DOI: 10.5061/dryad.tb2rbp09k
title: Data from: Genetic and phenotypic differentiation in a Neotropical passerine with a disjunct distribution in the Andean and Atlantic forests (Thamnophilus ruficapillus)

description: The Andean and Atlantic forests are separated by the open vegetation
 corridor, which acts as a geographic barrier. However, these forests
 experienced cycles of connection and isolation in the past, which shaped
 the phylogeographic patterns of their biotas. We analysed the evolutionary
 history of the Rufous-capped Antshrike (Thamnophilus ruficapillus), a
 species with a disjunct distribution in the Atlantic and Andean forests
 and thus an appropriate model to study the effect of the open vegetation
 corridor and the Andes on the diversification of the Neotropical avifauna.
 We performed a phylogenetic/phylogeographic analysis, including the five
 subspecies, using mitochondrial and nuclear genomic DNA, and studied their
 differences in vocalizations and plumage coloration. Both the
 mitochondrial and nuclear DNA evidenced a marked phylogeographic structure
 with three differentiated lineages that diverged without signs of gene
 flow in the Pleistocene (1.0–1.7 Mya): one in the Atlantic Forest and two
 in the Andean forest. However, the two Andean lineages do not coincide
 with the two disjunct areas of distribution of the species in the Andes.
 Vocalizations were significantly different between most subspecies, but
 their pattern of differentiation was discordant with that of the nuclear
 and mitochondrial DNA. In fact, we did not find song differentiation
 between the subspecies of the Atlantic Forest and that of the northwestern
 Bolivian Andes, even though they differ genetically and belong to
 different lineages. Consistently, no differences were found in plumage
 coloration between the subspecies of the Atlantic Forest and that of the
 southern Andes. Our results suggest a complex evolutionary history in this
 species, which differentiated both due to dispersion across the open
 vegetation corridor, likely during a period of connection between the
 Andean and Atlantic forests, and the effect of the Bolivian Altiplano as a
 geographic barrier. In both cases, Pleistocene climatic oscillations
 appear to have influenced the species diversification.
description: We extracted genomic DNA from fresh tissue samples using DNeasy
 tissue extraction kit (Qiagen, CA, USA). To generate the genomic data we
 used double-digest restriction site-associated DNA sequencing (ddRADseq)
 following the protocol of Peterson et al. (2012) with modifications
 described in Thrasher et al. (2018). Briefly, for each sample we isolated
 ~500 ng of DNA, at a standardized concentration of 20 ng/µl, and digested
 it with the restriction enzymes SbfI High Fidelity (8 base bp recognition
 site; 5’-CCTGCAGG-3’) and MspI (4 base bp recognition site; 5’-CCGG-3’)
 (New England BioLabs, MA, USA). The digested DNA was ligated to P1 and P2
 adapters on both ends (sequences are available in Peterson et al. 2012).
 We size-selected groups of 20 uniquely barcoded samples (index groups),
 retaining fragments between 400 and 700 bp using Blue Pippin (Sage
 Science, MA, USA). To incorporate the full Illumina TruSeq primer
 sequences and unique indexing primers into each library, we performed low
 cycle number PCR with Phusion High-Fidelity DNA Polymerase (New England
 BioLabs), with the following thermocycling profile: 98°C for 30 sec
 followed by 11 cycles at 98°C for 5 sec, 60°C for 25 sec, and 72°C for 10
 sec with a final extension at 72°C for 5 min. We visualized the product of
 this amplification on a 1% agarose gel and performed a final 0.73 AMPure
 cleanup to eliminate DNA fragments smaller than 200 bp. Finally,
 sequencing was performed on an Illumina HiSeq 2500 lane at the Cornell
 University Biotechnology Resource Center as part of a larger sequencing
 batch, obtaining single-end 150-bp sequences, with an average mean
 sequencing depth of 151 reads per locus per individual (range:
 104–220). We demultiplexed, trimmed, filtered reads,
 assembled loci and called single nucleotide polymorphism (SNPs) with
 Ipyrad 0.7.28 (Eaton and Overcast 2020). We discarded sequences when a
 single base had a Phred quality score below 10 or more than 5% of bases
 had a Phred quality score below 20. Additional filtering was applied to
 only retain reads that did not have enzyme cleavage sites, adaptor
 sequences or index sequences. We used the default settings within Ipyrad
 to assemble loci and call SNPs: cluster threshold was set to 0.85, maximum
 fraction of heterozygous bases allowed in consensus sequences was set to
 0.05, maximum number of SNPs allowed in a final locus was set to 0.2,
 maximum number of ambiguous sites was set to 0.05 and maximum number of
 shared polymorphic sites in a locus was set to 0.5. Finally, we only kept
 loci that were present in at least 80% of the samples.
description: Thamnophilus ruficapillus ddRAD data \--- LOCI file containing the ddRAD
 data of 18 samples of the Rufous-capped Antshrike (*Thamnophilus
 ruficapillus*) collected in Bolivia, Argentina, and Uruguay. Title of the
 manuscript: Genetic and phenotypic differentiation in a Neotropical
 passerine with a disjunct distribution in the Andean and Atlantic forests
 (Thamnophilus ruficapillus). Authors: Belén Bukowski, Leonardo Campagna,
 Gustavo S. Cabanne, Pablo L. Tubaro and Darío A. Lijtmaer.