Assembling the marine metagenome, one cell at a time.

The difficulty associated with the cultivation of most microorganisms and the complexity of natural microbial assemblages, such as marine plankton or human microbiome, hinder genome reconstruction of representative taxa using cultivation or metagenomic approaches. Here we used an alternative, single...

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Autores principales:	Tanja Woyke, Gary Xie, Alex Copeland, José M González, Cliff Han, Hajnalka Kiss, Jimmy H Saw, Pavel Senin, Chi Yang, Sourav Chatterji, Jan-Fang Cheng, Jonathan A Eisen, Michael E Sieracki, Ramunas Stepanauskas
Formato:	article
Lenguaje:	EN
Publicado:	Public Library of Science (PLoS) 2009
Materias:	Medicine R Science Q
Acceso en línea:	https://doaj.org/article/27edcef369fd44bd9c535ef4ad449ad4
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spelling	oai:doaj.org-article:27edcef369fd44bd9c535ef4ad449ad42021-11-25T06:23:06ZAssembling the marine metagenome, one cell at a time.1932-620310.1371/journal.pone.0005299https://doaj.org/article/27edcef369fd44bd9c535ef4ad449ad42009-01-01T00:00:00Zhttps://www.ncbi.nlm.nih.gov/pmc/articles/pmid/19390573/pdf/?tool=EBIhttps://doaj.org/toc/1932-6203The difficulty associated with the cultivation of most microorganisms and the complexity of natural microbial assemblages, such as marine plankton or human microbiome, hinder genome reconstruction of representative taxa using cultivation or metagenomic approaches. Here we used an alternative, single cell sequencing approach to obtain high-quality genome assemblies of two uncultured, numerically significant marine microorganisms. We employed fluorescence-activated cell sorting and multiple displacement amplification to obtain hundreds of micrograms of genomic DNA from individual, uncultured cells of two marine flavobacteria from the Gulf of Maine that were phylogenetically distant from existing cultured strains. Shotgun sequencing and genome finishing yielded 1.9 Mbp in 17 contigs and 1.5 Mbp in 21 contigs for the two flavobacteria, with estimated genome recoveries of about 91% and 78%, respectively. Only 0.24% of the assembling sequences were contaminants and were removed from further analysis using rigorous quality control. In contrast to all cultured strains of marine flavobacteria, the two single cell genomes were excellent Global Ocean Sampling (GOS) metagenome fragment recruiters, demonstrating their numerical significance in the ocean. The geographic distribution of GOS recruits along the Northwest Atlantic coast coincided with ocean surface currents. Metabolic reconstruction indicated diverse potential energy sources, including biopolymer degradation, proteorhodopsin photometabolism, and hydrogen oxidation. Compared to cultured relatives, the two uncultured flavobacteria have small genome sizes, few non-coding nucleotides, and few paralogous genes, suggesting adaptations to narrow ecological niches. These features may have contributed to the abundance of the two taxa in specific regions of the ocean, and may have hindered their cultivation. We demonstrate the power of single cell DNA sequencing to generate reference genomes of uncultured taxa from a complex microbial community of marine bacterioplankton. A combination of single cell genomics and metagenomics enabled us to analyze the genome content, metabolic adaptations, and biogeography of these taxa.Tanja WoykeGary XieAlex CopelandJosé M GonzálezCliff HanHajnalka KissJimmy H SawPavel SeninChi YangSourav ChatterjiJan-Fang ChengJonathan A EisenMichael E SierackiRamunas StepanauskasPublic Library of Science (PLoS)articleMedicineRScienceQENPLoS ONE, Vol 4, Iss 4, p e5299 (2009)
institution	DOAJ
collection	DOAJ
language	EN
topic	Medicine R Science Q
spellingShingle	Medicine R Science Q Tanja Woyke Gary Xie Alex Copeland José M González Cliff Han Hajnalka Kiss Jimmy H Saw Pavel Senin Chi Yang Sourav Chatterji Jan-Fang Cheng Jonathan A Eisen Michael E Sieracki Ramunas Stepanauskas Assembling the marine metagenome, one cell at a time.
description	The difficulty associated with the cultivation of most microorganisms and the complexity of natural microbial assemblages, such as marine plankton or human microbiome, hinder genome reconstruction of representative taxa using cultivation or metagenomic approaches. Here we used an alternative, single cell sequencing approach to obtain high-quality genome assemblies of two uncultured, numerically significant marine microorganisms. We employed fluorescence-activated cell sorting and multiple displacement amplification to obtain hundreds of micrograms of genomic DNA from individual, uncultured cells of two marine flavobacteria from the Gulf of Maine that were phylogenetically distant from existing cultured strains. Shotgun sequencing and genome finishing yielded 1.9 Mbp in 17 contigs and 1.5 Mbp in 21 contigs for the two flavobacteria, with estimated genome recoveries of about 91% and 78%, respectively. Only 0.24% of the assembling sequences were contaminants and were removed from further analysis using rigorous quality control. In contrast to all cultured strains of marine flavobacteria, the two single cell genomes were excellent Global Ocean Sampling (GOS) metagenome fragment recruiters, demonstrating their numerical significance in the ocean. The geographic distribution of GOS recruits along the Northwest Atlantic coast coincided with ocean surface currents. Metabolic reconstruction indicated diverse potential energy sources, including biopolymer degradation, proteorhodopsin photometabolism, and hydrogen oxidation. Compared to cultured relatives, the two uncultured flavobacteria have small genome sizes, few non-coding nucleotides, and few paralogous genes, suggesting adaptations to narrow ecological niches. These features may have contributed to the abundance of the two taxa in specific regions of the ocean, and may have hindered their cultivation. We demonstrate the power of single cell DNA sequencing to generate reference genomes of uncultured taxa from a complex microbial community of marine bacterioplankton. A combination of single cell genomics and metagenomics enabled us to analyze the genome content, metabolic adaptations, and biogeography of these taxa.
format	article
author	Tanja Woyke Gary Xie Alex Copeland José M González Cliff Han Hajnalka Kiss Jimmy H Saw Pavel Senin Chi Yang Sourav Chatterji Jan-Fang Cheng Jonathan A Eisen Michael E Sieracki Ramunas Stepanauskas
author_facet	Tanja Woyke Gary Xie Alex Copeland José M González Cliff Han Hajnalka Kiss Jimmy H Saw Pavel Senin Chi Yang Sourav Chatterji Jan-Fang Cheng Jonathan A Eisen Michael E Sieracki Ramunas Stepanauskas
author_sort	Tanja Woyke
title	Assembling the marine metagenome, one cell at a time.
title_short	Assembling the marine metagenome, one cell at a time.
title_full	Assembling the marine metagenome, one cell at a time.
title_fullStr	Assembling the marine metagenome, one cell at a time.
title_full_unstemmed	Assembling the marine metagenome, one cell at a time.
title_sort	assembling the marine metagenome, one cell at a time.
publisher	Public Library of Science (PLoS)
publishDate	2009
url	https://doaj.org/article/27edcef369fd44bd9c535ef4ad449ad4
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Assembling the marine metagenome, one cell at a time.

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