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促进基于高通量样本收集的基因组学:DNA提取和文库构建方法的比较

Facilitating high throughput collections-based genomics: a comparison of DNA extraction and library building methods.

作者信息

Marsh William A, Hall Andie, Barnes Ian, Price Ben

机构信息

Natural History Museum, Cromwell Road, South Kensington, SW7 5BD, London, UK.

出版信息

Sci Rep. 2025 Feb 19;15(1):6013. doi: 10.1038/s41598-025-88443-0.

Abstract

While DNA barcoding methods are an increasingly important tool in biological conservation, the resource requirements of constructing reference libraries frequently reduce their efficacy. One efficient way of sourcing taxonomically validated DNA for reference libraries is to use museum collections. However, DNA degradation intrinsic to historical museum specimens can, if not addressed in the wet lab, lead to low quality data generation and severely limit scientific output. Several DNA extraction and library build methods that are designed to work with degraded DNA have been developed, although the ability to implement these methods at scale and at low cost has yet to be formally addressed. Here, the performance of widely used DNA extraction and library build methods are compared using museum specimens. We find that while our selected DNA extraction methods do not significantly differ in DNA yield, the Santa Cruz Reaction (SCR) library build method is not only the most effective at retrieving degraded DNA from museum specimens but also easily implemented at high throughput for low cost. Results highlight the importance of lab protocol on data yield. An optimised "sample to sequencing" high-throughput protocol which incorporates SCR is included to allow for easy uptake by the wider scientific community.

摘要

虽然DNA条形码方法在生物保护中是一种日益重要的工具,但构建参考文库所需的资源常常会降低其功效。为参考文库获取经过分类学验证的DNA的一种有效方法是利用博物馆藏品。然而,如果在湿实验室中不解决历史博物馆标本固有的DNA降解问题,可能会导致生成低质量数据,并严重限制科学产出。尽管尚未正式解决大规模且低成本实施这些方法的能力问题,但已经开发了几种旨在处理降解DNA的DNA提取和文库构建方法。在这里,我们使用博物馆标本比较了广泛使用的DNA提取和文库构建方法的性能。我们发现,虽然我们选择的DNA提取方法在DNA产量上没有显著差异,但圣克鲁斯反应(SCR)文库构建方法不仅在从博物馆标本中检索降解DNA方面最有效,而且易于以高通量低成本实施。结果突出了实验室方案对数据产量的重要性。文中包含了一个优化的“样本到测序”高通量方案,该方案纳入了SCR,以便更广泛的科学界能够轻松采用。

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/472a/11839992/7d2af4aaf497/41598_2025_88443_Fig1_HTML.jpg

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