Moleculo Long-Read Sequencing Facilitates Assembly and Genomic Binning from Complex Soil Metagenomes

ABSTRACT Soil metagenomics has been touted as the “grand challenge” for metagenomics, as the high microbial diversity and spatial heterogeneity of soils make them unamenable to current assembly platforms. Here, we aimed to improve soil metagenomic sequence assembly by applying the Moleculo synthetic...

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Autores principales:	Richard Allen White, Eric M. Bottos, Taniya Roy Chowdhury, Jeremy D. Zucker, Colin J. Brislawn, Carrie D. Nicora, Sarah J. Fansler, Kurt R. Glaesemann, Kevin Glass, Janet K. Jansson
Formato:	article
Lenguaje:	EN
Publicado:	American Society for Microbiology 2016
Materias:	de novo assembly Moleculo metagenomic assembly metagenomic binning soil metagenomics Microbiology QR1-502
Acceso en línea:	https://doaj.org/article/b105ad52844f467796127d5775060196
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spelling	oai:doaj.org-article:b105ad52844f467796127d57750601962021-12-02T18:15:43ZMoleculo Long-Read Sequencing Facilitates Assembly and Genomic Binning from Complex Soil Metagenomes10.1128/mSystems.00045-162379-5077https://doaj.org/article/b105ad52844f467796127d57750601962016-06-01T00:00:00Zhttps://journals.asm.org/doi/10.1128/mSystems.00045-16https://doaj.org/toc/2379-5077ABSTRACT Soil metagenomics has been touted as the “grand challenge” for metagenomics, as the high microbial diversity and spatial heterogeneity of soils make them unamenable to current assembly platforms. Here, we aimed to improve soil metagenomic sequence assembly by applying the Moleculo synthetic long-read sequencing technology. In total, we obtained 267 Gbp of raw sequence data from a native prairie soil; these data included 109.7 Gbp of short-read data (~100 bp) from the Joint Genome Institute (JGI), an additional 87.7 Gbp of rapid-mode read data (~250 bp), plus 69.6 Gbp (>1.5 kbp) from Moleculo sequencing. The Moleculo data alone yielded over 5,600 reads of >10 kbp in length, and over 95% of the unassembled reads mapped to contigs of >1.5 kbp. Hybrid assembly of all data resulted in more than 10,000 contigs over 10 kbp in length. We mapped three replicate metatranscriptomes derived from the same parent soil to the Moleculo subassembly and found that 95% of the predicted genes, based on their assignments to Enzyme Commission (EC) numbers, were expressed. The Moleculo subassembly also enabled binning of >100 microbial genome bins. We obtained via direct binning the first complete genome, that of “Candidatus Pseudomonas sp. strain JKJ-1” from a native soil metagenome. By mapping metatranscriptome sequence reads back to the bins, we found that several bins corresponding to low-relative-abundance Acidobacteria were highly transcriptionally active, whereas bins corresponding to high-relative-abundance Verrucomicrobia were not. These results demonstrate that Moleculo sequencing provides a significant advance for resolving complex soil microbial communities. IMPORTANCE Soil microorganisms carry out key processes for life on our planet, including cycling of carbon and other nutrients and supporting growth of plants. However, there is poor molecular-level understanding of their functional roles in ecosystem stability and responses to environmental perturbations. This knowledge gap is largely due to the difficulty in culturing the majority of soil microbes. Thus, use of culture-independent approaches, such as metagenomics, promises the direct assessment of the functional potential of soil microbiomes. Soil is, however, a challenge for metagenomic assembly due to its high microbial diversity and variable evenness, resulting in low coverage and uneven sampling of microbial genomes. Despite increasingly large soil metagenome data volumes (>200 Gbp), the majority of the data do not assemble. Here, we used the cutting-edge approach of synthetic long-read sequencing technology (Moleculo) to assemble soil metagenome sequence data into long contigs and used the assemblies for binning of genomes. Author Video: An author video summary of this article is available.Richard Allen WhiteEric M. BottosTaniya Roy ChowdhuryJeremy D. ZuckerColin J. BrislawnCarrie D. NicoraSarah J. FanslerKurt R. GlaesemannKevin GlassJanet K. JanssonAmerican Society for Microbiologyarticlede novo assemblyMoleculometagenomic assemblymetagenomic binningsoil metagenomicsMicrobiologyQR1-502ENmSystems, Vol 1, Iss 3 (2016)
institution	DOAJ
collection	DOAJ
language	EN
topic	de novo assembly Moleculo metagenomic assembly metagenomic binning soil metagenomics Microbiology QR1-502
spellingShingle	de novo assembly Moleculo metagenomic assembly metagenomic binning soil metagenomics Microbiology QR1-502 Richard Allen White Eric M. Bottos Taniya Roy Chowdhury Jeremy D. Zucker Colin J. Brislawn Carrie D. Nicora Sarah J. Fansler Kurt R. Glaesemann Kevin Glass Janet K. Jansson Moleculo Long-Read Sequencing Facilitates Assembly and Genomic Binning from Complex Soil Metagenomes
description	ABSTRACT Soil metagenomics has been touted as the “grand challenge” for metagenomics, as the high microbial diversity and spatial heterogeneity of soils make them unamenable to current assembly platforms. Here, we aimed to improve soil metagenomic sequence assembly by applying the Moleculo synthetic long-read sequencing technology. In total, we obtained 267 Gbp of raw sequence data from a native prairie soil; these data included 109.7 Gbp of short-read data (~100 bp) from the Joint Genome Institute (JGI), an additional 87.7 Gbp of rapid-mode read data (~250 bp), plus 69.6 Gbp (>1.5 kbp) from Moleculo sequencing. The Moleculo data alone yielded over 5,600 reads of >10 kbp in length, and over 95% of the unassembled reads mapped to contigs of >1.5 kbp. Hybrid assembly of all data resulted in more than 10,000 contigs over 10 kbp in length. We mapped three replicate metatranscriptomes derived from the same parent soil to the Moleculo subassembly and found that 95% of the predicted genes, based on their assignments to Enzyme Commission (EC) numbers, were expressed. The Moleculo subassembly also enabled binning of >100 microbial genome bins. We obtained via direct binning the first complete genome, that of “Candidatus Pseudomonas sp. strain JKJ-1” from a native soil metagenome. By mapping metatranscriptome sequence reads back to the bins, we found that several bins corresponding to low-relative-abundance Acidobacteria were highly transcriptionally active, whereas bins corresponding to high-relative-abundance Verrucomicrobia were not. These results demonstrate that Moleculo sequencing provides a significant advance for resolving complex soil microbial communities. IMPORTANCE Soil microorganisms carry out key processes for life on our planet, including cycling of carbon and other nutrients and supporting growth of plants. However, there is poor molecular-level understanding of their functional roles in ecosystem stability and responses to environmental perturbations. This knowledge gap is largely due to the difficulty in culturing the majority of soil microbes. Thus, use of culture-independent approaches, such as metagenomics, promises the direct assessment of the functional potential of soil microbiomes. Soil is, however, a challenge for metagenomic assembly due to its high microbial diversity and variable evenness, resulting in low coverage and uneven sampling of microbial genomes. Despite increasingly large soil metagenome data volumes (>200 Gbp), the majority of the data do not assemble. Here, we used the cutting-edge approach of synthetic long-read sequencing technology (Moleculo) to assemble soil metagenome sequence data into long contigs and used the assemblies for binning of genomes. Author Video: An author video summary of this article is available.
format	article
author	Richard Allen White Eric M. Bottos Taniya Roy Chowdhury Jeremy D. Zucker Colin J. Brislawn Carrie D. Nicora Sarah J. Fansler Kurt R. Glaesemann Kevin Glass Janet K. Jansson
author_facet	Richard Allen White Eric M. Bottos Taniya Roy Chowdhury Jeremy D. Zucker Colin J. Brislawn Carrie D. Nicora Sarah J. Fansler Kurt R. Glaesemann Kevin Glass Janet K. Jansson
author_sort	Richard Allen White
title	Moleculo Long-Read Sequencing Facilitates Assembly and Genomic Binning from Complex Soil Metagenomes
title_short	Moleculo Long-Read Sequencing Facilitates Assembly and Genomic Binning from Complex Soil Metagenomes
title_full	Moleculo Long-Read Sequencing Facilitates Assembly and Genomic Binning from Complex Soil Metagenomes
title_fullStr	Moleculo Long-Read Sequencing Facilitates Assembly and Genomic Binning from Complex Soil Metagenomes
title_full_unstemmed	Moleculo Long-Read Sequencing Facilitates Assembly and Genomic Binning from Complex Soil Metagenomes
title_sort	moleculo long-read sequencing facilitates assembly and genomic binning from complex soil metagenomes
publisher	American Society for Microbiology
publishDate	2016
url	https://doaj.org/article/b105ad52844f467796127d5775060196
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_version_	1718378353944166400

Moleculo Long-Read Sequencing Facilitates Assembly and Genomic Binning from Complex Soil Metagenomes

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