Data from: Developing nuclear DNA phylogenetic markers in the angiosperm genus Leucadendron (Proteaceae): a next-generation sequencing transcriptomic approach

Despite the recent advances in generating molecular data, reconstructing species-level phylogenies for non-models groups remains a challenge. The use of a number of independent genes is required to resolve phylogenetic relationships, especially for groups displaying low polymorphism. In such cases, low-copy nuclear exons and non-coding regions, such as 3′ untranslated regions (3′-UTRs) or introns, constitute a potentially interesting source of nuclear DNA variation. Here, we present a methodology meant to identify new nuclear orthologous markers using both public-nucleotide databases and transcriptomic data generated for the group of interest by using next generation sequencing technology. To identify PCR primers for a non-model group, the genus Leucadendron (Proteaceae), we adopted a framework aimed at minimizing the probability of paralogy and maximizing polymorphism. We anchored when possible the right-hand primer into the 3′-UTR and the left-hand primer into the coding region. Seven new nuclear markers emerged from this search strategy, three of those included 3′-UTRs. We further compared the phylogenetic potential between our new markers and the ribosomal internal transcribed spacer region (ITS). The sequenced 3′-UTRs yielded higher polymorphism rates than the ITS region did. We did not find strong incongruences with the phylogenetic signal contained in the ITS region and the seven new designed markers but they strongly improved the phylogeny of the genus Leucadendron. Overall, this methodology is efficient in isolating orthologous loci and is valid for any non-model group given the availability of transcriptomic data.尽管分子数据生成技术近年取得诸多进展，但针对非模式类群重建物种级系统发育树仍是一项难题。厘清系统发育关系需借助多个独立基因，对于多态性较低的类群而言尤为如此。在此类场景下，低拷贝核外显子及非编码区域（如3′非翻译区（3′ untranslated regions, 3′-UTRs）或内含子），堪称核DNA变异的潜在优质来源。本研究提出一套方法，可同时利用公共核苷酸数据库与针对目标类群生成的转录组数据（借助下一代测序技术），筛选全新的核直系同源标记。为给非模式类群——山龙眼科（Proteaceae）Leucadendron属——设计聚合酶链式反应（PCR）引物，本研究采用了一套旨在降低旁系同源概率、最大化多态性的分析框架。我们尽可能将反向引物（右向引物）锚定至3′-UTR区域，正向引物（左向引物）锚定至编码区。通过该筛选策略共获得7个全新核标记，其中3个包含3′-UTR区域。本研究进一步对比了新开发标记与核糖体内部转录间隔区（ribosomal internal transcribed spacer region, ITS）的系统发育应用潜力。测序获得的3′-UTR区域多态性率高于ITS区域。本研究未发现新开发的7个标记与ITS区域所包含的系统发育信号存在显著冲突，但这些标记显著优化了Leucadendron属的系统发育重建结果。总体而言，该方法可高效分离直系同源基因座，且在具备目标类群转录组数据的前提下，适用于所有非模式类群。