DOI: 10.5061/dryad.pg4f4qrrq
title: Data from: Speciation with gene flow in an island endemic hummingbird

description: We find evidence for speciation in streamertail hummingbirds (Trochilus
 polytmus and T. scitulus), Jamaican endemic taxa thought to represent an
 exception to the rule that, for birds, speciation cannot progress in situ
 on small islands. Our analysis shows that divergent selection acting on
 male bill color, a sexual ornament that is red in polytmus and black in
 scitulus, plays a pivotal role as a reproductive barrier. We sequenced
 6,451 single nucleotide polymorphisms (SNPs) and the mitochondrial control
 region. Low genomic divergence was detected across the two datasets,
 consistent with a demographic history of recent speciation or extensive
 gene flow between previously diverged taxa. Such low background divergence
 offers little support for the presence of post-mating reproductive
 incompatibilities. Yet, a geographic cline analysis reveals narrow clines
 in a handful of traits likely to be under selection. In particular, the
 cline width for male bill color is only 2.3 km, marking it as one of the
 narrowest phenotypic clines observed in an avian hybrid zone. Notably,
 cline centers for individual genomic and phenotypic traits converged in
 the Rio Grande Valley, suggesting that this landscape feature played a
 role in stabilizing the hybrid zone’s position despite the fact that it
 does not impose a physical dispersal barrier to these highly volant
 hummingbirds. Pre-mating selection for bill color is the most likely
 driver of divergence in Jamaican streamertails.  
description: <strong>Laboratory Methods and Bioinformatics
 Processing: </strong>
 <em>Genotyping-by-sequencing</em>
 SNPs were generated using a genotyping-by-sequencing method by
 the Cornell Institute of Genomic Diversity (IGD). Briefly, DNA extracts
 for 160 <em>Trochilus</em> individuals was digested with PstI
 (CTGCAG), and the resulting fragments were ligated to sample-specific
 adapters and common adapters. Fragments were then pooled, cleaned, and
 amplified using an 18-cycle PCR. Final libraries were sequenced as
 single-end, 100-base pairs reads in one lane of an Illumina HiSeq 2000
 (Illumina, San Diego, CA, USA). SNP calling was
 performed using UNEAK, a reference-free SNP calling pipeline specific to
 GBS data that is an extension of the Java program TASSEL. UNEAK filters to
 remove reads that lack barcodes, cut sites, or that have ‘N’s present in
 the first part of the sequence following the barcode. High quality reads
 were then clustered into tags. Unique tags were merged and their counts in
 the whole sample of individuals were stored. Pairwise alignments of tags
 with a 1-bp mismatch were considered as candidate SNPs. Further filtering
 was performed to remove rare or singleton tags that likely result from
 sequencing error. We performed additional filtering steps using custom
 perl scripts to filter loci that are likely reverse complements of other
 loci in the dataset and to remove paralogous loci. The
 raw GBS reads were processed using the reference-free SNP calling pipeline
 UNEAK, an extension of the Java program ‘tassel’ (54). The UNEAK pipeline
 remove reads that lack barcodes, cut sites, or that have ‘N’s present in
 the first part of the sequence following the barcode. High quality reads
 were then clustered into tags. Unique tags in the pooled sample of
 individuals were compiled and counted. Pairwise alignments of tags with a
 1-bp mismatch were considered as candidate SNPs. Further filtering was
 performed to remove rare or singleton tags that likely result from
 sequencing error. Running the initial UNEAK pipeline
 yielded 97,432 SNPs. Filtering for missing data ( &gt; 20%) reduced
 the number of SNPs to 32,949. We then used custom perl scripts to filter
 potential reverse complements or loci that presumably contained paralogous
 reads, which reduced the total number of SNPs to 6,451 SNPs. Two
 individuals (635683, 636151) had ambiguous calls (‘N’s) for the majority
 of SNPs (&gt; 90%) and were removed from the dataset.  In addition, 14
 outliers in the multidimensional scaling and PCA plots were pruned because
 they were potentially biased by sequencing error: 633563, 633565, 633590,
 633591, 633600, 633603, 633623, 635683, 635691, 636149, 636157, 636159,
 636163, and 63617. The final dataset contains 6,451 high-quality SNPs in
 145 individuals. <em>MtCR
 haplotypes</em> Total DNA was extracted from
 approximately 25 mg of muscle tissue from 168 individuals using a DNeasy
 tissue kit (Qiagen, Valencia, California). A segment of the mitochondrial
 control region was amplified using a standard protocol for polymerase
 chain reaction (primers: H1251, 5'-TCTTGGCATCTTCAGTGCCRTGC-3'
 and TrochL-CR1: 5'-TGGCTAACGCGGAGCATACAATCTC-3'. Sequencing
 products were purified Sephadex and loaded on an ABI Prism 3100 Genetic
 Analyzer. Sequences were aligned in Sequencher 4.1 (Gene Codes, Ann Arbor,
 Michigan). When direct sequencing revealed more than one heterozygous site
 within a sequence, haplotypes were resolved probabilistically using
 PHASE.
description: # Data from: Speciation with gene flow in an island endemic hummingbird ##
 Authors of the Dataset: Caroline D. Judy, PhD, Department of Vertebrate
 Zoology, National Museum of Natural History, Smithsonian Institution
 [caroline.duffie@gmail.com](mailto:caroline.duffie@gmail.com) Gary R.
 Graves, John McCormack, Katherine Faust Stryjewski, Robb T. Brumfield
 \========================================== ## Description of the data
 files --- Three genomic data tables are archived in this repository (.csv,
 .fasta). --- ### museumID\_raw\.analysis.SNPs.csv A data matrix containing
 6,451 individual SNPs in 145 individuals. Variable names and descriptions:
 * Column1: MuseumID - a unique identifier associated with each vouchered
 specimen that can be cross referenced in Table S1. * Column2: phenotype -
 description of bill color based on morphological assessement Row1[,3:6453]
 labels for individual SNP IDs. ### SNP\_Loci.fasta A fasta file containing
 SNP loci. Variable names and descriptions: * Loci (rows) are labeled with
 their SNP IDs. ### mtcr\_museumID\_label.fasta A fasta file containing
 sequenced haplotypes from the mitochondrial control region. Variable names
 and descriptions: * Individuals (rows) are labeled with a MuseumID, a
 unique identifier associated with each vouchered. * specimen that can be
 cross referenced in Table S1.