worker of Cataglyphis niger
For six individuals from each of the 47 nests, DNA was extracted from the head and abdomen (QIAGEN AllPrep DNA/RNA Mini kit) and the thorax was used for the chemical analysis of CHCs (see below). A uniquely barcoded genomic library was constructed for each sample following the standard protocol of the Illumina TruSeq Nano DNA Library Prep kit. Briefly, a Covaris S220 sonicator was used for DNA shearing aiming for 550bp inserts, ends were repaired, and the fragments were size-selected and purified using magnetic beads (supplied in kit). Then, an adenosine was added to the 3 end of the blunt fragments, individually barcoded adaptors were ligated, and ligated fragments were amplified (eight PCR cycles). Libraries were pooled at equal quantities as three separate pools, each of which was sequenced in two lanes of a paired-end, 150bp reads, on a HiSeq X Ten Illumina sequencer (six lanes in total).