title: Poultry Litter Ash Plot 2  Unbalanced 2019   Fungi ITS Library
design: Soil sampling was conducted 51 days after fertiliser application following cutting of grass silage (July 2019). Nine sample cores were taken to a depth of 10cm per plot and stored at -20C until ready for analysis. A total of 40 composite samples were obtained (8 treatments X 5 replicate plots). Defrosted samples were sieved through a 2mm mesh and homogenised. For Fungi analysis, 0.25g sub-samples were obtained. Total DNA was extracted from 0.25g soil sub-samples using Qiagen DNeasy PowerSoil Pro kit, as per manufacturers instructions. Total DNA quality and quantity was assessed by NanoDrop instrument and agarose gel electrophoeresis using 1% agarose gels, before outsourcing to a sequencing company.Fungil ITS V4-V5 region were sequenced using 515F and 907R  primer pairs, respectively, on Illumina paired-end platform