Improved recovery of ancient DNA from subfossil wood – application to the world’s oldest Late Glacial pine forest. "Binz" subfossil pine ancient DNA

Ancient DNA from historical and subfossil wood has the potential to address important ecological and evolutionary questions, yet its extraction and analysis has not become routine, mainly because contamination of the wood with modern plant material can complicate the verification of genetic information. Here, we use sapwood tissue from 24 subfossil pines that were growing ~13,000 years BP in Zurich, Switzerland. We develop and evaluate protocols to eliminate surface contamination, and we test ancient DNA authenticity based on plastid DNA metabarcoding and the assessment of post-mortem DNA damage. A novel approach of laser irradiation coupled with bleaching and surface removal was most efficient in eliminating contaminating DNA. DNA metabarcoding confirmed which ancient DNA samples repeatedly amplified pine DNA and were free of exogenous plant taxa. Pine DNA sequences of these samples showed a high degree of cytosine to thymine mismatches, typical of post-mortem damage. This study attests that stringent decontamination of wood surfaces together with techniques of verifying DNA allow access to endogenous ancient DNA from the oldest, yet well preserved, Late Glacial pine forest, with the potential for genome-scale studies of many sources of relict wood, including different species from environments around the world.