An evaluation of transcriptome-based exon capture for frog phylogenomics across multiple scales of divergence (Class: Amphibia, Order: Anura)

Abstract

Custom sequence capture experiments are becoming an efficient approach for gathering large sets of orthologous markers with targeted levels of informativeness in non-model organisms. Transcriptome-based exon capture utilizes transcript sequences to design capture probes, often with the aid of a reference genome to identify intron-exon boundaries and exclude shorter exons ( 200 bp), with no major reduction in coverage towards the ends of exons. We observed significant differences in the performance of blocking oligos for target enrichment and non-target depletion during captures, and observed differences in PCR duplication rates that can be attributed to the number of individuals pooled for capture reactions. We explicitly tested the effects of phylogenetic distance on capture sensitivity, specificity, and missing data, and provide a baseline estimate of expectations for these metrics based on nuclear pairwise differences among samples. We provide recommendations for transcriptome-based exon capture design based on our results, and describe multiple pipelines for data assembly and analysis.