title: Fungal ITS2, isolated from soil
design: DNA was extracted from 0.25 g of sieved soil using the DNeasy PowerSoil Kit (Qiagen, Hilden, Germany). DNA concentrations were determined using a NanoDrop 2000 Micro-Volume UV-Vis spectrophotometer (Thermo Scientific, Wilmington, Delaware, USA). DNA samples were diluted to 10 ng/uL prior to PCR amplification. Amplicons of the V4 region of the bacterial 16S rRNA gene were generated using 515f (5-GTGCCAGCMGCCGCGGTAA-3; Caporaso et al., 2011) and 806r (5'-GGACTACHVGGGTWTCTAAT-3'; Caporaso et al., 2011). Amplicons of the internal transcribed spacer between the 5.8S and 28S regions (ITS2) of the fungal rRNA gene were generated using fITS7 (5-GTGARTCATCGAATCTTTG-3; Ihrmark et al. 2012) and ITS4 (5'-TCCTCCGCTTATTGATATGC-3'; White et al. 1990). All amplicons were purified using the Agencourt AMpure XP system (Beckman Coulter, Lane Cove, NSW, Australia) and genomic libraries were prepared using the Nextera XT Index Kit (Illumina, San Diego, CA, USA). Paired-end (2 x 251 bases) sequencing was performed on the Illumina MiSeq platform.