IMIS - Marine Onderzoeksgroepen

Microbial mat samples were collected in the littoral or deeper parts of the euphotic zone of the lakes using a spatula or gravity corer, respectively. The upper 1 cm of the core was aseptically removed and kept dark and cool until transfer to -20°C.
Study Extent: Samples were taken from benthic microbial mats (upper 1 cm), collected in 233 lakes covering 17.35° latitude in the Northern Hemisphere (61.39°N to 78.74°N) and 37.16° in the Southern Hemisphere (46.84°S to 84°S), including the major biogeographical regions in Antarctica, Sub-Antarctica and the Arctic.
Quality Control

1. To assess overall sequence quality and estimate the parameters for bioinformatics processing, each run also included two replicates of positive control sample (mock community) containing bacterial or eukaryote taxa on the respective bacterial and eukaryote runs. Method step description: Extracellular proteins and DNA from 1.5-3 g subsamples were removed (Corinaldesi et al., 2005), after which genomic DNA was extracted using a phenol-chloroform based protocol (Zwart et al., 1998). For Bacteria, the V1-V3 region of the 16S SSU rRNA gene was targeted for bacteria with domain-specific universal primer sets (Edwards et al., 1989; Cleenwerck et al., 2007), while for eukaryotes, V4 region of the 18S ssu rRNA gene was targeted with the primers of Stoeck et al. (2010). . The polymerase chain reaction was performed in duplicate to level out stochastic artefacts, and amplicon libraries were barcoded using the NEXTERA XT index kit (Illumina Inc.) according to manufacturer's instructions. Libraries were sequenced on an Illumina MiSeq machine (300 bp, Paired-end), while each run was spiked with 20% PhiX DNA (Illumina Inc.) to reduce cluster density.