Direct PCR Approach to Accelerate Analyses of Human-Associated Microbial Communities Flores, Gilberto E. Henley, Jessica B. Fierer, Noah PLoS One Research Article Since the composition of the human microbiome is highly variable both within and between individuals, researchers are increasingly reliant on high-throughput molecular approaches to identify linkages between the composition of these communities and human health. While new sequencing technologies have made it increasingly feasible to analyze large numbers of human-associated samples, the extraction of DNA from samples often remains a bottleneck in the process. Here we tested a direct PCR approach using the Extract-N-Amp Plant PCR Kit to accelerate the 16S rRNA gene-based analyses of human-associated bacterial communities, directly comparing this method to a more commonly-used approach whereby DNA is first extracted and purified from samples using a series of steps prior to PCR amplification. We used both approaches on replicate samples collected from each of five body habitats (tongue surface, feces, forehead skin, underarm skin, and forearm skin) from four individuals. With the exception of the tongue samples, there were few significant differences in the estimates of taxon richness or phylogenetic diversity obtained using the two approaches. Perhaps more importantly, there were no significant differences between the methods in their ability resolve body habitat differences or inter-individual differences in bacterial community composition and the estimates of the relative abundances of individual taxa were nearly identical with the two methods. Overall, the two methods gave very similar results and the direct PCR approach is clearly advantageous for many studies exploring the diversity and composition of human-associated bacterial communities given that large numbers of samples can be processed far more quickly and efficiently. Public Library of Science 2012-09-04 /pmc/articles/PMC3433448/ /pubmed/22962617 http://dx.doi.org/10.1371/journal.pone.0044563 Text en © 2012 Flores et al http://creativecommons.org/licenses/by/4.0/ This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are properly credited.
spellingShingle Research Article
Flores, Gilberto E.
Henley, Jessica B.
Fierer, Noah
A Direct PCR Approach to Accelerate Analyses of Human-Associated Microbial Communities
title A Direct PCR Approach to Accelerate Analyses of Human-Associated Microbial Communities
title_full A Direct PCR Approach to Accelerate Analyses of Human-Associated Microbial Communities
title_fullStr A Direct PCR Approach to Accelerate Analyses of Human-Associated Microbial Communities
title_full_unstemmed A Direct PCR Approach to Accelerate Analyses of Human-Associated Microbial Communities
title_short A Direct PCR Approach to Accelerate Analyses of Human-Associated Microbial Communities
title_sort direct pcr approach to accelerate analyses of human-associated microbial communities
topic Research Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3433448/
https://www.ncbi.nlm.nih.gov/pubmed/22962617
http://dx.doi.org/10.1371/journal.pone.0044563
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