Background Plankton seascape genomics studies have revealed different trends from large-scale weak differentiation to microscale structures. Previous studies have underlined the influence of the environment and seascape on species differentiation and adaptation. However, these studies have generally focused on a few single species, sparse molecular markers, or local scales. Here, we investigated the genomic differentiation of plankton at the macro-scale in a holistic approach using Tara Oceans metagenomic data together with a reference-free computational method. Results We reconstructed the F ST -based genomic differentiation of 113 marine planktonic taxa occurring in the North and South Atlantic Oceans, Southern Ocean, and Mediterranean Sea. These taxa belong to various taxonomic clades spanning Metazoa, Chromista, Chlorophyta, Bacteria, and viruses. Globally, population genetic connectivity was significantly higher within oceanic basins and lower in bacteria and unicellular eukaryotes than in zooplankton. Using mixed linear models, we tested six abiotic factors influencing connectivity, including Lagrangian travel time, as proxies of oceanic current effects. We found that oceanic currents were the main population genetic connectivity drivers, together with temperature and salinity. Finally, we classified the 113 taxa into parameter-driven groups and showed that plankton taxa belonging to the same taxonomic rank such as phylum, class or order presented genomic differentiation driven by different environmental factors. Conclusion Our results validate the isolation-by-current hypothesis for a non-negligible proportion of taxa and highlight the role of other physicochemical parameters in large-scale plankton genetic connectivity. The reference-free approach used in this study offers a new systematic framework to analyse the population genomics of non-model and undocumented marine organisms from a large-scale and holistic point of view.