Nextera XT DNA Library Preparation

Filters and cells possibly detached from the filters were subjected to three successive rinsing/centrifugation steps using a phosphate buffer solution (PBS) to remove the RNAlater reagent. Total nucleic acids were extracted using the Power Water RNA isolation kit (Machery-Nagel), following manufacturer instructions, except that the DNAse treatment step was omitted (DNA and RNA were co-extracted but only the DNA fraction was further analyzed in the present study). Libraries of total genomic DNA were prepared using Nextera XT preparation kit (Illumina) following the manufacturer’s instructions, except that we increased the number of PCR amplifications to 13 cycles as indicated for low DNA input (0.1 ng/µl in our case)^{53 (link)}. Libraries were purified using AMPure XP beads (New England Biolabs) and their concentration was normalized following Illumina Nextera XT protocol. Bioanalyzer and High Sensitivity DNA Kit (Agilent) were used to check the fragment size distribution of each library. Libraries were sequenced on an Illumina Miseq sequencer based at the Environmental Microbial Genomics group in Laboratoire Ampère (Ecole Centrale de Lyon) with 2 × 250 bp chemistry (MiSeq Reagent Kit v2).