Data from: Optimizing methods for PCR-based analysis of predation

Molecular methods have become an important tool for studying feeding interactions under natural conditions. Despite their growing importance, many methodological aspects have not yet been evaluated but need to be considered to fully exploit the potential of this approach. Using feeding experiments with high alpine carabid beetles and lycosid spiders, we investigated how PCR annealing temperature affects prey DNA detection success and how post-PCR visualization methods differ in their sensitivity. Moreover, the replicability of prey DNA detection among individual PCR assays was tested using beetles and spiders that had digested their prey for extended times postfeeding. By screening all predators for three differently sized prey DNA fragments (range 116–612 bp), we found that only in the longest PCR product, a marked decrease in prey detection success occurred. Lowering maximum annealing temperatures by 4 °C resulted in significantly increased prey DNA detection rates in both predator taxa. Among the three post-PCR visualization methods, an eightfold difference in sensitivity was observed. Repeated screening of predators increased the total number of samples scoring positive, although the proportion of samples testing positive did not vary significantly between different PCRs. The present findings demonstrate that assay sensitivity, in combination with other methodological factors, plays a crucial role to obtain robust trophic interaction data. Future work employing molecular prey detection should thus consider and minimize the methodologically induced variation that would also allow for better cross-study comparisons.分子生物学方法已成为研究自然条件下取食相互作用的重要工具。尽管其重要性与日俱增，但诸多方法学层面的问题尚未得到系统评估，而若要充分挖掘该方法的应用潜力，这些问题均需纳入考量。本研究以高山步甲（carabid beetles）与狼蛛（lycosid spiders）为对象开展取食实验，探究了聚合酶链式反应（Polymerase Chain Reaction，PCR）退火温度对猎物DNA检测成功率的影响，以及不同PCR后可视化方法的灵敏度差异。此外，本研究还针对取食后已消化猎物较长时间的步甲与狼蛛样本，检验了单次PCR实验中猎物DNA检测的可重复性。研究人员针对所有捕食者样本，筛选了三种大小各异的猎物DNA片段（长度范围116–612 bp），结果发现仅在最长的PCR扩增产物中，猎物检测成功率出现了显著下降。将最高退火温度降低4℃后，两类捕食者类群的猎物DNA检出率均显著提升。在三种PCR后可视化方法中，灵敏度差异可达8倍。对捕食者样本的重复检测提升了呈阳性的样本总数，尽管不同PCR实验间的阳性样本占比并无显著差异。本研究结果表明，检测方法灵敏度结合其他方法学因素，对于获取可靠的营养级相互作用数据至关重要。未来采用分子猎物检测技术的研究，应考量并尽可能降低方法学层面引入的变异，这也将有助于实现更具可比性的跨研究比较。