Data from: Effect of marker choice and thermal cycling protocol on zooplankton DNA metabarcoding studies

DNA metabarcoding is a promising approach for rapidly surveying biodiversity and is likely to become an important tool for measuring ecosystem responses to environmental change. Metabarcoding markers need sufficient taxonomic coverage to detect groups of interest, sufficient sequence divergence to resolve species, and will ideally indicate relative abundance of taxa present. We characterized zooplankton assemblages with three different metabarcoding markers (nuclear 18S rDNA, mitochondrial COI, and mitochondrial 16S rDNA) to compare their performance in terms of taxonomic coverage, taxonomic resolution, and correspondence between morphology- and DNA-based identification. COI amplicons sequenced on separate runs showed that operational taxonomic units representing >0.1% of reads per sample were highly reproducible, although slightly more taxa were detected using a lower annealing temperature. Mitochondrial COI and nuclear 18S showed similar taxonomic coverage across zooplankton phyla. However, mitochondrial COI resolved up to threefold more taxa to species compared to 18S. All markers revealed similar patterns of beta-diversity, although different taxa were identified as the greatest contributors to these patterns for 18S. For calanoid copepod families, all markers displayed a positive relationship between biomass and sequence reads, although the relationship was typically strongest for 18S. The use of COI for metabarcoding has been questioned due to lack of conserved primer-binding sites. However, our results show the taxonomic coverage and resolution provided by degenerate COI primers, combined with a comparatively well-developed reference sequence database, make them valuable metabarcoding markers for biodiversity assessment.

DNA 元条形码（DNA metabarcoding）是一种极具前景的生物多样性快速调查手段，有望成为衡量生态系统对环境变化响应的重要工具。元条形码标记需具备足够的分类学覆盖范围以检出目标类群、足够的序列差异以区分物种，且理想情况下可反映现存类群的相对丰度。本研究采用三类不同的元条形码标记（核18S rDNA、线粒体COI以及线粒体16S rDNA）对浮游动物群落进行表征，以比较三者在分类学覆盖范围、分类分辨率以及形态学鉴定与基于DNA的鉴定结果一致性方面的表现。在独立测序批次中获得的COI扩增子数据显示，占单样本序列读段数0.1%以上的操作分类单元（operational taxonomic unit, OTU）具有极高的可重复性；尽管采用较低退火温度可检测到略多的类群。线粒体COI与核18S在浮游动物各门的分类学覆盖范围上表现相近，但COI对物种的分辨能力可达18S的三倍。所有标记均呈现出相似的β多样性模式，但针对18S的分析中，被认定为该模式最大贡献者的类群有所不同。对于哲水蚤目（calanoid copepod）科类群，所有标记均显示生物量与序列读段数之间存在正相关关系，不过该相关性在18S标记中通常最为显著。此前因缺乏保守的引物结合位点，COI用于元条形码研究的可行性曾受到质疑，但本研究结果表明，简并COI引物所提供的分类学覆盖范围与分辨率，结合相对完善的参考序列数据库，使其成为生物多样性评估中极具价值的元条形码标记。