It is now widely agreed that the Native American founders originated from a Beringian source population ~15-18 thousand years ago (kya) and rapidly populated all of the New World, probably mainly ...following the Pacific coastal route. However, details about the migration into the Americas and the routes pursued on the continent still remain unresolved, despite numerous genetic, archaeological, and linguistic investigations. To examine the pioneering peopling phase of the South American continent, we screened literature and mtDNA databases and identified two novel mitochondrial DNA (mtDNA) clades, here named D1g and D1j, within the pan-American haplogroup D1. They both show overall rare occurrences but local high frequencies, and are essentially restricted to populations from the Southern Cone of South America (Chile and Argentina). We selected and completely sequenced 43 D1g and D1j mtDNA genomes applying highest quality standards. Molecular and phylogeographic analyses revealed extensive variation within each of the two clades and possibly distinct dispersal patterns. Their age estimates agree with the dating of the earliest archaeological sites in South America and indicate that the Paleo-Indian spread along the entire longitude of the American double continent might have taken even <2000 yr. This study confirms that major sampling and sequencing efforts are mandatory for uncovering all of the most basal variation in the Native American mtDNA haplogroups and for clarification of Paleo-Indian migrations, by targeting, if possible, both the general mixed population of national states and autochthonous Native American groups, especially in South America.
Abstract Insights into the human mitochondrial phylogeny have been primarily achieved by sequencing full mitochondrial genomes (mtGenomes). In forensic genetics (partial) mtGenome information can be ...used to assign haplotypes to their phylogenetic backgrounds, which may, in turn, have characteristic geographic distributions that would offer useful information in a forensic case. In addition and perhaps even more relevant in the forensic context, haplogroup-specific patterns of mutations form the basis for quality control of mtDNA sequences. The current method for establishing (partial) mtDNA haplotypes is Sanger-type sequencing (STS), which is laborious, time-consuming, and expensive. With the emergence of Next Generation Sequencing (NGS) technologies, the body of available mtDNA data can potentially be extended much more quickly and cost-efficiently. Customized chemistries, laboratory workflows and data analysis packages could support the community and increase the utility of mtDNA analysis in forensics. We have evaluated the performance of mtGenome sequencing using the Personal Genome Machine (PGM) and compared the resulting haplotypes directly with conventional Sanger-type sequencing. A total of 64 mtGenomes (>1 million bases) were established that yielded high concordance with the corresponding STS haplotypes (<0.02% differences). About two-thirds of the differences were observed in or around homopolymeric sequence stretches. In addition, the sequence alignment algorithm employed to align NGS reads played a significant role in the analysis of the data and the resulting mtDNA haplotypes. Further development of alignment software would be desirable to facilitate the application of NGS in mtDNA forensic genetics.
Pan-American mitochondrial DNA (mtDNA) haplogroup C1 has been recently subdivided into three branches, two of which (C1b and C1c) are characterized by ages and geographical distributions that are ...indicative of an early arrival from Beringia with Paleo-Indians. In contrast, the estimated ages of C1d--the third subset of C1--looked too young to fit the above scenario. To define the origin of this enigmatic C1 branch, we completely sequenced 63 C1d mitochondrial genomes from a wide range of geographically diverse, mixed, and indigenous American populations. The revised phylogeny not only brings the age of C1d within the range of that of its two sister clades, but reveals that there were two C1d founder genomes for Paleo-Indians. Thus, the recognized maternal founding lineages of Native Americans are at least 15, indicating that the overall number of Beringian or Asian founder mitochondrial genomes will probably increase extensively when all Native American haplogroups reach the same level of phylogenetic and genomic resolution as obtained here for C1d.
Highlights • We present a midi amplicon protocol involving two PCR multiplexes and amplicon sizes of 300–500 bp for massively parallel sequencing (MPS) of entire mitochondrial (mt)genomes from hair ...shaft samples. • We evaluate alternative protocols using larger amplification products (2–3 kbp) as well as MPS from hair samples without prior amplification. • Our study demonstrates that massively mtGenome sequencing is feasible from hair samples, which would significantly increase the discrimination power in forensic mtDNA testing.
The high number of matching haplotypes of the most common mitochondrial (mt)DNA lineages are considered to be the greatest limitation for forensic applications. This study investigates the potential ...to solve this constraint by massively parallel sequencing a large number of mitogenomes that share the most common West Eurasian mtDNA control region (CR) haplotype motif (263G 315.1C 16519C). We augmented a pilot study on 29 to a total of 216 Italian mitogenomes that represents the largest set of the most common CR haplotype compiled from a single country. The extended population sample confirmed and extended the huge coding region diversity behind the most common CR motif. Complete mitogenome sequencing allowed for the detection of 163 distinct haplotypes, raising the power of discrimination from 0 (CR) to 99.6% (mitogenome). The mtDNAs were clustered into 61 named clades of haplogroup H and did not reveal phylogeographic trends within Italy. Rapid individualization approaches for investigative purposes are limited to the most frequent H clades of the dataset, viz. H1, H3, and H7.
Immigrants from diverse origins have arrived in Paraguay and produced important demographic changes in a territory initially inhabited by indigenous Guarani. Few studies have been performed to ...estimate the proportion of Native ancestry that is still preserved in Paraguay and the role of females and males in admixture processes. Therefore, 548 individuals from eastern Paraguay were genotyped for three marker sets: mtDNA, Y-SNPs and autosomal AIM-InDels. A genetic homogeneity was found between departments for each set of markers, supported by the demographic data collected, which showed that only 43% of the individuals have the same birthplace as their parents. The results show a sex-biased intermarriage, with higher maternal than paternal Native American ancestry. Within the native mtDNA lineages in Paraguay (87.2% of the total), most haplogroups have a broad distribution across the subcontinent, and only few are concentrated around the Paraná River basin. The frequency distribution of the European paternal lineages in Paraguay (92.2% of the total) showed a major contribution from the Iberian region. In addition to the remaining legacy of the colonial period, the joint analysis of the different types of markers included in this study revealed the impact of post-war migrations on the current genetic background of Paraguay.
Highlights • 588 complete mtDNA haplotypes from three U.S. population groups are reported. • Nearly all haplotypes (>90%) were unique within each population sample. • Point heteroplasmy was observed ...in 23.8% of individuals. • The Sanger-based datasets were developed to current forensic quality standards. • The haplotypes can serve as a benchmark for evaluation of future mtGenome datasets.
Kaspar Hauser’s parentage has been the subject of research and debate for nearly 200 years. As for his possible aristocratic descent through the House of Baden, there is suspicion that he was swapped ...as a baby, kidnapped, and kept in isolation to bring a collateral lineage to the throne. In the last 28 years, various genetic analyses have been carried out to investigate this possible aristocratic origin. Previous results using less sensitive Sanger and electrophoresis-based methods were contradictory, and moreover, the authenticity of some samples was disputed, thus leaving the question open. Our analyses using modern capture- and whole genome-based massively parallel sequencing techniques reveal that the mitochondrial DNA haplotypes in different samples attributed to Kaspar Hauser were identical, demonstrating authenticity for the first time, and clearly different from the mitochondrial lineage of the House of Baden, which rules out a maternal relationship and thus the widely believed “Prince theory”.
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•Independent genetic mtDNA analyses in three laboratories gave identical results•Hair and blood samples attributed to Kaspar Hauser yielded the same mtDNA•Kaspar Hauser’s mtDNA is unambiguously different from the “Baden lineage”
Techniques in genetics; Molecular genetics; History
•The first forensic mtDNA population sample for Côte d’Ivoire is presented.•The first forensic mtDNA population sample for Rwanda is presented.•A mixed forensic mtDNA population sample for West ...Africa is presented.•The sample may serve as database for several yet uncovered countries of West Africa.•A strategy for use of impaired but precious population samples is presented.
This study provides 398 novel complete mitochondrial control region sequences that augment the still underrepresented data from Africa by three datasets: a mixed West African sample set deriving from 12 countries (n = 145) and datasets from Côte d’Ivoire (Ivory Coast) (n = 100) as well as Rwanda (n = 153). The analysis of mtDNA variation and genetic comparisons with published data revealed low random match probabilities in all three datasets and typical West African and East African diversity, respectively. Genetic parameters indicate that the presented mixed West African dataset may serve as first forensic mtDNA control region database for West Africa in general. In addition, a strategy for responsible forensic application of precious mtDNA population samples potentially containing close maternal relatives is outlined. The datasets will be uploaded to the forensic mtDNA database EMPOP (https://empop.online) upon publication.
Abstract Forensic mitochondrial DNA (mtDNA) testing requires appropriate, high quality reference population data for estimating the rarity of questioned haplotypes and, in turn, the strength of the ...mtDNA evidence. Available reference databases (SWGDAM, EMPOP) currently include information from the mtDNA control region; however, novel methods that quickly and easily recover mtDNA coding region data are becoming increasingly available. Though these assays promise to both facilitate the acquisition of mitochondrial genome (mtGenome) data and maximize the general utility of mtDNA testing in forensics, the appropriate reference data and database tools required for their routine application in forensic casework are lacking. To address this deficiency, we have undertaken an effort to: (1) increase the large-scale availability of high-quality entire mtGenome reference population data, and (2) improve the information technology infrastructure required to access/search mtGenome data and employ them in forensic casework. Here, we describe the application of a data generation and analysis workflow to the development of more than 400 complete, forensic-quality mtGenomes from low DNA quantity blood serum specimens as part of a U.S. National Institute of Justice funded reference population databasing initiative. We discuss the minor modifications made to a published mtGenome Sanger sequencing protocol to maintain a high rate of throughput while minimizing manual reprocessing with these low template samples. The successful use of this semi-automated strategy on forensic-like samples provides practical insight into the feasibility of producing complete mtGenome data in a routine casework environment, and demonstrates that large (>2 kb) mtDNA fragments can regularly be recovered from high quality but very low DNA quantity specimens. Further, the detailed empirical data we provide on the amplification success rates across a range of DNA input quantities will be useful moving forward as PCR-based strategies for mtDNA enrichment are considered for targeted next-generation sequencing workflows.
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