Propylene glycol-based antifreeze is an effective preservative for DNA metabarcoding of benthic arthropods

Preservation of DNA in bulk environmental samples is conventionally achieved using ethanol; however, transportation restrictions on ethanol, particularly from remote locations, are problematic, and ethanol requires a lengthy evaporation period to avoid polymerase chain reaction inhibition. We examined the efficacy of an easily accessible, non-toxic, propylene glycol-based antifreeze as an alternative to molecular-grade ethanol for preserving macroinvertebrate DNA from bulk-benthos DNA samples. We used 2 processing methods (no evaporation of preservative vs full evaporation) to test the differences in both cytochrome oxidase I (COI) exact sequence variants (ESVs) and COI taxonomic orders detected in both ethanol- and antifreeze-preserved samples. Our results suggest that antifreeze is a suitable alternative to ethanol for preservation of DNA in freshly collected samples (e.g., up to 3 d) because of the comparable ESV richness detected in antifreeze-preserved samples. We have demonstrated that by using antifreeze, it is possible to achieve sufficient taxonomic coverage and assess macroinvertebrate assemblages within bulk-benthos DNA samples. The application of this non-regulated preservative is particularly important for remote sampling (i.e., only air accessible) and sampling for community-based biomonitoring projects within Indigenous territories where alcohol is prohibited or not available.

Automated summary

The study found that propylene glycol-based antifreeze is an effective alternative to ethanol for preserving DNA in bulk environmental samples, with comparable DNA sequence variants detected in antifreeze-preserved samples as in ethanol-preserved samples. This non-toxic liquid shows good performance in DNA preservation and is particularly suitable for environments where ethanol is not available or prohibited, such as remote areas and Indigenous territories.