DOI: 10.5061/dryad.prr4xgxk1
title: Microsatellites genotyping for common and lesser cuckoos

description: In a parasite species, the dispersal of individuals should be dependent on
 host species to which they are specialized; thus, any sexual/individual
 difference in host specificity may influence their dispersal patterns, and
 hence, population genetic structures. However, such predictions remain
 poorly verified in generalist avian brood parasites that is composed of
 multiple lineages of host-specific races. Here, we show the dispersal
 consequences inferred from spatial genetic structures and their
 association with host specificity in brood parasitic common cuckoos
 Cuculus canorus in which female-specific host race formation has been
 widely accepted. Genetic sampling from adult cuckoos confirmed restricted
 dispersal in both sexes and resultant genetic structures between
 populations where different host species breed allopatrically, whereas it
 was not the case between distant areas inhabited by the same host species.
 Contrary to the female host race hypothesis, our results demonstrate that
 male cuckoos may also have host specificity and disperse accordingly,
 conclusively allowing us to hypothesize the formation of host race
 including both sexes.
description: Using mist nets with song playbacks and dummy cuckoos, we
 captured common cuckoos in MA and JJ during the breeding season (May-July)
 in 2013 and 2014, respectively, and in YP in 2018 (Fig. 1). For lesser
 cuckoos which only commonly occur in the southern part of Korea, the
 catching was conducted in MA and JJ in 2013 and 2014, respectively. We
 captured in total 110 common cuckoos and 74 lesser cuckoos: 25 (6 females
 and 19 males), 46 (6 females and 40 males) and 39 common cuckoos (8
 females and 31 males) in MA, YP and JJ, respectively, and 21 (6 females
 and 15 males) and 53 lesser cuckoos (9 females and 44 males) in MA and JJ,
 respectively. Blood samples (20-30 µl) were collected from the brachial
 wing vein of these birds and maintained in 1.5ml tubes with absolute
 ethanol at room temperature until further processing. Each bird was banded
 with a metal ring to prevent potential duplicate sampling. After sample
 collection, the cuckoos were safely released at the place where they were
 captured. DNA was extracted from blood samples using
 the ammonium acetate method (Nicholls<i> </i>et
 al<i>.</i> 2000). For microsatellite genotyping, we initially
 attempted to characterize both species of cuckoos at 14 microsatellites
 (Ccμ 01, 02, 09, 13, 60, 88, 100, 108, 119, 137, Clu 02, 03, 05, Cba 08)
 that were employed in previous studies on the common cuckoo
 (Gibbs<i> </i>et al<i>.</i> 2000; Fossøy<i>
 </i>et al<i>.</i> 2011). The polymerase chain reaction
 (PCR) and processing of the PCR products were outsourced to SolGent co.,
 Ltd, Daejeon, Korea. PCR was conducted in an ABI 9700 thermal cycler
 (Applied Biosystems) according to the following conditions: 95 °C
 denaturation for 15 min, followed by 35 cycles of denaturing at 95 °C for
 20 s, annealing at 54.0 °C for 40 s and then 72 °C for 60 s, and final
 extension at 72 °C for 3 min. The PCR products were run in a GeneScan™ 500
 ROX™ and an ABI 3130XL DNA Analyzer (Applied Biosystems) and the data
 obtained were scored in ABI GeneMapper v. 4.0 (Applied Biosystems) by one
 person (the first author) to ensure consistency. For
 mtDNA analysis, a 411-basepair portion of the left-hand hypervariable
 region of the mtDNA control region in both common and lesser cuckoo was
 sequenced. We used two <i>Cuculus</i>-specific primers
 (CCRL1A, 5’-CAT GAT ACA TTA CAT GTA TGC CTG-3’ and CCRH1, 5’-CTG AAA TAG
 TAT GAA TGT ATC TGT G-3’) for PCR as described by Gibbs<i>
 </i>et al. (2000), which successfully amplified the target region of
 mtDNA in both species. PCR and sequencing of the PCR products were
 outsourced to SolGent co., Ltd, Daejeon, Korea. DNA amplification was
 performed using an ABI 9700 thermal cycler (Applied Biosystems) with the
 same condition described above. The purified PCR products were sequenced
 using a BigDye Terminator v3.1 cycle sequencing ready reaction Kit and an
 ABI 3730XL DNA Analyzer (Applied Biosystems). DNA was sequenced from both
 ends using the same primers as in PCR. Chromatograms were edited and
 assembled by Sequencher 4.9 (Gene Codes Corporation). The edited sequences
 were aligned with Clustral W (Thompson<i> </i>et
 al<i>.</i> 1994) and manually adjusted using MEGA 6.0
 (Tamura<i> </i>et al<i>.</i> 2013).
description: The data contains two files that were used in the associated
 manuscript.  1) The results of microsatellite gentyping
 for common cuckoos (11 markers) and lesser cuckoos (10 markers) according
 to sex and capture sites (CJ: common cuckoo captured in Jeju; CM: common
 cuckoo captured in Muan; CY: common cuckoo captured in Yangpyeong; PJ:
 lesser cuckoo captured in Jeju; PM: lesser cuckoo captured in
 Muan). 2) The frequency of mtDNA haplotypes according
 to capture sites in common cuckoos and lesser cuckoos. The exact sequences
 of the haplotypes can be found in the supplementary files of the
 associated manuscript.