Protists are extremely diverse and they play key roles in food webs and biogeochemical processes in aquatic ecosystems. Characterizing the biodiversity of protists has become in recent years a major goal in the ecological studies of protists. This project aims at discovering the overall diversity and main ecological structure of picoplanktonic eukaryotes in the Gulf of gabès and investigating their relationships to environmental factors. In order to achieve this objective, 12 samples were selected from the coastal and the open-sea areas in the Gulf of Gabès, at different depths and from different cruises (June 2008, April 2009 and November 2009)to account for the whole area and were processed to high-throughput sequencing. The eukaryotic V4 region of the 18S rRNA (about 400 bp) was PCR amplified from environmental DNA using the forward primer V4F and the reverse primer V4R (Stoeck et al. 2010. PCR reactions and Illumina MiSeq sequencing were performed at the Research and Testing Laboratories (RTL, Lubbock, TX, USA).