Christine Keyser

Ancient DNA provides new insights into the history of south Siberian Kurgan people

To help unravel some of the early Eurasian steppe migration movements, we determined the Y-chromosomal and mitochondrial haplotypes and haplogroups of 26 ancient human specimens from the Krasnoyarsk area dated from between the middle of the second millennium BC. to the fourth century AD. In order to go further in the search of the geographic origin and physical traits of these south Siberian specimens, we also typed phenotype-informative single nucleotide polymorphisms. Our autosomal, Y-chromosomal and mitochondrial DNA analyses reveal that whereas few specimens seem to be related matrilineally or patrilineally, nearly all subjects belong to haplogroup R1a1-M17 which is thought to mark the eastward migration of the early Indo-Europeans. Our results also confirm that at the Bronze and Iron Ages, south Siberia was a region of overwhelmingly predominant European settlement, suggesting an eastward migration of Kurgan people across the Russo-Kazakh steppe. Finally, our data indicate that at the Bronze and Iron Age timeframe, south Siberians were blue (or green)-eyed, fair-skinned and light-haired people and that they might have played a role in the early development of the Tarim Basin civilization. To the best of our knowledge, no equivalent molecular analysis has been undertaken so far.

RNA/DNA co-analysis from human menstrual blood and vaginal secretion stains: Results of a fourth and fifth collaborative EDNAP exercise

The European DNA Profiling Group (EDNAP) organized a fourth and fifth collaborative exercise on RNA/DNA co-analysis for body fluid identification and STR profiling. The task was to identify dried menstrual blood and vaginal secretion stains using specific RNA biomarkers, and additionally test 3 housekeeping genes for their suitability as reference genes. Six menstrual blood and six vaginal secretion stains, two dilution series (1/4-1/64 pieces of a menstrual blood/vaginal swab) and, optionally, bona fide or mock casework samples of human or non-human origin were analyzed by 24 participating laboratories, using RNA extraction or RNA/DNA co-extraction methods. Two novel menstrual blood mRNA multiplexes were used: MMP triplex (MMP7, MMP10, MMP11) and MB triplex (MSX1, LEFTY2, SFRP4) in conjunction with a housekeeping gene triplex (B2M, UBC, UCE). Two novel mRNA multiplexes and a HBD1 singleplex were used for the identification of vaginal secretion: Vag triplex (MYOZ1, CYP2B7P1 and MUC4) and a Lactobacillus-specific Lacto triplex (Ljen, Lcris, Lgas). The laboratories used different chemistries and instrumentation and all were able to successfully isolate and detect mRNA in dried stains. The simultaneous extraction of RNA and DNA allowed for positive identification of the tissue/fluid source of origin by mRNA profiling as well as a simultaneous identification of the body fluid donor by STR profiling, also from old and compromised casework samples. The results of this and the previous collaborative RNA exercises support RNA profiling as a reliable body fluid identification method that can easily be combined with current STR typing technology.

Comparison between morphological and genetic data to estimate biological relationship: The case of the Egyin Gol necropolis (Mongolia)

Osseous and dental nonmetric (discrete) traits have long been used to assess population variability and affinity in anthropological and archaeological contexts. However, the full extent to which nonmetric traits can reliably be used as a proxy for genetic data when assessing close or familial relationships is currently poorly understood. This study represents the unique opportunity to directly compare genetic and nonmetric data for the same individuals excavated from the Egyin Gol necropolis, Mongolia. These data were analyzed to consider the general efficacy of nonmetric traits for detecting familial groupings in the absence of available genetic data. The results showed that the Egyin Gol population is quite homogenous both metrically and genetically confirming a previous suggestion that the same people occupied the necropolis throughout the five centuries of its existence. Kinship analysis detected the presence of potential family burials in the necropolis. Moreover, individuals buried in one sector of the necropolis were differentiated from other sectors on the basis of nonmetric data. This separation is likely due to an outside Turkish influence in the paternal line, as indicated by the results of Y-chromosome analysis. Affinity matrices based on nonmetric and genetic data were correlated demonstrating the potential of nonmetric traits for detecting relationships in the absence of genetic data. However, the strengths of the correlations were relatively low, cautioning against the use of nonmetric traits when the resolution of the familial relationships is low.

Variola Virus in a 300-Year-Old Siberian Mummy

This letter describes a distant lineage of the variola virus (the agent of smallpox) that was identified in a mummy found buried in the Siberian permafrost.

Efficiency evaluation of a DNA extraction and purification protocol on archival formalin-fixed and paraffin-embedded tissue.

Formalin-fixed and paraffin-embedded tissue (FF-PET) is an invaluable resource for retrospective molecular genetic studies, but the extraction of high-quality genomic DNA from FF-PET is still a problematic issue. Despite the range of DNA extraction methods currently in use, the association of phenol-chloroform extraction and silica-based purification protocols, reported in ancient DNA studies on archaeological bones, has, to our knowledge, not been used for DNA extraction from FF-PET yet. The present study compared the efficiency of three DNA extraction and purification protocols from two different FF-PET substrates, heart and liver, by using quantitative PCR and multiplex amplification. We showed that the method, using phenol-chloroform and the QIAamp DNA mini Kit (Qiagen), was the most effective DNA extraction and purification method and that the DNA quantity extracted from liver is statistically more important than that extracted from heart. Autosomal STR typing by multiplex amplifications gave partial allelic profiles with only small size products (less than 300 bases) amplified, suggesting that DNA extracted from FF-PET was degraded. In conclusion, the protocol presented here, previously described in studies on ancient bones, should find application in different molecular studies involving FF-PET material.

Genetic analysis of human remains from a double inhumation in a frozen kurgan in Kazakhstan (Berel site, Early 3rd Century BC).

Abstract. The discovery of a big barrow of the Saka period in eastern Kazakhstan between the Russian and the Chinese borders provided the opportunity to excavate a frozen burial site. In the burial chamber, there was a wooden sarcophagus with two human bodies. The skeletons of these two individuals, a man and a woman, were well preserved. A genetic study based on STRs and mitochondrial DNA analyses was undertaken in order to determine whether these human remains belonged to close relatives. Results were obtained for all the markers. Nevertheless, nuclear STRs did not allow a clear conclusion concerning the relationship, but analysis of mitochondrial DNA showed that these skeletons were not close relatives.

Application of the iPLEXTM Gold SNP genotyping method for the analysis of Amerindian ancient DNA samples: Benefits for ancient population studies

Important developments in the matrix‐assisted laser desorption/ionization time‐of‐flight mass spectrometry (MALDI‐TOF MS) technique have generated new perspectives regarding SNP genotyping, which are particularly promising for ancient population‐based studies. The main aim of the present study was to investigate the application of a MALDI‐TOF MS‐based SNP genotyping technique, called iPLEX® Gold, to analyze Amerindian ancient DNA samples. The first objective was to test the sensitivity of the method, which is recommended for DNA quantities between 10 and 5 ng, for ancient biological samples containing DNA molecules that were degraded and present in minute quantities. The second objective was to detail the advantages of this technique for studies on ancient populations. Two multiplexes were designed, allowing the major Amerindian mitochondrial and Y haplogroups to be determined simultaneously. This analysis has never been described before. Results demonstrated the reliability and accuracy of the method; data were obtained for both mitochondrial and nuclear DNA using picogram (pg) quantities of nucleic acid. This technique has the advantages of both MS and minisequencing techniques; thus, it should be included in the protocols for future ancient DNA studies.

Matrix-Assisted Laser Desorption Ionization-Time of Flight Mass Spectrometry-Based Single Nucleotide Polymorphism Genotyping Assay Using iPLEX Gold Technology for Identification of Mycobacterium tuberculosis Complex Species and Lineages

ABSTRACT The major goal of the present study was to investigate the potential use of a novel single nucleotide polymorphism (SNP) genotyping technology, called iPLEX Gold (Sequenom), for the simultaneous analysis of 16 SNPs that have been previously validated as useful for identification of Mycobacterium tuberculosis complex (MTBC) species and classification of MTBC isolates into distinct genetic lineages, known as principal genetic groups (PGGs) and SNP cluster groups (SCGs). In this context, we developed a 16-plex iPLEX assay based on an allele-specific-primer single-base-extension reaction using the iPLEX Gold kit (Sequenom), followed by matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) analysis on the commercially available Sequenom MassARRAY platform. This assay was tested on a panel of 55 well-characterized MTBC strains that were also genotyped for the same loci using the previously reported SNaPshot assay, as well as 10 non-MTBC mycobacteria and 4 bacteria not belonging to the genus Mycobacterium. All MTBC samples were successfully analyzed with the iPLEX assay, which yielded clear allelic data for 99.9% of the SNPs (879 out of 880). No false-positive results were obtained with the negative controls. Compared to the SNaPshot assay, the newly developed 16-plex iPLEX assay produced fully concordant results that allowed reliable differentiation of MTBC species and recognition of lineages, thus demonstrating its potential value in diagnostic, epidemiological, and evolutionary applications. Compared to the SNaPshot approach, the implementation of the iPLEX technology could offer a higher throughput and could be a more flexible and cost-effective option for microbiology laboratories.

Human evolution in Siberia: from frozen bodies to ancient DNA

BackgroundThe Yakuts contrast strikingly with other populations from Siberia due to their cattle- and horse-breeding economy as well as their Turkic language. On the basis of ethnological and linguistic criteria as well as population genetic studies, it has been assumed that they originated from South Siberian populations. However, many questions regarding the origins of this intriguing population still need to be clarified (e.g. the precise origin of paternal lineages and the admixture rate with indigenous populations). This study attempts to better understand the origins of the Yakuts by performing genetic analyses on 58 mummified frozen bodies dated from the 15th to the 19th century, excavated from Yakutia (Eastern Siberia).ResultsHigh quality data were obtained for the autosomal STRs, Y-chromosomal STRs and SNPs and mtDNA due to exceptional sample preservation. A comparison with the same markers on seven museum specimens excavated 3 to 15 years ago showed significant differences in DNA quantity and quality. Direct access to ancient genetic data from these molecular markers combined with the archaeological evidence, demographical studies and comparisons with 166 contemporary individuals from the same location as the frozen bodies helped us to clarify the microevolution of this intriguing population.ConclusionWe were able to trace the origins of the male lineages to a small group of horse-riders from the Cis-Baïkal area. Furthermore, mtDNA data showed that intermarriages between the first settlers with Evenks women led to the establishment of genetic characteristics during the 15th century that are still observed today.

Paleogenetical Study of Pre-Columbian Samples From Pampa Grande (Salta, Argentina)

Ancient DNA recovered from 21 individuals excavated from burial sites in the Pampa Grande (PG) region (Salta province) of North-Western Argentina (NWA) was analyzed using various genetic markers (mitochondrial DNA, autosomal STRs, and Y chromosomal STRs). The results were compared to ancient and modern DNA from various populations in the Andean and North Argentinean regions, with the aim of establishing their relationships with PG. The mitochondrial haplogroup frequencies described (11% A, 47% B, and 42% D) presented values comparable to those found for the ancient Andean populations from Peru and San Pedro de Atacama. On the other hand, mitochondrial and Y chromosomal haplotypes were specific to PG, as they did not match any other of the South American populations studied. The described genetic diversity indicates homogeneity in the genetic structure of the ancient Andean populations, which was probably facilitated by the intense exchange network in the Andean zone, in particular among Tiwanaku, San Pedro de Atacama, and NWA. The discovery of haplotypes unique to PG could be due to a loss of genetic diversity caused by recent events affecting the autochthonous populations (establishment of the Inca Empire in the region, colonization by the Europeans).