DNA recovery and identification from stone tool microcracks

Graduation date: 2003The studies described here introduce a model for residue preservation on stone\ud tools. They simulate stone tool manufacture in order to define parameters important\ud for the study of DNA residues. Microscopic examination of stone tools has identified\ud microcracks that trap DNA and protein from animal blood cells. Thorough\ud investigation of different methods to recover residues from stone tools shows that\ud surface washing leaves DNA and protein, trapped in subsurface microcracks.\ud However, other extraction techniques are able to release 60-80% of DNA and protein\ud residues.\ud Previous research documents the identification of protein from stone tools\ud sonicated in 5% ammonium hydroxide, but it remains untested whether the same\ud treatment yields useable DNA. Using this treatment, I identified 13-year-old DNA\ud residues from experimentally manufactured stone tools. In addition, results clearly\ud indicate that washing procedures typically used to curate stone tools removed only a\ud small fraction of the DNA deposited during animal butchery.\ud Twenty-four pieces of chipped stone recovered from the Bugas-Holding site\ud were studied to explore the validity of ancient DNA residue identifications. Nine tools\ud yielded DNA residues. Modern humans did not touch three of these tools, which\ud suggests that the DNA recovered from them was present prior to excavation. One\ud tool, which was handled by excavators without gloves, harbored DNA from three\ud species, and these templates competed during PCR. On at least two tools, handling\ud after excavation introduced animal DNA unrelated to tool use. Careful testing of\ud Bugas-Holding chipped stone suggests that stone tools may harbor both ancient and\ud modern DNA, and that investigators must take great care to exclude modern DNA\ud from ancient specimens.\ud Ultimately, I developed and streamlined a method to analyze DNA-containing\ud residues preserved on stone tools. This led to several technical improvements in\ud ancient DNA residue analysis. These include a more effective DNA recovery\ud protocol, methods to measure sensitivity and inhibition of PCR in each sample, and\ud strategies to surmount competition between templates during amplification, which can\ud occur in samples that contain DNA from multiple species. These new developments\ud will help future investigators achieve the full potential of ancient DNA residue\ud analysis