Data from: Species tree estimation of North American chorus frogs (Hylidae: Pseudacris) with parallel tagged amplicon sequencing

The field of phylogenetics is changing rapidly with the application of high-throughput sequencing to non-model organisms. Cost-effective use of this technology for phylogenetic studies, which often include a relatively small portion of the genome but several taxa, requires strategies for genome partitioning and sequencing multiple individuals in parallel. In this study we estimated a multilocus phylogeny for the North American chorus frog genus Pseudacris using anonymous nuclear loci that were recently developed using a reduced representation library approach. We sequenced 27 nuclear loci and three mitochondrial loci for 44 individuals on 1/3 of an Illumina MiSeq run, obtaining 96.5% of the targeted amplicons at less than 20% of the cost of traditional Sanger sequencing. We found heterogeneity among gene trees, although four major clades (Trilling Frog, Fat Frog, crucifer, and West Coast) were consistently supported, and we resolved the relationships among these clades for the first time with strong support. We also found discordance between the mitochondrial and nuclear datasets that we attribute to mitochondrial introgression and a possible selective sweep. Bayesian concordance analysis in BUCKy and species tree analysis in *BEAST produced largely similar topologies, although we identify taxa that require additional investigation in order to clarify taxonomic and geographic range boundaries. Overall, we demonstrate the utility of a reduced representation library approach for marker development and parallel tagged sequencing on an Illumina MiSeq for phylogenetic studies of non-model organisms.

随着高通量测序（high-throughput sequencing）技术应用于非模式生物（non-model organisms），系统发育学（phylogenetics）领域正经历快速变革。针对通常仅覆盖基因组较小片段、却涉及多个类群的系统发育研究，要经济高效地应用该技术，需开发基因组分区策略并实现多个个体的平行测序。本研究利用新近通过简化基因组文库方法（reduced representation library approach）开发的匿名核基因座，对北美雨蛙属（Pseudacris）构建了多位点系统发育树。我们在一台Illumina MiSeq测序仪的1/3测序通量中，为44个个体完成了27个核基因座与3个线粒体基因座的测序，以不足传统桑格测序（Sanger sequencing）20%的成本，成功获取了96.5%的目标扩增子。研究发现基因树（gene trees）间存在异质性，但4个主要演化支（clades）——鸣蛙支系、胖蛙支系、十字支系与西海岸支系——始终得到有力支持，并首次以高置信度解析了这些演化支间的系统发育关系。我们还发现线粒体数据集与核数据集之间存在不一致性，该现象可归因于线粒体基因渐渗（mitochondrial introgression）与潜在的选择性清除（selective sweep）。借助BUCKy开展贝叶斯一致性分析（Bayesian concordance analysis），通过*BEAST进行物种树分析（species tree analysis），二者得到的拓扑结构（topologies）整体相似；不过我们也识别出部分类群需进一步研究，以明确其分类学地位与地理分布范围边界。综上，本研究证实了简化基因组文库方法用于分子标记开发，以及在Illumina MiSeq平台上开展平行标签测序，在非模式生物的系统发育研究中具备应用价值。