Cargando…
Analysis of the Cystic Fibrosis Lung Microbiota via Serial Illumina Sequencing of Bacterial 16S rRNA Hypervariable Regions
The characterization of bacterial communities using DNA sequencing has revolutionized our ability to study microbes in nature and discover the ways in which microbial communities affect ecosystem functioning and human health. Here we describe Serial Illumina Sequencing (SI-Seq): a method for deep se...
Autores principales: | , , , , , , , , , , , , , |
---|---|
Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
Public Library of Science
2012
|
Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3462755/ https://www.ncbi.nlm.nih.gov/pubmed/23056217 http://dx.doi.org/10.1371/journal.pone.0045791 |
_version_ | 1782245205479522304 |
---|---|
author | Maughan, Heather Wang, Pauline W. Diaz Caballero, Julio Fung, Pauline Gong, Yunchen Donaldson, Sylva L. Yuan, Lijie Keshavjee, Shaf Zhang, Yu Yau, Yvonne C. W. Waters, Valerie J. Tullis, D. Elizabeth Hwang, David M. Guttman, David S. |
author_facet | Maughan, Heather Wang, Pauline W. Diaz Caballero, Julio Fung, Pauline Gong, Yunchen Donaldson, Sylva L. Yuan, Lijie Keshavjee, Shaf Zhang, Yu Yau, Yvonne C. W. Waters, Valerie J. Tullis, D. Elizabeth Hwang, David M. Guttman, David S. |
author_sort | Maughan, Heather |
collection | PubMed |
description | The characterization of bacterial communities using DNA sequencing has revolutionized our ability to study microbes in nature and discover the ways in which microbial communities affect ecosystem functioning and human health. Here we describe Serial Illumina Sequencing (SI-Seq): a method for deep sequencing of the bacterial 16S rRNA gene using next-generation sequencing technology. SI-Seq serially sequences portions of the V5, V6 and V7 hypervariable regions from barcoded 16S rRNA amplicons using an Illumina short-read genome analyzer. SI-Seq obtains taxonomic resolution similar to 454 pyrosequencing for a fraction of the cost, and can produce hundreds of thousands of reads per sample even with very high multiplexing. We validated SI-Seq using single species and mock community controls, and via a comparison to cystic fibrosis lung microbiota sequenced using 454 FLX Titanium. Our control runs show that SI-Seq has a dynamic range of at least five orders of magnitude, can classify >96% of sequences to the genus level, and performs just as well as 454 and paired-end Illumina methods in estimation of standard microbial ecology diversity measurements. We illustrate the utility of SI-Seq in a pilot sample of central airway secretion samples from cystic fibrosis patients. |
format | Online Article Text |
id | pubmed-3462755 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2012 |
publisher | Public Library of Science |
record_format | MEDLINE/PubMed |
spelling | pubmed-34627552012-10-10 Analysis of the Cystic Fibrosis Lung Microbiota via Serial Illumina Sequencing of Bacterial 16S rRNA Hypervariable Regions Maughan, Heather Wang, Pauline W. Diaz Caballero, Julio Fung, Pauline Gong, Yunchen Donaldson, Sylva L. Yuan, Lijie Keshavjee, Shaf Zhang, Yu Yau, Yvonne C. W. Waters, Valerie J. Tullis, D. Elizabeth Hwang, David M. Guttman, David S. PLoS One Research Article The characterization of bacterial communities using DNA sequencing has revolutionized our ability to study microbes in nature and discover the ways in which microbial communities affect ecosystem functioning and human health. Here we describe Serial Illumina Sequencing (SI-Seq): a method for deep sequencing of the bacterial 16S rRNA gene using next-generation sequencing technology. SI-Seq serially sequences portions of the V5, V6 and V7 hypervariable regions from barcoded 16S rRNA amplicons using an Illumina short-read genome analyzer. SI-Seq obtains taxonomic resolution similar to 454 pyrosequencing for a fraction of the cost, and can produce hundreds of thousands of reads per sample even with very high multiplexing. We validated SI-Seq using single species and mock community controls, and via a comparison to cystic fibrosis lung microbiota sequenced using 454 FLX Titanium. Our control runs show that SI-Seq has a dynamic range of at least five orders of magnitude, can classify >96% of sequences to the genus level, and performs just as well as 454 and paired-end Illumina methods in estimation of standard microbial ecology diversity measurements. We illustrate the utility of SI-Seq in a pilot sample of central airway secretion samples from cystic fibrosis patients. Public Library of Science 2012-10-02 /pmc/articles/PMC3462755/ /pubmed/23056217 http://dx.doi.org/10.1371/journal.pone.0045791 Text en © 2012 Maughan 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 Maughan, Heather Wang, Pauline W. Diaz Caballero, Julio Fung, Pauline Gong, Yunchen Donaldson, Sylva L. Yuan, Lijie Keshavjee, Shaf Zhang, Yu Yau, Yvonne C. W. Waters, Valerie J. Tullis, D. Elizabeth Hwang, David M. Guttman, David S. Analysis of the Cystic Fibrosis Lung Microbiota via Serial Illumina Sequencing of Bacterial 16S rRNA Hypervariable Regions |
title | Analysis of the Cystic Fibrosis Lung Microbiota via Serial Illumina Sequencing of Bacterial 16S rRNA Hypervariable Regions |
title_full | Analysis of the Cystic Fibrosis Lung Microbiota via Serial Illumina Sequencing of Bacterial 16S rRNA Hypervariable Regions |
title_fullStr | Analysis of the Cystic Fibrosis Lung Microbiota via Serial Illumina Sequencing of Bacterial 16S rRNA Hypervariable Regions |
title_full_unstemmed | Analysis of the Cystic Fibrosis Lung Microbiota via Serial Illumina Sequencing of Bacterial 16S rRNA Hypervariable Regions |
title_short | Analysis of the Cystic Fibrosis Lung Microbiota via Serial Illumina Sequencing of Bacterial 16S rRNA Hypervariable Regions |
title_sort | analysis of the cystic fibrosis lung microbiota via serial illumina sequencing of bacterial 16s rrna hypervariable regions |
topic | Research Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3462755/ https://www.ncbi.nlm.nih.gov/pubmed/23056217 http://dx.doi.org/10.1371/journal.pone.0045791 |
work_keys_str_mv | AT maughanheather analysisofthecysticfibrosislungmicrobiotaviaserialilluminasequencingofbacterial16srrnahypervariableregions AT wangpaulinew analysisofthecysticfibrosislungmicrobiotaviaserialilluminasequencingofbacterial16srrnahypervariableregions AT diazcaballerojulio analysisofthecysticfibrosislungmicrobiotaviaserialilluminasequencingofbacterial16srrnahypervariableregions AT fungpauline analysisofthecysticfibrosislungmicrobiotaviaserialilluminasequencingofbacterial16srrnahypervariableregions AT gongyunchen analysisofthecysticfibrosislungmicrobiotaviaserialilluminasequencingofbacterial16srrnahypervariableregions AT donaldsonsylval analysisofthecysticfibrosislungmicrobiotaviaserialilluminasequencingofbacterial16srrnahypervariableregions AT yuanlijie analysisofthecysticfibrosislungmicrobiotaviaserialilluminasequencingofbacterial16srrnahypervariableregions AT keshavjeeshaf analysisofthecysticfibrosislungmicrobiotaviaserialilluminasequencingofbacterial16srrnahypervariableregions AT zhangyu analysisofthecysticfibrosislungmicrobiotaviaserialilluminasequencingofbacterial16srrnahypervariableregions AT yauyvonnecw analysisofthecysticfibrosislungmicrobiotaviaserialilluminasequencingofbacterial16srrnahypervariableregions AT watersvaleriej analysisofthecysticfibrosislungmicrobiotaviaserialilluminasequencingofbacterial16srrnahypervariableregions AT tullisdelizabeth analysisofthecysticfibrosislungmicrobiotaviaserialilluminasequencingofbacterial16srrnahypervariableregions AT hwangdavidm analysisofthecysticfibrosislungmicrobiotaviaserialilluminasequencingofbacterial16srrnahypervariableregions AT guttmandavids analysisofthecysticfibrosislungmicrobiotaviaserialilluminasequencingofbacterial16srrnahypervariableregions |