DNA diagenesis and palaeogenetic analysis: Critical assessment and methodological progress
Abstract Palaeogenetic data obtained from fossilizing or fossil bones and teeth are of great importance to studies of vertebrate evolution, human biological and cultural evolution, plant and animal domestication and reconstructions of palaeoenvironment and palaeoecology. These studies are based on the retrieval of DNA preserved in fossilizing bones and teeth. DNA is present in fossils, if at all, in only very small amounts, which makes its amplification with PCR necessary for detailed sequence analysis. Erroneous nucleotides can be incorporated during in vitro amplification either because of post-mortem base damage of the original DNA template or simply because the fidelity of DNA polymerases is not absolute and can be decreased by suboptimal buffer conditions or possibly by compounds in the fossil extracts. These erroneously introduced nucleotides can be mistaken for authentic mutations of the ancient sequence compared to the closest extant sequence. Moreover, contamination by modern DNA, which is not chemically modified and therefore a better substrate for the Taq polymerase, can also lead to erroneous results. Here, we will present the procedures that we have developed in order to (i) ensure negligible mutagenicity of the PCR reaction, (ii) eliminate contamination by DNA molecules originating from previous PCR reactions and cloning procedures, (iii) prevent contamination with modern DNA of fossil bones and teeth during and after their excavation, and (iv) prevent degradation of ancient DNA after excavation. Finally, we will discuss our results on DNA preservation as a function of the taphonomy of the skeletal part that is analyzed and of the depositional context of preservation.