Analysis of radiocarbon, stable isotopes and DNA in teeth to facilitate identification of unknown decedents.

The characterization of unidentified bodies or suspected human remains is a frequent and important task for forensic investigators. However, any identification method requires clues to the person's identity to allow for comparisons with missing persons. If such clues are lacking, information about the year of birth, sex and geographic origin of the victim, is particularly helpful to aid in the identification casework and limit the search for possible matches. We present here results of stable isotope analysis of (13)C and (18)O, and bomb-pulse (14)C analyses that can help in the casework. The (14)C analysis of enamel provided information of the year of birth with an average absolute error of 1.8±1.3 years. We also found that analysis of enamel and root from the same tooth can be used to determine if the (14)C values match the rising or falling part of the bomb-curve. Enamel laydown times can be used to estimate the date of birth of individuals, but here we show that this detour is unnecessary when using a large set of crude (14)C data of tooth enamel as a reference. The levels of (13)C in tooth enamel were higher in North America than in teeth from Europe and Asia, and Mexican teeth showed even higher levels than those from USA. DNA analysis was performed on 28 teeth, and provided individual-specific profiles in most cases and sex determination in all cases. In conclusion, these analyses can dramatically limit the number of possible matches and hence facilitate person identification work.

Finding the founder of Stockholm - a kinship study based on Y-chromosomal, autosomal and mitochondrial DNA.

Historical records claim that Birger Magnusson (died 1266), famous regent of Sweden and the founder of Stockholm, was buried in Varnhem Abbey in Västergötland. After being lost for centuries, his putative grave was rediscovered during restoration work in the 1920s. Morphological analyses of the three individuals in the grave concluded that the older male, the female and the younger male found in the grave were likely to be Birger, his second wife Mechtild of Holstein and his son Erik from a previous marriage. More recent evaluations of the data from the 1920s seriously questioned these conclusions, ultimately leading to the reopening and reexamination of the grave in 2002. Ancient DNA-analyses were performed to investigate if the relationship between the three individuals matched what we would expect if the individuals were Birger, Erik and Mechtild. We used pyrosequencing of Y-chromosomal and autosomal SNPs and compared the results with haplogroup frequencies of modern Swedes to investigate paternal relations. Possible maternal kinship was investigated by deep FLX-sequencing of overlapping mtDNA amplicons. The authenticity of the sequences was examined using data from independent extractions, massive clonal data, the c-statistics, and real-time quantitative data. We show that the males carry the same Y-chromosomal haplogroup and thus we cannot reject a father-son type of relation. Further, as shown by the mtDNA analyses, none of the individuals are maternally related. We conclude that the graves indeed belong to Birger, Erik and Mechtild, or to three individuals with the exact same kind of biological relatedness.

More on contamination: the use of asymmetric molecular behavior to identify authentic ancient human DNA.

Authentication of ancient human DNA results is an exceedingly difficult challenge due to the presence of modern contaminant DNA sequences. Nevertheless, the field of ancient human genetics generates huge scientific and public interest, and thus researchers are rarely discouraged by problems concerning the authenticity of such data. Although several methods have been developed to the purpose of authenticating ancient DNA (aDNA) results, while they are useful in faunal research, most of the methods have proven complicated to apply to ancient human DNA. Here, we investigate in detail the reliability of one of the proposed criteria, that of appropriate molecular behavior. Using real-time polymerase chain reaction (PCR) and pyrosequencing, we have quantified the relative levels of authentic aDNA and contaminant human DNA sequences recovered from archaeological dog and cattle remains. In doing so, we also produce data that describes the efficiency of bleach incubation of bone powder and its relative detrimental effects on contaminant and authentic ancient DNA. We note that bleach treatment is significantly more detrimental to contaminant than to authentic aDNA in the bleached bone powder. Furthermore, we find that there is a substantial increase in the relative proportions of authentic DNA to contaminant DNA as the PCR target fragment size is decreased. We therefore conclude that the degradation pattern in aDNA provides a quantifiable difference between authentic aDNA and modern contamination. This asymmetrical behavior of authentic and contaminant DNA can be used to identify authentic haplotypes in human aDNA studies.

Extensive human DNA contamination in extracts from ancient dog bones and teeth.

Ancient DNA (aDNA) sequences, especially those of human origin, are notoriously difficult to analyze due to molecular damage and exogenous DNA contamination. Relatively few systematic studies have focused on this problem. Here we investigate the extent and origin of human DNA contamination in the most frequently used sources for aDNA studies, that is, bones and teeth from museum collections. To distinguish contaminant DNA from authentic DNA we extracted DNA from dog (Canis familiaris) specimens. We monitored the presence of a 148-bp human-specific and a 152-bp dog-specific mitochondrial DNA (mtDNA) fragment in DNA extracts as well as in negative controls. The total number of human and dog template molecules were quantified using real-time polymerase chain reaction (PCR), and the sequences were characterized by amplicon cloning and sequencing. Although standard precautions to avoid contamination were taken, we found that all samples from the 29 dog specimens contained human DNA, often at levels exceeding the amount of authentic ancient dog DNA. The level of contaminating human DNA was also significantly higher in the dog extracts than in the negative controls, and an experimental setup indicated that this was not caused by the carrier effect. This suggests that the contaminating human DNA mainly originated from the dog bones rather than from laboratory procedures. When cloned, fragments within a contaminated PCR product generally displayed several different sequences, although one haplotype was often found in majority. This leads us to believe that recognized criteria for authenticating aDNA cannot separate contamination from ancient human DNA the way they are presently used.