title: Whole-genome amplification (WGA) of marine photosynthetic eukaryote populations.
abstract: Metagenomics approaches have been developing rapidly in marine sciences. However, the application of these approaches to marine eukaryotes, and in particular to the smallest ones, is challenging because marine microbial communities are dominated by prokaryotes. One way to circumvent this problem is to separate eukaryotic cells using techniques such as single-cell pipetting or flow cytometry sorting. However, the number of cells that can be recovered by such techniques remains low and genetic material needs to be amplified before metagenomic sequencing can be undertaken. In this methodological study, we tested the application of whole-genome amplification (WGA) to photosynthetic eukaryotes. We performed various optimization steps both on a mixture of known microalgal strains and on natural photosynthetic eukaryote populations sorted by flow cytometry. rRNA genes were used as markers for assessing the efficiency of different protocols. Our data indicate that WGA is suitable for the amplification of photosynthetic eukaryote genomes, but that biases are induced, reducing the diversity of the initial population. Nonetheless, this approach appears to be suitable for obtaining metagenomics data on microbial eukaryotic communities.
authors.affiliation: UPMC (Paris-06) and CNRS, UMR 7144, Station Biologique de Roscoff, BP74, Roscoff, France. c.lepere@warwick.ac.uk
authors.forename: Cécile
authors.initials: C
authors.lastname: Lepere
authors.forename: Mikihide
authors.initials: M
authors.lastname: Demura
authors.forename: Masanobu
authors.initials: M
authors.lastname: Kawachi
authors.forename: Sarah
authors.initials: S
authors.lastname: Romac
authors.forename: Ian
authors.initials: I
authors.lastname: Probert
authors.forename: Daniel
authors.initials: D
authors.lastname: Vaulot
mesh_terms: D018744:DNA, Plant / Q000235:genetics; D005434:Flow Cytometry; D020459:Genes, rRNA; D056186:Metagenomics / Q000379:methods*; D058086:Microalgae / Q000235:genetics*; D021141:Nucleic Acid Amplification Techniques / Q000379:methods*; D010788:Photosynthesis; D010802:Phylogeny; D016133:Polymerase Chain Reaction; D015203:Reproducibility of Results; D012623:Seawater / Q000382:microbiology; D014871:Water Microbiology
chemical_list: D018744:DNA, Plant