“An Optimized Method for the Extraction of Ancient Eukaryote DNA from Marine Sediments”, 2020-04-11 (; backlinks):
Marine sedimentary ancient DNA (sedaDNA) provides a powerful means to reconstruct marine paleo-communities across the food web. However, currently there are few optimized sedaDNA extraction protocols available to maximize the yield of small DNA fragments typical of ancient DNA (aDNA) across a broad diversity of eukaryotes.
We compared 7 combinations of sedaDNA extraction treatments and sequencing library preparations using marine sediments collected at a water depth of 104 m off Maria Island, Tasmania, in 2018. These 7 methods contrasted frozen versus refrigerated sediment, bead-beating induced cell lysis versus ethylenediaminetetraacetic acid (EDTA) incubation, DNA binding in silica spin columns versus in silica-solution, diluted versus undiluted DNA in shotgun library preparations to test potential inhibition issues during amplification steps, and size-selection of low molecular-weight (LMW) DNA to increase the extraction efficiency of sedaDNA.
Maximum efficiency was obtained from frozen sediments subjected to a combination of EDTA incubation and bead-beating, DNA binding in silica-solution, and undiluted DNA in shotgun libraries, across 45 marine eukaryotic taxa. We present an optimized extraction protocol integrating these steps, with an optional post-library LMW size-selection step to retain DNA fragments of ≤500 base pairs. We also describe a stringent bioinformatic filtering approach for metagenomic data and provide a comprehensive list of contaminants as a reference for future sedaDNA studies.
The new extraction and data-processing protocol should improve quantitative paleo-monitoring of eukaryotes from marine sediments, as well as other studies relying on the detection of highly fragmented and degraded eukaryote DNA in sediments.
[Keywords: ancient DNA, diatoms, dinoflagellates, haptophytes, Maria Island, metagenomics, plankton, seafloor, Tasmania]