Low hybridization temperatures improve target capture success of invertebrate loci

Abstract

Target capture approaches are widely used in phylogenomic studies, yet few experimental comparisons of critical parameters, e.g. hybridization temperature, have been published. Even fewer studies have focused on invertebrates where bait-target divergences may be high. Most capture studies use a fixed hybridization temperature of 65°C to maximize the proportion of on-target data, but lower temperatures, which might improve locus recovery, are not commonly employed. We used fresh and degraded specimens of leaf-footed bugs and relatives (Hemiptera: Coreoidea) to investigate the effect of hybridization temperature on capture success of previously published ultraconserved elements (UCE) targeted by baits derived from divergent hemipteran genomes and other loci targeted by newly designed baits derived from less divergent coreoid transcriptomes. We found touchdown capture approaches with lower hybridization temperatures generally resulted in lower proportions of on-target reads and lower coverage but were associated with more assembled contigs and improved recovery of targeted UCE loci. Low hybridization temperatures were also associated with increased numbers of putative paralogs of targeted UCE loci and recovery of well-known loci (from off-target reads) with historical uses in Sanger-based molecular phylogenetic studies. Hybridization temperatures did not generally affect recovery of newly targeted loci, which we attributed to lower bait-target divergences (compared to higher divergences between UCE baits and targets) and greater bait tiling density. Thus, optimizing in vitro target capture conditions to accommodate low hybridization temperatures can provide a cost-effective, widely applicable solution to improve recovery of protein-coding loci in invertebrates, while retrieving other potentially useful data for downstream comparative analyses.