Here we present physico-chemical data collected during two research cruises conducted to and across the Ross Sea, Antarctica in the summer of 2018 (February-March) and 2019 (January-February). The dataset includes measurements of temperature, salinity, oxygen, par and transmissivity obtained with a Sea-Bird Electronics (SBE) 911plus CTD. The CTD sensor was configured with SBE 3plus, SBE 4, and SBE 43 dual sensors for the parameters above, in addition to a seapoint fluorescence sensor, and a photosynthetically active radiation (PAR) sensor (Biospherical Instruments QCP‐2300L‐HP). These data were used to provide oceanographic context to DNA metabarcoding analysis of 18S rRNA V4 region that was carried out on DNA samples collected in parallel to nutrient and chlorophyll-a samples. Fastq samples from DNA metabarcoding analysis and the associated metadata (including nutrients, Chlorophyll-a, and size-fractionated chlorophyll-a) were deposited to GenBank under project numbers PRJNA756172 (2018 cruise) and PRJNA974160 (2019 cruise). The study resulting from this analysis has been submitted to Limnology and Oceanography.