2.2 NextRAD sequencing
Genomic DNA was converted into nextRAD genotyping-by-sequencing libraries (SNPsaurus, LLC, Eugene, OR, USA) as in Russello, Waterhouse, Etter, and Johnson, (2015). Genomic DNAs were first fragmented with Nextera DNA Flex reagent (Illumina, Inc., San Diego, CA, USA), which also ligates short adapter sequences to the ends of the fragments. The Nextera reaction was scaled at a 1/5th reaction, and 5 ng of genomic DNA was used due to the presence of inhibitors in the samples. Fragmented DNAs were then PCR amplified for 26 cycles at 73ÂșC, with one of the primers matching the adapter and extending 9 nucleotides into the genomic DNA with the selective sequence GTGTAGAGC. Thus, only fragments starting with a sequence that matched the selective sequence of the primer were efficiently amplified. NextRAD libraries were sequenced in a 150 bp single-end reads mode in a lane of a HiSeq 4000 (SNPSaurus at University of Oregon). Raw sequence data are available at the National Centre for Biotechnology Information (NCBI), under the BioProject accession number PRJNA593002.