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<p>We derived inbred lines from the Raleigh, USA population by 20 generations of full-sib mating. We used the C(2L)RM-P1, b1; C(2R)RM-SKIA, cn1bw1 compound autosome (CA) stock for fitness assays. P-element mutations and co-isogenic control lines were a gift of H. Bellen (Howard Hughes Medical Institute, Baylor College of Medicine). We reared flies on cornmeal-molasses-agar medium at 25 °C, 60–75% relative humidity and a 12-h light-dark cycle unless otherwise specified.</p>
<p>Organismal phenotypes.</p>
<p>For the starvation stress resistance group, we placed ten same-sex, 2-d-old flies in vials containing 1.5% agar and 5 ml water, and scored survival every eight hours (N = 5 vials/sex/line). For the chill coma recovery group, we placed 3- to 7-d-old flies in empty vials on ice for three hours, and recorded the time for each individual to right itself after transfer to room temperature (N = 20 flies/sex/line). For longevity, we placed five 1- to 2-d-old same-sex virgin flies into vials containing 5 ml medium, and recorded survival every two days (N = 5 vials/sex/line). For locomotor reactivity, we placed single 3- to 7-d-old flies into vials containing 5 ml medium. The following day, between 8 am and 12 pm, we mechanically disturbed each fly19, and recorded the total activity in the 45 s immediately following the disturbance. We obtained two replicate measurements of 20 flies/sex/replicate/line. For the copulation latency group, we aspirated pairs of 3- to 7-d-old virgin flies into vials containing 5 ml medium between 8 am and 12 pm, and recorded the number of minutes until initiation of copulation, for a maximum of 120 min (N = 24 pairs/line). For the reproductive fitness group, we used the competitive index technique45, 46. We reared all wild-type and CA parents in constant density (10 pairs) vials. We placed six 3- to 4-d-old virgin CA males and females and three 3- to 4-d-old wild-type males and females in a vial containing 10 ml medium, discarding the flies after 7 d. The competitive index was the ratio of the number of wild type to the total number of progeny emerging by day 17 (N = 20 replicate vials/line).</p>
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