Species determination in plankton net (20 µm) concentrates by DNA sequencing during the PHYCOB cruise in the Southwestern Black Sea in 2021@en

At each station two vertical net tows were taken from the water column using a 20 μm plankton net (438-030, Hydro-Bios, Kiel, Germany) for analyses of plankton and phycotoxins. The depth of the hauls was fixed to 0 – 30 m. The collected net tow concentrates were adjusted to 1 L with filtered seawater. 150 mL aliquots were filtered under gentle vacuum (< 0.2 bar) through 1 μm pore-size polycarbonate filters (Whatman, USA) for DNA analysis. DNA from the filters was immediately extracted by using 5% Chelex buffer (Tanabe et al. 2016). For detection of eukaryotic species, universal primers for 18S rRNA gene V7-V9 variable region (18S-V7F: TGGAGYGATHTGTCTGGTTDATTCCG and 18S-V9R: TCACCTACGGAWACCTTGTTACG; modified from Tanabe et al. 2016) were used. The procedures and techniques, applicable to the treatment of the obtained sequences, selection and taxonomic identification of operational taxonomic units (OTUs), were administered according to the workflow described in Dzhembekova et al. (2017). Taxonomic assignment was performed using BLAST against a sequence database downloaded from GenBank.