title: ITS: Alterations to grassland soil nutrient supply and plant consumer manipulations jointly influence fungal necromass decay
design: We analyzed microbial communities in soil and on necromass collected at 6- and 32-days post-burial, representing early and intermediate decay stages. For these molecular-based analyses, total genomic DNA was extracted using a Qiagen DNeasy PowerSoil Kit (Qiagen, Benelux BV), including a 30-second bead beating step with 2-mm sterile glass beads (BioSpec Products, Bartlesville, OK, USA) to homogenize the necromass before cell lysis. To assess the composition of the fungal decomposer communities, ITS (fungal) DNA amplicons were generated at the University of Minnesota Genomics Center (UMGC, Minneapolis, MN). ITS2 amplicons were generated using a dual-index technique described in Gohl et al. (2016), with fungal-specific primers targeting the ITS2 region (forward primer 5.8SR, TCGATGAAGAACGCAGCG and the reverse primer ITS4, TCCTCCGCTTATTGATATGC). For PCR, 25 cycles were used for amplicon generation. Samples were sequenced on a full NextSeq run (2  300 bp Illumina chemistry).