RNA-seq of Southern Ocean and Ross Sea samples
Total RNA with was extracted from the filters using the NucleoMag RNA kit (Macherey-Nagel, Dren, Germany). rRNA was depleted using Ribo-Zero Magnetic kit (Illumina, La Jolla, USA) with a modified Removal Solution consisting of plant, bacterial, and human/mouse/rat solutions (2:1:1 ratio). cDNA was synthesized by the Ovation RNA-Seq System V2 (Tecan, Redwood City, USA), which was then fragmented to the target size of 400bp using the Covaris E210 focused ultrasonicator. Libraries were prepared using the Ovation Ultralow V2 system (Tecan) and purified by AMPure XP beads (Beckman Coulter Life Sciences, Brea, USA). Libraries were subjected to paired-end 2150 bp sequencing on a NovaSeq 6000 instrument (Illumina) to an average of 24 million reads per library.