A Versatile and Highly Efficient Toolkit Including 102 Nuclear Markers for Vertebrate Phylogenomics, Tested by Resolving the Higher Level Relationships of the Caudata
A Versatile and Highly Efficient Toolkit Including 102 Nuclear Markers for Vertebrate Phylogenomics, Tested by Resolving the Higher Level Relationships of the Caudata
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多功能且高效的工具包，包括 102 个用于脊椎动物系统基因组学的核标记，通过解决有尾目的高级关系进行测试
DOI：
10.1093/molbev/mst122
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发表时间：
2013-10-01
影响因子：
10.7
通讯作者：
Zhang, Peng
中科院分区：
文献类型：
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作者：
Shen, Xing Xing;Liang, Dan;Zhang, Peng
为脊椎动物生命树的任何部分解决困难的节点通常需要分析大量的基因座。开发对感兴趣的群体可行的分子标记往往是系统发育研究的瓶颈。在此，基于套式聚合酶链式反应(PCR)策略，我们提出了一个包括102个核蛋白编码基因座(NPCL)标记的通用工具包，用于脊椎动物系统学研究。102个NPCL标记具有广泛的进化速率，这使得它们对广泛的时间深度很有用。与以前开发的所有NPCL试剂盒相比，新的NPCL工具包具有三个重要优势：1)该试剂盒适用于脊椎动物，在16个差异很大的被测脊椎动物物种中的PCR成功率为94.6%；2)90%以上的PCR反应产生预期大小的强而单一的条带，可以直接测序；以及3)所有的Cleanup PCR反应都可以用两个特定的通用引物进行测序。为了测试其实际的系统发育效用，该工具包中的30个NPCL被用来研究活着的火蜥蜴的更高级别的关系。在对19个火蜥蜴和几个外群物种进行的639个目标PCR反应中，632个(98.9%)成功，602个(94.1%)直接测序。对这一30个基因座的数据集的连接和物种树分析产生了一个完全解析的系统发育图，并表明隐鳃总科(隐鳃总科+小蜥科)首先在火蜥蜴树内分枝，然后是Sirenidae。我们的实验测试和对特定案例的演示表明，我们的NPCL工具包是一种高度可靠、快速和经济高效的脊椎动物系统学研究方法，因此有可能加速完成脊椎动物生命树的许多部分。
Resolving difficult nodes for any part of the vertebrate tree of life often requires analyzing a large number of loci. Developing molecular markers that are workable for the groups of interest is often a bottleneck in phylogenetic research. Here, on the basis of a nested polymerase chain reaction (PCR) strategy, we present a universal toolkit including 102 nuclear protein-coding locus (NPCL) markers for vertebrate phylogenomics. The 102 NPCL markers have a broad range of evolutionary rates, which makes them useful for a wide range of time depths. The new NPCL toolkit has three important advantages compared with all previously developed NPCL sets: 1) the kit is universally applicable across vertebrates, with a PCR success rate of 94.6% in 16 widely divergent tested vertebrate species; 2) more than 90% of PCR reactions produce strong and single bands of the expected sizes that can be directly sequenced; and 3) all cleanup PCR reactions can be sequenced with only two specific universal primers. To test its actual phylogenetic utility, 30 NPCLs from this toolkit were used to address the higher level relationships of living salamanders. Of the 639 target PCR reactions performed on 19 salamanders and several outgroup species, 632 (98.9%) were successful, and 602 (94.1%) were directly sequenced. Concatenation and species-tree analyses on this 30-locus data set produced a fully resolved phylogeny and showed that Cryptobranchoidea (Cryptobranchidae + Hynobiidae) branches first within the salamander tree, followed by Sirenidae. Our experimental tests and our demonstration for a particular case show that our NPCL toolkit is a highly reliable, fast, and cost-effective approach for vertebrate phylogenomic studies and thus has the potential to accelerate the completion of many parts of the vertebrate tree of life.