Streamlined extraction of nucleic acids and metabolites from low- and high-biomass samples using isopropanol and matrix tubes

An essential aspect of population-based research is collecting samples outside of a clinical setting. This is crucial because microbial populations are highly dynamic, varying significantly across hosts, environments, and time points, a variability that clinical sample collection alone cannot fully capture. At-home sample collection enables the inclusion of a larger and more diverse group of participants, accounting for differences in ethnicity, age, and other factors. However, managing large studies is challenging due to the complexities involved in sample acquisition, processing, and analysis. Building on our previous work demonstrating the effectiveness of single 1 mL barcoded, racked Matrix Tubes in reducing sample processing time and well-to-well contamination for paired DNA and metabolite extraction, we further validate this method against a previously benchmarked plate-based approach using the same extraction reagents. This validation focuses on samples from the built environment, human skin, human saliva, and feces from mice and humans. Importantly, we explore the impact of using a mix of bead sizes during bead-beating for cell lysis, demonstrating that it enhances taxonomic recovery compared to a single bead size. Finally, we assess the potential of 95% isopropanol for room-temperature sample preservation. Our results show that isopropanol performs comparably to 95% ethanol in many cases, suggesting it is viable as an alternative when ethanol is unavailable. Beyond minimizing contamination, halving processing time, eliminating human error during sample plating, and streamlining metadata curation, the Matrix tube approach produces metabolomic, 16S, and shotgun metagenomic data consistent with the Plate-based Method for both high- and low-biomass samples.