radseq of Cladonia perforata: CordellNW_3
DNA Extraction and RAD Library Preparation & Sequencing  To assess genetic variation in C. perforata, up to eight thalli from each population were selected at random from sampled thalli for high-throughput sequencing. DNA extraction, RAD library preparation, and sequencing were performed by the Biotechnology Center Next Gen Sequencing facility at the University of Wisconsin (https://biotech.wisc.edu/dna-seq/). DNA was extracted using the DNeasy mericon Food Kit (Qiagen, Hilden, Germany), and DNA concentration was verified using the Quant-iTTM PicoGreen dsDNA Kit (Life Technologies, Grand Island, NY, USA).  RAD libraries were prepared following Elshire et al. (2011), with minor modifications. Specifically, 50 ng of DNA was digested using the 5-bp cutter ApeKI (New England Biolabs, Ipswich, MA, USA), barcoded adapters were added by ligation with T4 ligase (New England Biolabs, Ipswich, MA, USA), and samples were then pooled and amplified to generate library quantities suitable for sequencing. Adapter dimers were removed by SPRI bead purification. The quality and quantity of libraries were assessed using the Agilent Bioanalyzer High Sensitivity Chip (Agilent Technologies, Inc., Santa Clara, CA, USA) and the Qubit dsDNA HS Assay Kit (Life Technologies, Grand Island, NY, USA), respectively. RAD libraries were sequenced on an Illumina NovaSeq 6000 platform (Illumina Inc., San Diego, CA, USA) at one million reads per sample. Processed data were returned in the FASTQ format, with Phred quality scores for all bases.