Pre-sampling conditions affect salivary genomic DNA yield and human DNA fraction: a single-donor pilot study

As saliva-based genomic analyses expand, the effect of common pre-sampling behaviors on salivary genomic DNA (gDNA) recovery remains poorly characterized. In this single-donor pilot study, we tested eight pre-sampling conditions combining tooth brushing, gargling, and water intake. Saliva was collected in triplicate under each condition, cells were isolated, and gDNA was extracted using the Qiagen Kit. Three-way ANOVA showed that brushing was the dominant factor, accounting for 62.9% of total variation in DNA yield. Gargling had a smaller but significant effect (5.6%), while water intake had no significant effect. DNA purity, assessed by A260/280 ratio, remained stable across all conditions. To assess the human-to-bacterial DNA ratio, we performed qPCR using RPPH1 (human) and 16S rRNA (bacterial) markers. Brushing increased the human DNA fraction but reduced the absolute amount of human DNA recovered, indicating a purity-yield trade-off. These single-donor pilot findings offer preliminary, quantitative guidance for pre-sampling conditions in saliva-based genomic analyses, to be validated in multi-donor cohorts.