IMIS - Marine Onderzoeksgroepen

Samples (1 g) were taken aseptically, and were preserved using the LifeGuard™ Soil Preservation Solution (MO BIO Laboratories, Carlsbad, CA, USA).
Study Extent: Soil crust samples were collected during expeditions to the Arctic and Antarctica in August 2014 and February 2015, respectively. Arctic samples were taken near the station Ny-Ålesund. The Antarctic samples were collected close by the Spanish Juan Carlos I Antarctic Base at Livingston Island, South Shetland Islands.
Method step description:

1. The Arctic samples (NA) were processed according to Rippin et al. (2016) using the CTAB protocol, DNase I (Thermo Fisher Scientific, Waltham, MA, USA) and the RNeasy MinElute Cleanup Kit (Qiagen, Hilden, Germany). Due to low RNA yields, a total of six biological replicates were extracted and combined to obtain three pooled replicates. RNA from three biological replicates, collected at Livingston Island (Gr1), was isolated using the Spectrum™ Plant Total RNA Kit (Sigma-Aldrich, St. Louis, MO, USA), treated with DNase I (Thermo Fisher Scientific, Waltham, MA, USA) and purified using the RNeasy MinElute Cleanup Kit (Qiagen, Hilden, Germany) as described by Rippin et al. (2016). Single samples yielded sufficient amounts of RNA.
2. All RNA samples were further processed by Eurofins Genomics (Ebersberg, Germany). The processing included quality control utilizing the BioAnalyzer (Agilent Technologies, Santa Clara, CA, USA) and library preparation for both triplicates. Eukaryotic mRNA was enriched using oligo-(dT) beads, fragmented and, subsequently, cDNA was synthesized using random hexamers. Finally, Illumina compatible adapters were ligated. The libraries of the individual samples NÅ and Gr1 were applied to an Illumina HiSeq 2500, all triplicates multiplexed on one lane, using 125 bp paired-end and single-end mode, respectively. For sample NÅ, the HiSeq Control Software 2.2.58, RTA 1.18.64 and bcl2fastq-1.8.4 were used while the detected signals from sample Gr1 were processed by operating the HiSeq Control Software 2.2.38, RTA 1.18.61 and bcl2fastq-1.8.4 (Illumina, San Diego, CA, USA).