title: 16S amplicon sea ice
design: Total genomic DNA was extracted directly from the polycarbonate filters (n = 54) in three batches (Supplementary Table 1) using the Power Soil Pro kit and following the instructions of the manufacturer (Qiagen). Quantitative Polymerase Chain Reaction (qPCR) using an MX3005p qPCR instrument (Agilent, Santa Clara, CA, United States) was performed to quantify the amount of bacterial 16S rRNA and eukaryotic 18S rRNA gene copies gene copies. For bacteria we targeted partial 16S rRNA gene sequences using universal primers Bac908F (5-AAC TCA AAK GAA TTG ACG GG-3) and Bac1075R (5-CAC GAG CTG ACG ACA RCC-3)48. For eukaryotic 18S rRNA sequences we used primers Euk345F (5- AAGGAAGGCAGCAGGCG-3) and Euk499R (5- CACCAGACTTGCCCTCYAAT-3)49.  The qPCR mixture contained 2 l template DNA, 1 l of each primer (10 pmol l-1), 2 l BSA (10 ng l-1), 10 l LightCycler Mastermix (Roche) and 4 l dH2O to a final volume of 20 l. Samples were run in triplicate including triplicate negative qPCR controls without addition of template DNA. The thermal conditions were as follows: For 16S quantification we used 95 C for 5 min, 45 cycles with denaturation at 95 C for 30 s, annealing at 55 C for 30 s, elongation at 72 C for 15 s, followed by fluorescent acquisition 80 C at 5 s. A melting curve was produced using 1 cycle at 95 C at 30 s, 60 C at 1 min and 95 C at 30 s. For the 18S quantification we used 95 C for 5 min, 40 cycles with denaturation at 95 C for 30 s, annealing at 60 C for 30 s, elongation at 72 C for 20 s, followed by fluorescent acquisition 79 C at 15 s. A melting curve was produced using 1 cycle at 95 C at 30 s, 55 C at 1 min and 95 C at 30 s. The V3 and V4 of the 16S rRNA gene was amplified with primers Bac341F (5-CCT ACG GGN GGC WGC AG-3) and Bac805R (5-GAC TAC HVG GGT ATC TAA TCC-3). The 16S rRNA gene amplification was performed according to a modified Illumina protocol. The PCR mixture contained 2 l template DNA, 12.5 l 2  KAPA HiFi HotStart polymerase (Kapa Biosystems, Inc., Wilmington, MA, United States), 0.2 M forward primer and 0.2 M reverse primer and dH2O to a final volume of 25 ul. The thermal cycling was run with an initial denaturation step at 95C for 3 min, 30 cycles with denaturation at 95C for 30 s, annealing at 55C for 30 s, elongation at 72C for 30 s and a final elongation at 72C for 5 min. The 18S rRNA library preparation of DNA was performed as follows: Primers TareukFWD1 (5- CCAGCASCYGCGGTAATTCC-3) and TAReukREV3 (5- ACTTTCGTTCTTGATYRA-3) were used to amplify the V4 RNA loop (Stoeck et al. 2010). The PCR mixture contained 4 l template DNA. The thermal cycling was run with touchdown PCR approach with an initial denaturation step at 95 C for 3 min, 10 cycles with denaturation at 95 C for 30 s, annealing at 57 C for 30 s, elongation at 72 C for 30 s, followed by 15 cycles with denaturation at 95 C for 30 s, annealing at 47 C for 30 s, elongation at 72 C and a final elongation step at 72 C for 5 min. The PCR products were cleaned using 30 l AMPure XP magnetic beads. The second round of PCR was run for 10 cycles to incorporate overhang adapters and was run with the same conditions as the previous PCR with an annealing temperature of 57 C. Products were cleaned, and the Nextera XT Index primers were incorporated in a third PCR reaction which was run for 8 cycles following the previous condition and an annealing temperature of 55 C. The PCR products were quantified using a Quant-iT dsDNA BR assay kit on a FLUOstar Omega fluorometric microplate reader (BMG LABTECH, Ortenberg, Germany), diluted and pooled together in equimolar ratios. The pool was quantified using the Quant-iT dsDNA BR assay kit on a Qubit fluorometer (Thermo Fisher Scientific, Waltham MA) and sequenced on the Illumina MiSeq platform (Illumina, San Diego, CA) which produces two 300-bp long paired-end reads.