Sample dissection and DNA extraction in the lab
Feather dissections were performed by taking the cells from the basal tip of the calamus, following Morin et al. (1994). The lysis was performed by adding 400 µl lysis buffer and 40 µl protease K, and incubating samples at 56ºC overnight. DNA was extracted using an automated Chemagic 360 instrument (Perkin Elmer) from a total of 957 samples corresponding to 905 feathers from fledglings, and 49 feathers and 3 tissue samples from adult individuals. A duplicated sample collected from a recaptured individual was included as a genotyping control. The quality and concentration of the extracted DNA was assessed using a NanoDrop spectrophotometer (Thermo Fisher Scientific). Laboratory procedures were performed at the SVGM laboratories (Molecular Genetics Veterinary Service) of the Faculty of Veterinary of the Autonomous University of Barcelona. Some feather samples, particularly those belonging to adults, had low DNA concentrations (below 50 ng/μL, which is the recommended DNA concentration to ensure proper sample genotyping according to the Thermo Fisher Scientific protocol for QuantStudio™ OpenArray® PCR Plates). For such low-quality samples (N=90), liquid was evaporated using a SpeedVac Vaccum Concentrator centrifuge to increase DNA concentrations. This process was carried out at the Centre for Research in Agricultural Genomics (CRAG) facilities. Further details on sample quality, applied thresholds and genotyping success related with DNA concentration are shown in Section S1 in Supplementary Information (see also Table S1, Fig. S2 and Fig. S3).