Methodology for Single-Cell Genetic Analysis of Planktonic Foraminifera for Studies of Protist Diversity and Evolution

• Published in: Frontiers in Marine Science
• Main Authors: Weiner, Agnès K.M., Morard, Raphael, Weinkauf, Manuel, Darling, Kate F., Andre, Aurore, Quillévéré, Frédéric, Ujiié, Yurika, Douady, Christophe Jean, de Vargas, Colomban, Kucera, Michal
• Other Authors: Japan Agency for Marine-Earth Science and Technology (JAMSTEC), Center for Marine Environmental Sciences Bremen (MARM), University of Bremen, Université de Genève = University of Geneva (UNIGE), School of Geosciences Edinburgh, University of Edinburgh (Edin.), School of Geography and Geosciences St Andrews, University of St Andrews Scotland, Université de Reims Champagne-Ardenne (URCA), Laboratoire de Géologie de Lyon - Terre, Planètes, Environnement (LGL-TPE), École normale supérieure de Lyon (ENS de Lyon)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Institut national des sciences de l'Univers (INSU - CNRS)-Université Jean Monnet - Saint-Étienne (UJM)-Centre National de la Recherche Scientifique (CNRS), Center for Advanced Marine Core Research, Kochi University, Laboratoire d'Ecologie des Hydrosystèmes Naturels et Anthropisés (LEHNA), Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-École Nationale des Travaux Publics de l'État (ENTPE)-Centre National de la Recherche Scientifique (CNRS), Institut universitaire de France (IUF), Ministère de l'Education nationale, de l’Enseignement supérieur et de la Recherche (M.E.N.E.S.R.), Evolution des Protistes et Ecosystèmes Pélagiques (EPEP), Adaptation et diversité en milieu marin (AD2M), Station biologique de Roscoff Roscoff (SBR), Université Pierre et Marie Curie - Paris 6 (UPMC)-Centre National de la Recherche Scientifique (CNRS)-Université Pierre et Marie Curie - Paris 6 (UPMC)-Centre National de la Recherche Scientifique (CNRS)-Centre National de la Recherche Scientifique (CNRS)-Station biologique de Roscoff Roscoff (SBR), Université Pierre et Marie Curie - Paris 6 (UPMC)-Centre National de la Recherche Scientifique (CNRS)-Université Pierre et Marie Curie - Paris 6 (UPMC)-Centre National de la Recherche Scientifique (CNRS)-Centre National de la Recherche Scientifique (CNRS), ANR-11-BTBR-0008,OCEANOMICS,Biotechnologies et bioressources pour la valorisation des écosystèmes marins planctoniques(2011)
• Format: Article in Journal/Newspaper
• Language: English
• Published: HAL CCSD 2016
• Subjects: methods; cryptic diversity; comparability; foraminifera; protists; single-cell genetics; laboratory protocols; standardization; [SDE.BE]Environmental Sciences/Biodiversity and Ecology; Planktonic foraminifera
• Online Access: https://sde.hal.science/hal-01420794; https://sde.hal.science/hal-01420794/document; https://sde.hal.science/hal-01420794/file/fmars-03-00255.pdf; https://doi.org/10.3389/fmars.2016.00255

International audience Single-cell genetic analysis is an essential method to investigate the biodiversity and evolutionary ecology of marine protists. In protist groups that do not reproduce under laboratory conditions, this approach provides the only means to directly associate molecular sequences with cell morphology. The resulting unambiguous taxonomic identification of the DNA sequences is a prerequisite for barcoding and analyses of environmental metagenomic data. Extensive single-cell genetic studies have been carried out on planktonic foraminifera over the past 20 years to elucidate their phylogeny, cryptic diversity, biogeography, and the relationship between genetic and morphological variability. In the course of these investigations, it has become evident that genetic analysis at the individual specimen level is confronted by innumerable challenges ranging from the negligible amount of DNA present in the single cell to the substantial amount of DNA contamination introduced by endosymbionts or food particles. Consequently, a range of methods has been developed and applied throughout the years for the genetic analysis of planktonic foraminifera in order to enhance DNA amplification success rates. Yet, the description of these methods in the literature rarely occurred with equivalent levels of detail and the different approaches have never been compared in terms of their efficiency and reproducibility. Here, aiming at a standardization of methods, we provide a comprehensive review of all methods that have been employed for the single-cell genetic analysis of planktonic foraminifera. We compile data on success rates of DNA amplification and use these to evaluate the effects of key parameters associated with the methods of sample collection, storage and extraction of single-cell DNA. We show that the chosen methods influence the success rates of single-cell genetic studies, but the differences between them are not sufficient to hinder comparisons between studies carried out by different methods. The ...