Inferring Phylogenies from RAD Sequence Data

Abstract

Reduced-representation genome sequencing represents a new source of data for systematics, and its potential utility in interspecific phylogeny reconstruction has not yet been explored. One approach that seems especially promising is the use of inexpensive short-read technologies (e.g., Illumina, SOLiD) to sequence restriction-site associated DNA (RAD) – the regions of the genome that flank the recognition sites of restriction enzymes. In this study, we simulated the collection of RAD sequences from sequenced genomes of different taxa (Drosophila, mammals, and yeasts) and developed a proof-of-concept workflow to test whether informative data could be extracted and used to accurately reconstruct “known” phylogenies of species within each group. The workflow consists of three basic steps: first, sequences are clustered by similarity to estimate orthology; second, clusters are filtered by taxonomic coverage; and third, they are aligned and concatenated for “total evidence” phylogenetic analysis. We evaluated the performance of clustering and filtering parameters by comparing the resulting topologies with well-supported reference trees and we were able to identify conditions under which the reference tree was inferred with high support. For Drosophila, whole genome alignments allowed us to directly evaluate which parameters most consistently recovered orthologous sequences. For the parameter ranges explored, we recovered the best results at the low ends of sequence similarity and taxonomic representation of loci; these generated the largest supermatrices with the highest proportion of missing data. Applications of the method to mammals and yeasts were less successful, which we suggest may be due partly to their much deeper evolutionary divergence times compared to Drosophila (crown ages of approximately 100 and 300 versus 60 Mya, respectively). RAD sequences thus appear to hold promise for reconstructing phylogenetic relationships in younger clades in which sufficient numbers of orthologous restriction sites are retained across species.

Double‐digest RADseq loci using standard Illumina indexes improve deep and shallow phylogenetic resolution of Lophodermium, a widespread fungal endophyte of pine needles

“…Although clustering reads or loci by similarity is an imperfect solution to identifying homologous loci, our results agree with conclusions from Rubin et al. (2012), showing that clusters that may include a mixture of orthologous and paralogous loci still contain substantial phylogenetic signal that can produce correct topologies with moderate to high accuracy.…”

“…Trimming reads to shorter lengths or targeting shorter reads will likely minimize loss of clusters or loci due to length variation among and within samples of different species. However, using longer reads produces more informative sites that may compensate for having a reduced number of shared loci and can potentially resolve deeper divergences (e.g., >300 million years) better than shorter reads (Rubin et al., 2012). Longer reads can also be more helpful for downstream population genetic analyses, such as genome mapping, haplotype reconstruction, and linkage to adaptive loci.…”

“…nov .). Clustering thresholds between 55 and 70% sequence similarity are considered to reconstruct the most accurate phylogenetic topologies for recent species divergences (e.g., <60 million years) but is highly recommended to be tested at various clustering values (Rubin, Ree, & Moreau, 2012). As our reads are longer than typical ddRAD reads sequenced on the HiSeq, we tested clustering at 85% threshold (DaCosta & Sorenson, 2014; Heffelfinger et al., 2014) allowing 15 Ns as well as at a lower 70% threshold allowing 30 Ns.…”

Genomic signatures of population bottleneck and recovery in Northwest Atlantic pinnipeds

“…These studies, using both simulated and empirical data, have quantified the number of retained homologous loci among species with varying divergence dates. Simulations suggest that hundreds of orthologous, phylogenetically informative loci can be identified in species with divergence times of up to 60 million years (Cariou, Duret, & Charlat, 2016a,2016b; Rubin, Ree, & Moreau, 2012), but empirical studies have successfully applied RADseq to phylogenetic studies of species only from more recent radiations (<100,000 years; Keller et al., 2013; Wagner et al., 2013) and find that the number of shared SNPs drops, sometimes quickly, as divergence dates increase (Lexer et al., 2013; Pante et al., 2015). Among our two studied seal species, harbor and gray, that diverged approximately 5 mya (Arnason et al., 2006), we found over 100,000 homologous RAD loci.…”

Genetic relationships among Eriobotrya species revealed by genome‐wide RAD sequence data

“…RAD‐Seq is suitable for fine‐scale linkage mapping (Scaglione et al., 2015; Wang, Fang, Xin, Wang, & Li, 2012), population genetics (Hohenlohe et al., 2011; Andersen et al. 2012), phylogenetics, and phylogeography (Cruaud et al., 2014; Rubin, Ree, & Moreau, 2012; Takahashi & Moreno, 2015; Valdisser et al., 2016). RAD‐Seq has also been used to generate large SNP datasets for many plants (Torres‐Martínez & Emery, 2016; Wang, Jin, Zhang, Shen, & Lin, 2015).…”