Density separation of petrous bone powders for optimized ancient DNA yields
- Daniel M. Fernandes1,2,3,
- Kendra A. Sirak4,5,
- Olivia Cheronet1,3,
- Mario Novak6,
- Florian Brück1,
- Evelyn Zelger1,
- Alejandro Llanos-Lizcano1,
- Anna Wagner1,
- Anna Zettl1,
- Kirsten Mandl1,
- Kellie Sara Duffet Carlson1,3,
- Victoria Oberreiter1,3,
- Kadir T. Özdoğan1,7,
- Susanna Sawyer1,
- Francesco La Pastina8,
- Emanuela Borgia9,
- Alfredo Coppa1,3,10,
- Miroslav Dobeš11,
- Petr Velemínský12,
- David Reich4,5,
- Lynne S. Bell13 and
- Ron Pinhasi1,3
- 1Department of Evolutionary Anthropology, University of Vienna, 1030 Vienna, Austria;
- 2CIAS, Department of Life Sciences, University of Coimbra, 3000-456 Coimbra, Portugal;
- 3Human Evolution and Archaeological Sciences Forschungsverbund, University of Vienna, 1030 Vienna, Austria;
- 4Department of Genetics, Harvard Medical School, Boston, Massachusetts 02115, USA;
- 5Department of Human Evolutionary Biology, Harvard University, Cambridge, Massachusetts 02138, USA;
- 6Centre for Applied Bioanthropology, Institute for Anthropological Research, 10000 Zagreb, Croatia;
- 7Department of History and Art History, Utrecht University, 3512 BS Utrecht, The Netherlands;
- 8Department of Archaeology, University of Cambridge, Cambridge CB2 3DZ, United Kingdom;
- 9Dipartimento di Scienze dell'Antichità, Sapienza Università di Roma, Rome 00185, Italy;
- 10Dipartimento di Biologia Ambientale, Sapienza Università di Roma, Rome 00185, Italy;
- 11Institute of Archaeology of the Czech Academy of Sciences, Prague 118 00, Czech Republic;
- 12Department of Anthropology, National Museum, Prague 115 79, Czech Republic;
- 13Centre for Forensic Research, School of Criminology, Simon Fraser University, Burnaby, British Columbia V5A 1S6, Canada
Abstract
Density separation is a process routinely used to segregate minerals, organic matter, and even microplastics, from soils and sediments. Here we apply density separation to archaeological bone powders before DNA extraction to increase endogenous DNA recovery relative to a standard control extraction of the same powders. Using nontoxic heavy liquid solutions, we separated powders from the petrous bones of 10 individuals of similar archaeological preservation into eight density intervals (2.15 to 2.45 g/cm3, in 0.05 increments). We found that the 2.30 to 2.35 g/cm3 and 2.35 to 2.40 g/cm3 intervals yielded up to 5.28-fold more endogenous unique DNA than the corresponding standard extraction (and up to 8.53-fold before duplicate read removal), while maintaining signals of ancient DNA authenticity and not reducing library complexity. Although small 0.05 g/cm3 intervals may maximally optimize yields, a single separation to remove materials with a density above 2.40 g/cm3 yielded up to 2.57-fold more endogenous DNA on average, which enables the simultaneous separation of samples that vary in preservation or in the type of material analyzed. While requiring no new ancient DNA laboratory equipment and fewer than 30 min of extra laboratory work, the implementation of density separation before DNA extraction can substantially boost endogenous DNA yields without decreasing library complexity. Although subsequent studies are required, we present theoretical and practical foundations that may prove useful when applied to other ancient DNA substrates such as teeth, other bones, and sediments.
Footnotes
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[Supplemental material is available for this article.]
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Article published online before print. Article, supplemental material, and publication date are at https://www.genome.org/cgi/doi/10.1101/gr.277714.123.
- Received January 18, 2023.
- Accepted April 11, 2023.
This article is distributed exclusively by Cold Spring Harbor Laboratory Press for the first six months after the full-issue publication date (see https://genome.cshlp.org/site/misc/terms.xhtml). After six months, it is available under a Creative Commons License (Attribution-NonCommercial 4.0 International), as described at http://creativecommons.org/licenses/by-nc/4.0/.
