DOI: 10.5061/dryad.sbcc2fr8f
title: Data from: Hybrid enrichment of adaptive variation revealed by genotype-environment associations in montane sedges

description: The role of hybridization in diversification is complex and may result in
 many possible outcomes. Not only can hybridization produce new lineages,
 but those lineages may contain unique combinations of adaptive genetic
 variation derived from parental taxa that allow hybrid-origin lineages to
 occupy unique environmental space relative to one (or both) parents. We
 document such a case of hybridization between two sedge
 species, Carex nova and Carex nelsonii (Cyperaceae), that occupy partially overlapping environmental space in the southern Rocky Mountains, USA. In the region hypothesized to be the origin of the hybrid lineage, one parental taxon (C. nelsonii) is at the edge of its environmental tolerance. Hybrid-origin individuals display mixed ancestry between the parental taxa – of nearly 7,000 unlinked loci sampled, almost 30% showed evidence of excess ancestry from one parental lineage – approximately half displayed a genomic background skewed towards one parent, and half skewed towards the other. To test whether excess ancestry loci may have conferred an adaptive advantage to the hybrid-origin lineage, we conducted genotype-environment association analyses on different combinations of loci – with and without excess ancestry – and with multiple contrasts between the hybrids and parental taxa. Loci with skewed ancestry showed significant environmental associations distinguishing the hybrid lineage from one parent (C. nelsonii), whereas loci with relatively equal representation of parental ancestries showed no such environmental associations. Moreover, the overwhelming majority of candidate adaptive loci with respect to environmental gradients also had excess ancestry from a parental lineage, implying these loci have facilitated the persistence of the hybrid lineage in an environment unsuitable to at least one parent.
description: <em>Field sampling and genomic
 characterization</em> Samples were
 field-collected from eight <em>C. nelsonii</em> localities,
 15 <em>C. nova</em> localities, and five putative hybrid sites
 (Fig. 1). Between two and ten individuals (mean = 8.5) separated by at
 least five meters were collected at each sampling locality (Supplemental
 Table S1, Supplemental Table S2). Putative hybrid-origin individuals
 collected and keyed out to <em>C. nelsonii </em>were only
 identified as hybrid-origin during the course of population genetic
 analyses. A total of 237 individuals – 54 <em>C.
 nelsonii</em>, 136 <em>C. nova</em>, and 47 putative
 hybrids – were used for genomic analyses (Fig. 1, Supplemental Table S1,
 Supplemental Table S2). We performed double-digest
 RAD-Seq (Peterson et al. 2012) to obtain anonymous genomic loci. We used
 two restriction enzymes, <em>Eco</em>RI
 and <em>Mse</em>I, to digest genomic DNA, and ligated Illumina
 adaptor sequences and unique 10 bp barcodes to restriction sites. The
 ligation products for each library were pooled and PCR-amplified for 12
 cycles, and 400-500 bp fragments were size selected using a Pippin Prep
 (Sage Science, Beverly, MA). All libraries were sequenced on single-end
 Illumina HiSeq 2500 runs at The Centre for Applied Genomics (Hospital for
 Sick Children, Toronto, Ontario). Raw sequencing reads were processed
 using ipyrad v9.42 (Eaton 2014, Eaton & Overcast 2020) in a single
 assembly that combined both species and the putative hybrids (Supplemental
 Table S2). The combined assembly approach facilitated the tracking of
 allele frequencies across all individuals and populations in downstream
 analyses. The ipyrad assembly was filtered using vcftools (Danacek et al.
 2011) to allow 40% missing data per site (--max-missing = 0.6) and require
 a minor allele count of two or greater (--mac = 2). The resulting data
 matrix consisted of 6,946 unlinked SNPs (i.e., one randomly selected SNP
 per RAD-Seq locus). Because we were interested in the effects of putative
 adaptive loci associated with hybridization and not loci associated with
 selection within one of the parental species, we used PCAdapt (Luu et al.
 2017) to remove <em>F<sub>ST</sub> </em>outliers
 identified within either <em>C. nelsonii </em>or <em>C.
 nova</em>; in total, we removed 206 loci, which left 6,740 unlinked
 SNPs that were used in most subsequent analyses, except where noted. To
 verify that
 removing <em>F<sub>ST</sub> </em>outliers did not
 bias our downstream results, we ran several analyses both with and
 without <em>F<sub>ST</sub> </em>outliers,
 including analyses of excess ancestry, all genetic diversity and
 differentiation statistics, and redundancy analyses, as noted
 below. A separate ipyrad assembly was performed to add
 an outgroup (<em>C. chalciolepis</em>) suitable for rooting
 the phylogeny of <em>C. nelsonii </em>+ <em>C.
 nova </em>+ hybrids and for use in downstream analyses. This
 assembly included the 237 individuals referenced above plus 152
 individuals of <em>C. chalciolepis</em> and leveraged the same
 filtering strategy as the first assembly. The resulting SNP data matrix
 consisted of 6,069 unlinked SNPs, which were used in the SVDQuartets and
 TREEMIX analyses. The authors of HyDe recommend including invariant sites
 in the genetic dataset, so we used all 69,373 sites from the assembly with
 the outgroup in the HyDe analysis.
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