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Restriction-site-associated genomic markers are a powerful tool for investigating evolutionary questions at the population level, but are limited in their utility at deeper phylogenetic scales where fewer orthologous loci are typically recovered across disparate taxa. While this limitation stems in part from mutations to restriction recognition sites that disrupt data generation, an additional source of data loss comes from the failure to identify homology during bioinformatic analyses. Clustering methods that allow for lower similarity thresholds and the inclusion of indel variation will perform better at assembling RADseq loci at the phylogenetic scale.

PyRAD: assembly of de novo RADseq loci for phylogenetic analyses