Maximizing Molecular Data From Low-Quality Fluid-Preserved Specimens in Natural History Collections
Maximizing Molecular Data From Low-Quality Fluid-Preserved Specimens in Natural History Collections
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DOI：
10.3389/fevo.2022.893088
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发表时间：
2022-06-30
影响因子：
3
通讯作者：
Ruane, Sara
中科院分区：
文献类型：
--
作者：
Bernstein, Justin M.;Ruane, Sara
在过去的十年中，博物馆基因组学研究主要集中在从液体保存的自然历史标本中获得足够质量和数量的DNA用于测序，主要用于系统研究。虽然这些研究为全世界许多物种的进化和生物多样性知识打开了一扇窗，但已发表的作品往往集中在这些DNA测序工作的成功上，这无疑比从自然历史收藏的标本中获得很少或有时没有DNA或无法使用的序列数据更少见。在这里，我们试图从115个新鲜的和41个降解的类蛇样本中获得DNA提取物并对其进行测序，以及从两种不为人知的蛇，Hydrablabes periops的降解样本中获得DNA提取物。水合蛇被认为至少属于两个不同的科（水合蛇科和Homalopsidae），由于没有已知的新鲜组织，棘手的博物馆标本目前提供了确定这种蛇的分类亲和力的唯一机会。虽然我们的目标是为这些样本生成目标捕获数据集，以便纳入更广泛的系统发育研究，但由于大量缺失数据，特别是使用与标准高质量样本相同的下游方法，结果并不理想。然而，我们并没有完全忽略结果，而是使用了参考文献和假参考文献的作图方法，以及系统发育分析，以最大限度地利用我们测序工作中的任何可用的分子数据，确定了H. periops的分类亲和力，并比较了新鲜和降解组织样本之间的测序成功。这导致了5个标本的线粒体基因组基本完整，以及数百到数千个核位点（超保守位点、锚定杂交富集位点和各种经常用于鳞状动物系统发育研究的位点）的保存，包括菲尔德自然历史博物馆收藏的一种圆形蛇标本。我们将H. periops数据与先前发表的基因组和sanger测序数据集结合起来，确认了该分类单元的家族名称，拒绝了先前的分类假设，并对Hydrablabes进行了生物地理学推断。第二个周期人标本，尽管在最初的原始测序结果和经过相同的程序之后看起来很相似，但几乎没有得到可用的分子数据。我们讨论了使用不同的管道和方法来最大限度地从这些数据中获得产品的成功和失败，并为其他希望在可能已降解DNA的标本上使用DNA测序工作的人提供了期望。
Over the past decade, museum genomics studies have focused on obtaining DNA of sufficient quality and quantity for sequencing from fluid-preserved natural history specimens, primarily to be used in systematic studies. While these studies have opened windows to evolutionary and biodiversity knowledge of many species worldwide, published works often focus on the success of these DNA sequencing efforts, which is undoubtedly less common than obtaining minimal or sometimes no DNA or unusable sequence data from specimens in natural history collections. Here, we attempt to obtain and sequence DNA extracts from 115 fresh and 41 degraded samples of homalopsid snakes, as well as from two degraded samples of a poorly known snake, Hydrablabes periops. Hydrablabes has been suggested to belong to at least two different families (Natricidae and Homalopsidae) and with no fresh tissues known to be available, intractable museum specimens currently provide the only opportunity to determine this snake's taxonomic affinity. Although our aim was to generate a target-capture dataset for these samples, to be included in a broader phylogenetic study, results were less than ideal due to large amounts of missing data, especially using the same downstream methods as with standard, high-quality samples. However, rather than discount results entirely, we used mapping methods with references and pseudoreferences, along with phylogenetic analyses, to maximize any usable molecular data from our sequencing efforts, identify the taxonomic affinity of H. periops, and compare sequencing success between fresh and degraded tissue samples. This resulted in largely complete mitochondrial genomes for five specimens and hundreds to thousands of nuclear loci (ultra-conserved loci, anchored-hybrid enrichment loci, and a variety of loci frequently used in squamate phylogenetic studies) from fluid-preserved snakes, including a specimen of H. periops from the Field Museum of Natural History collection. We combined our H. periops data with previously published genomic and Sanger-sequenced datasets to confirm the familial designation of this taxon, reject previous taxonomic hypotheses, and make biogeographic inferences for Hydrablabes. A second H. periops specimen, despite being seemingly similar for initial raw sequencing results and after being put through the same protocols, resulted in little usable molecular data. We discuss the successes and failures of using different pipelines and methods to maximize the products from these data and provide expectations for others who are looking to use DNA sequencing efforts on specimens that likely have degraded DNA.