Sequencing platform and library preparation choices impact viral metagenomes

Solonenko, Sergei A., Ignacio-Espinoza, J. Cesar, Alberti, Adriana, Cruaud, Corinne, Hallam, Steven, Konstantinidis, Kostas, Tyson, Gene, Wincker, Patrick and Sullivan, Matthew B. (2013) Sequencing platform and library preparation choices impact viral metagenomes. Bmc Genomics, 14 1: 320.1-320.12. doi:10.1186/1471-2164-14-320


Author Solonenko, Sergei A.
Ignacio-Espinoza, J. Cesar
Alberti, Adriana
Cruaud, Corinne
Hallam, Steven
Konstantinidis, Kostas
Tyson, Gene
Wincker, Patrick
Sullivan, Matthew B.
Title Sequencing platform and library preparation choices impact viral metagenomes
Journal name Bmc Genomics   Check publisher's open access policy
ISSN 1471-2164
Publication date 2013-05-01
Year available 2013
Sub-type Article (original research)
DOI 10.1186/1471-2164-14-320
Open Access Status DOI
Volume 14
Issue 1
Start page 320.1
End page 320.12
Total pages 12
Place of publication London, United Kingdom
Publisher BioMed Central
Language eng
Abstract Background: Microbes drive the biogeochemistry that fuels the planet. Microbial viruses modulate their hosts directly through mortality and horizontal gene transfer, and indirectly by re-programming host metabolisms during infection. However, our ability to study these virus-host interactions is limited by methods that are low-throughput and heavily reliant upon the subset of organisms that are in culture. One way forward are culture-independent metagenomic approaches, but these novel methods are rarely rigorously tested, especially for studies of environmental viruses, air microbiomes, extreme environment microbiology and other areas with constrained sample amounts. Here we perform replicated experiments to evaluate Roche 454, Illumina HiSeq, and Ion Torrent PGM sequencing and library preparation protocols on virus metagenomes generated from as little as 10pg of DNA.
Formatted abstract
Background: Microbes drive the biogeochemistry that fuels the planet. Microbial viruses modulate their hosts directly through mortality and horizontal gene transfer, and indirectly by re-programming host metabolisms during infection. However, our ability to study these virus-host interactions is limited by methods that are low-throughput and heavily reliant upon the subset of organisms that are in culture. One way forward are culture-independent metagenomic approaches, but these novel methods are rarely rigorously tested, especially for studies of environmental viruses, air microbiomes, extreme environment microbiology and other areas with constrained sample amounts. Here we perform replicated experiments to evaluate Roche 454, Illumina HiSeq, and Ion Torrent PGM sequencing and library preparation protocols on virus metagenomes generated from as little as 10pg of DNA.

Results: Using %G + C content to compare metagenomes, we find that (i) metagenomes are highly replicable, (ii) some treatment effects are minimal, e.g., sequencing technology choice has 6-fold less impact than varying input DNA amount, and (iii) when restricted to a limited DNA concentration (<1μg), changing the amount of amplification produces little variation. These trends were also observed when examining the metagenomes for gene function and assembly performance, although the latter more closely aligned to sequencing effort and read length than preparation steps tested. Among Illumina library preparation options, transposon-based libraries diverged from all others and adaptor ligation was a critical step for optimizing sequencing yields.

Conclusions: These data guide researchers in generating systematic, comparative datasets to understand complex ecosystems, and suggest that neither varied amplification nor sequencing platforms will deter such efforts.
Keyword Ocean Viruses
Dna
Amplification
Bias
Genomes
Q-Index Code C1
Q-Index Status Confirmed Code
Grant ID POSEIDON/ANR-09-BLAN-0348
287589
Institutional Status UQ

Document type: Journal Article
Sub-type: Article (original research)
Collections: Official 2014 Collection
School of Information Technology and Electrical Engineering Publications
 
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