High variability in SSU rDNA gene copy number among planktonic foraminifera revealed by single-cell qPCR@en

Metabarcoding has become the workhorse of community ecology. Sequencing a taxonomically informative DNA fragment from environmental samples gives fast access to community composition across taxonomic groups, but it relies on the assumption that the number of sequences for each taxon correlates with its abundance in the sampled community. However, gene copy number varies among and within taxa, and the extent of this variability must therefore be considered when interpreting community composition data derived from environmental sequencing. Here we measured with single-cell qPCR the SSU rDNA gene copy number of 139 specimens of five species of planktonic foraminifera. We found that the average gene copy number varied between of ~4 000 to ~50 000 gene copies between species, and individuals of the same species can carry between ~300 to more than 350 000 gene copies. This variability cannot be explained by differences in cell size and considering all plausible sources of bias, we conclude that this variability likely reflects dynamic genomic processes acting during the life cycle. We used the observed variability to model its impact on metabarcoding and found that the application of a correcting factor at species level may correct the derived relative abundances, provided sufficiently large populations have been sampled.元条形码（metabarcoding）技术已成为群落生态学的支柱性研究手段。通过对环境样本中携带分类学信息的DNA片段进行测序，可快速获取不同类群的群落组成信息，但该方法的核心前提假设为：每个类群的测序序列数与其在采样群落中的丰度呈正相关。然而，基因拷贝数在不同类群及同类群内部均存在变异，因此在解读基于环境测序得到的群落组成数据时，必须充分考量这种变异程度。本研究借助单细胞qPCR（single-cell qPCR）技术，测定了5种浮游有孔虫共计139个标本的小亚基核糖体DNA（SSU rDNA）基因拷贝数。结果显示，不同物种的平均基因拷贝数介于约4000至50000之间，而同一物种的不同个体，其基因拷贝数可从约300波动至350000以上。该变异无法通过细胞大小差异进行解释；在综合考量所有潜在偏倚来源后，我们认为这种变异大概率反映了生物生命周期中动态发生的基因组过程。我们利用观测到的基因拷贝数变异，对其在元条形码技术中的影响开展建模分析，结果表明：若采样种群规模足够大，通过在物种水平应用校正因子，即可有效校正得到的相对丰度数据。