The study aims at investigating the protist diversity in 17 aquatic locations in the region of the Vestfold hills, Eastern Antarctica (68°S, 78°E), including one marine site and 16 lakes, across a gradient of salinity spanning from 0 to 250 psu, and over the austral summer period (December 2008-February 2009). A total of 28 samples from the 17 locations were extracted using a CTAB method and amplified in two consecutive runs (HA6YNFP01.sff and HA6YNFP02.sff) using 14 MIDs each, over the v4 region of the 18S rRNA gene, and using the 454 pyrosequencing high-throughput sequencing method. The lakes comprise Lake Abraxas, Ace Lake, Crooked Lake, Deep Lake, Ekho Lake, Lake Hand, Highway Lake, Lebed Lake, Lake McNeil, Organic Lake, Pendant Lake, Rookery Lake, Lake Shield, Vereteno Lake, Lake Watts, and Lake Williams. Information on the tag annotation indicates respectively the run number (R1: run1 and R2: run2) and the 454-MID used (ALxx). Using the inferred diversity of protist in the Vestfold Hills region we further investigated the impact of environmental and historical factors on community structure (Logares et al 201x).