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Abstract

Biology

Single Cell Micro-aspiration as an Alternative Strategy to Fluorescence-activated Cell Sorting for Giant Virus Mixture Separation

Published: October 27th, 2019

DOI:

10.3791/60148

1Institut Hospitalo-Universitaire (IHU) - Méditerranée Infection, 2Microbes, Evolution, Phylogeny and Infection (MEΦI), Aix-Marseille Université UM63, Institut de Recherche pour le Développement IRD 198, Assistance Publique - Hôpitaux de Marseille (AP-HM)

Abstract

During the amoeba co-culture process, more than one virus may be isolated in a single well. We previously solved this issue by end point dilution and/or fluorescence activated cell sorting (FACS) applied to the viral population. However, when the viruses in the mixture have similar morphologic properties and one of the viruses multiplies slowly, the presence of two viruses is discovered at the stage of genome assembly and the viruses cannot be separated for further characterization. To solve this problem, we developed a single cell micro-aspiration procedure that allows for separation and cloning of highly similar viruses. In the present work, we present how this alternative strategy allowed us to separate the small viral subpopulations of Clandestinovirus ST1 and Usurpativirus LCD7, giant viruses that grow slowly and do not lead to amoebal lysis compared to the lytic and fast-growing Faustovirus. Purity control was assessed by specific gene amplification and viruses were produced for further characterization.

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Keywords Single Cell Micro aspiration

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