Nanopore sequencing of long ribosomal DNA amplicons enables portable and simple biodiversity assessments with high phylogenetic resolution across broad taxonomic scale
Nanopore sequencing of long ribosomal DNA amplicons enables portable and simple biodiversity assessments with high phylogenetic resolution across broad taxonomic scale
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DOI：
10.1093/gigascience/giz006
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发表时间：
2019-05-01
期刊：
影响因子：
9.2
通讯作者：
Prost, Stefan
中科院分区：
文献类型：
--
作者：
Krehenwinkel, Henrik;Pomerantz, Aaron;Prost, Stefan
背景：鉴于当前的生物多样性危机，DNA条形码正在发展成为量化全球生态系统状态变化的重要工具。目前的条形码协议通常依赖于短扩增子序列，这可以准确地识别群落中的生物实体，但在广泛的分类尺度上提供有限的系统发育分辨率。然而，群落的系统发育结构是生物多样性的重要组成部分。因此，需要一种结合强大的分类分配能力和高系统发育效用的条形码方法。一种可能的解决方案是在MinION平台（牛津纳米孔技术公司）上对长核糖体DNA （rDNA）扩增子进行测序。研究结果：利用多种动植物物种的数据集，以节肢动物为重点，我们组装了一个长rDNA条形码分析管道，并引入了一个新的软件（MiniBar）来解多重双索引纳米孔读取。我们发现在广泛的分类尺度上，长rDNA序列提供了极好的系统发育和分类分辨率。我们通过在偏远雨林中的小型移动实验室进行现场条形码，强调了我们方法的简单性。我们还测试了长rDNA扩增子通过元条形码分析群落多样性的效用，发现它们恢复了高度倾斜的多样性估计。结论：在MinION平台上对双索引长rDNA扩增子进行测序是一种简单、经济、便携和通用的真核生物DNA条形码方法。尽管使用长扩增子方法进行大量群落分析可能会引入偏差，但长rDNA扩增子方法为准确恢复生物群落的分类和系统发育多样性提供了强有力的工具。
Background: In light of the current biodiversity crisis, DNA barcoding is developing into an essential tool to quantify state shifts in global ecosystems. Current barcoding protocols often rely on short amplicon sequences, which yield accurate identification of biological entities in a community but provide limited phylogenetic resolution across broad taxonomic scales. However, the phylogenetic structure of communities is an essential component of biodiversity. Consequently, a barcoding approach is required that unites robust taxonomic assignment power and high phylogenetic utility. A possible solution is offered by sequencing long ribosomal DNA (rDNA) amplicons on the MinION platform (Oxford Nanopore Technologies). Findings: Using a dataset of various animal and plant species, with a focus on arthropods, we assemble a pipeline for long rDNA barcode analysis and introduce a new software (MiniBar) to demultiplex dual indexed Nanopore reads. We find excellent phylogenetic and taxonomic resolution offered by long rDNA sequences across broad taxonomic scales. We highlight the simplicity of our approach by field barcoding with a miniaturized, mobile laboratory in a remote rainforest. We also test the utility of long rDNA amplicons for analysis of community diversity through metabarcoding and find that they recover highly skewed diversity estimates. Conclusions: Sequencing dual indexed, long rDNA amplicons on the MinION platform is a straightforward, cost-effective, portable, and universal approach for eukaryote DNA barcoding. Although bulk community analyses using long-amplicon approaches may introduce biases, the long rDNA amplicons approach signifies a powerful tool for enabling the accurate recovery of taxonomic and phylogenetic diversity across biological communities.