Abstract
Ancient DNA (aDNA) recovered from archaeobotanical remains can provide key insights into many prominent archaeological research questions, including processes of domestication, past subsistence strategies, and human interactions with the environment. However, it is often difficult to isolate aDNA from ancient plant materials, and furthermore, such DNA extracts frequently contain inhibitory substances that preclude successful PCR amplification. In the age of high-throughput sequencing, this problem is even more significant because each additional endogenous aDNA molecule improves analytical resolution. Therefore, in this paper, we compare a variety of DNA extraction techniques on primarily desiccated archaeobotanical remains and identify which method consistently yields the greatest amount of purified DNA. In addition, we test five DNA polymerases to determine how well they replicate DNA extracted from non-charred ancient plant remains. Based upon the criteria of resistance to enzymatic inhibition, behavior in quantitative real-time PCR, replication fidelity, and compatibility with aDNA damage, we conclude these polymerases have nuanced properties, requiring researchers to make educated decisions as to which one to use for a given task. The experimental findings should prove useful to the aDNA and archaeological communities by guiding future research methodologies and ensuring precious archaeobotanical remains are studied in optimal ways, and may thereby yield important new perspectives on the interactions between humans and past plant communities.
Highlights
Ancient DNA studies have become an integral part of Quaternary research, providing invaluable anthropological and biological insights, on issues as diverse as human evolution [1], modern human migrations [2,3,4], plant and animal domestication [5,6], and paleoecology [7]
In addition to the issues of contamination and biomolecular degradation faced by all aDNA research [12], ancient plant materials frequently contain compounds that impede DNA extraction and enzymatic reactions, including the polymerase chain reaction (PCR)
Sediments adhering to charred cereals may contain humic acids that could inhibit PCR. We suggest it is worth conducting PCR with bovine serum albumin (BSA) to ensure enzymatic inhibition does not lead to false negative results
Summary
Ancient DNA (aDNA) studies have become an integral part of Quaternary research, providing invaluable anthropological and biological insights, on issues as diverse as human evolution [1], modern human migrations [2,3,4], plant and animal domestication [5,6], and paleoecology [7]. Research on plant aDNA from archaeological contexts is of particular interest because archaeobotanical remains can provide important data on subsistence patterns, human behavioral variability, domestication, and broader environmental issues [8,9,10]. Despite this rich potential, relatively few researchers have studied aDNA from plant materials [9,11]; the scarcity of this line of research can be partially attributed to the many methodological challenges posed by ancient plant materials. Even when DNA eluates are visually transparent, inhibitors may still be present, leading to PCR failures
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