Fly collection, DNA isolation and sequencing
We collected wild Drosophila at the four mountainous locations
across Arizona between the 22nd of August and the
11th of September 2017: the Southwest research station
in the Chiricahua mountains (CH, ~5,400 feet elevation,
31.871 latitude -109.237 longitude, 96 flies), in Prescott National
Forest (PR, ~7,900 feet elevation, 34.586 latitude
-112.559 longitude, 96 flies), Madera Canyon in the Santa Rita mountains
(SR, ~4,900 feet elevation, 31.729 latitude -110.881
longitude, 96 flies) and Miller Peak in the Huachuca mountains (HU,
~5,900 feet elevation, 31.632 latitude -110.340
longitude, 53 flies) (Coe et al. 2012). Baits consisted
of store-bought white button mushrooms (Agaricus bisporus ) placed
in large piles about 30cm in diameter, with at least 5 baits per
location. We used a sweep net to collect flies over the baits in either
the early morning or late afternoon between one and three days after the
bait was set. We sorted flies by sex and species at the University of
Arizona in Tucson, AZ and flash frozen at -80°C before shipping on dry
ice to the University of Kansas in Lawrence KS.
We sorted 343 flies (172 females and 171 males) which phenotypically
matched D. innubila. We then homogenized and extracted DNA using
the Qiagen Gentra Puregene Tissue kit (USA Qiagen Inc., Germantown, MD,
USA). We also prepared the DNA of 40 D. innubila collected in
2001 from CH. We prepared a genomic DNA library of these 383 DNA samples
using a modified version of the Nextera DNA library prep kit
(~ 350bp insert size) meant to conserve reagents. We
sequenced the libraries on four lanes of an Illumina HiSeq 4000 (150bp
paired end) (Supplementary Table 1, Data to be deposited in the NCBI
SRA).