Laboratory protocols for nuclear genomic markers
We extracted genomic DNA from fresh tissue samples using DNeasy tissue extraction kit (Qiagen, CA, USA). To generate the genomic data we used double-digest restriction site-associated DNA sequencing (ddRADseq) following the protocol of Peterson et al. (2012) with modifications described in Thrasher et al. (2018). Briefly, for each sample we isolated ~500 ng of DNA, at a standardized concentration of 20 ng/µl, and digested it with the restriction enzymes SbfI High Fidelity (8 base bp recognition site; 5’-CCTGCAGG-3’) and MspI (4 base bp recognition site; 5’-CCGG-3’) (New England BioLabs, MA, USA). The digested DNA was ligated to P1 and P2 adapters on both ends (sequences are available in Peterson et al. 2012). We size-selected groups of 20 uniquely barcoded samples (index groups), retaining fragments between 400 and 700 bp using Blue Pippin (Sage Science, MA, USA). To incorporate the full Illumina TruSeq primer sequences and unique indexing primers into each library, we performed low cycle number PCR with Phusion High-Fidelity DNA Polymerase (New England BioLabs), with the following thermocycling profile: 98°C for 30 sec followed by 11 cycles at 98°C for 5 sec, 60°C for 25 sec, and 72°C for 10 sec with a final extension at 72°C for 5 min. We visualized the product of this amplification on a 1% agarose gel and performed a final 0.73 AMPure cleanup to eliminate DNA fragments smaller than 200 bp. Finally, sequencing was performed on an Illumina HiSeq 2500 lane at the Cornell University Biotechnology Resource Center as part of a larger sequencing batch, obtaining single-end 150-bp sequences, with an average mean sequencing depth of 151 reads per locus per individual (range: 104-220).