Illumina Seq of Zooplankton: VCDT3
Samples were prepared for Illumina sequencing by COI gene and 18S-V4 gene amplification of the eukaryotic community. The DNA was amplified for each one of the hypervariable regions with specific primers and further reamplified in a limited-cycle PCR reaction to add sequencing adapters and dual indexes. Second PCR reactions added indexes and sequencing adapters to both ends of the amplified target regions according to manufacturers recommendations (Illumina, 2013). PCR products were then one-step purified and normalized using SequalPrep 96-well plate kit (ThermoFisher Scientific, Waltham, USA) (Comeau, Douglas, & Langille, 2017), pooled and pair end sequenced in the Illumina MiSeq sequencer with the MiSeq reagent Kit v3 (600 cycles), according to manufacturers instructions (Illumina, San Diego, CA, USA) at Genoinseq (Cantanhede, Portugal).