Restriction site-associated DNA sequencing (RAD-seq) and related methods have become relatively common approaches to resolve species-level phylogeny. It is not clear, however, whether RAD-seq data matrices are well suited to relaxed clock inference of divergence times, given the size of the matrices and the abundance of missing data. We investigated the sensitivity of Bayesian relaxed clock estimates of divergence times to alternative analytical decisions on an empirical RAD-seq phylogenetic matrix. We explored the relative contribution of secondary calibration strategies, amount of missing data, and the data partition analyzed to overall variance in divergence times inferred using BEAST MCMC analyses of Carex section Schoenoxiphium (Cyperaceae)—a recent radiation for which we have nearly complete species sampling of RAD-seq data. The crown node for Schoenoxiphium was estimated to be 15.22 (9.56–21.18) Ma using a single calibration point and low missing data, 11.93 (8.07–16.03) Ma using multiple calibration points and low missing data, and 8.34 (5.41–11.22) using multiple calibrations but high missing data. We found that using matrices with more than half of the individuals with missing data inferred younger mean ages for all nodes. Moreover, we have found that our molecular clock estimates are sensitive to the positions of the calibration(s) in our phylogenetic tree (using matrices with low missing data), especially when only a single calibration was applied to estimate divergence times. These results argue for sensitivity analyses and caution in interpreting divergence time estimates from RAD-seq data.

中文翻译：

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评估发散时间估计对 Schoenoxiphium 的 RAD-seq 系统发育中的位点采样、校准点和模型先验的敏感性

限制性位点相关 DNA 测序 (RAD-seq) 和相关方法已成为解决物种水平系统发育的相对常见的方法。然而，鉴于矩阵的大小和缺失数据的丰富性，尚不清楚 RAD-seq 数据矩阵是否非常适合散度时间的宽松时钟推断。我们研究了散度时间的贝叶斯松弛时钟估计对经验 RAD-seq 系统发育矩阵的替代分析决策的敏感性。我们探讨了二级校准策略、缺失数据量和数据分区的相对贡献，分析了使用Carex部分Schoenoxiphium 的BEAST MCMC 分析推断的发散时间的总体方差（莎草科）——最近的一种辐射，我们对 RAD-seq 数据进行了几乎完整的物种采样。Schoenoxiphium的冠节点使用单个校准点和低缺失数据估计为 15.22 (9.56-21.18) Ma，使用多个校准点和低缺失数据估计为 11.93 (8.07-16.03) Ma，使用多个校准但高缺失数据估计为 8.34 (5.41-11.22)数据。我们发现，使用包含超过一半数据缺失个体的矩阵推断出所有节点的平均年龄更年轻。此外，我们发现我们的分子钟估计对系统发育树中校准的位置敏感（使用具有低缺失数据的矩阵），尤其是当仅应用单个校准来估计发散时间时。这些结果支持敏感性分析，并在解释来自 RAD-seq 数据的发散时间估计时要谨慎。