Microvolume DNA extraction methods for microscale amplicon and metagenomic studies.

ABSTRACT: Investigating the composition and metabolic capacity of aquatic microbial assemblages usually requires the filtration of multi-litre samples, which are up to 1 million-fold larger than the microenvironments within which microbes are predicted to be spatially organised. To determine if community profiles can be reliably generated from microlitre volumes, we sampled seawater at a coastal and an oceanic site, filtered and homogenised them, and extracted DNA from bulk samples (2 L) and microvolumes (100, 10 and 1 μL) using two new approaches. These microvolume DNA extraction methods involve either physical or chemical lysis (through pH/thermal shock and lytic enzymes/surfactants, respectively), directly followed by the capture of DNA on magnetic beads. Downstream analysis of extracted DNA using both amplicon sequencing and metagenomics, revealed strong correlation with standard large volume approaches, demonstrating the fidelity of taxonomic and functional profiles of microbial communities in as little as 1 μL of seawater. This volume is six orders of magnitude smaller than most standard operating procedures for marine metagenomics, which will allow precise sampling of the heterogenous landscape that microbes inhabit.

SUBMITTER: Bramucci AR 

PROVIDER: S-EPMC9723667 | biostudies-literature | 2021 Dec

REPOSITORIES: biostudies-literature