ddRAD-seq
Double-digest restriction site-associated DNA sequencing (ddRAD-seq) and bioinformatic processing for SNP discovery and genotyping was adapted from Peterson et al. (2012) following Thrasher et al. (2018). The DNA concentration of sample extracts was determined using the Qubit dsDNA BR Assay Kit (ThermoFisher Scientific). Samples were then diluted or concentrated to 5-25 ng/l in order to obtain a final mass of 100-500 ng of DNA in a volume of 20 l. Briefly, the samples were digested with the restriction enzymes Sbfl-HF (NEB, R3642L) and Mspl (NEB, R0106), and the resulting fragments ligated to P1 (with a unique barcode) and P2 (with an index group barcode) adapters to the 5 and 3 ends, respectively. We pooled all samples, each containing a unique combination of barcodes for posterior bioinformatic demultiplexing. The DNA libraries were size-selected by BluePippin (Sage Science) to retain fragments between 400 and 700 bp and amplified using nine PCR cycles with Phusion DNA Polymerase (NEB). Following clean-up with AMPure beads, index groups were pooled in equimolar ratios and sequenced on one lane of an Illumina HiSeq 2500 at the Cornell University Biotechnology Resource Center. obtaining single-end 101 bp reads.