![]() Phylogenetic information can provide a valuable QC check of mtDNA data, identifying errors like artificial recombination and phantom mutations. Different patterns of LHP were observed across sequencing technologies, further supporting current guidelines to ignore LHP in database searches and match comparisons in forensic analyses. Since length heteroplasmy (LHP) can also complicate mtDNA data analysis, Paper II characterized LHP in data generated on two NGS platforms as well as with Sanger-type sequencing. Due to NUMT interference, a 10% variant frequency threshold was necessary to produce haplotypes consistent with high-quality mitogenome datasets. ![]() Paper I assessed the feasibility of generating forensic-quality mitogenome data from whole genome sequencing (WGS) data, which are valuable sources of mitogenome haplotypes for population studies. This thesis aimed to address the implementation challenges of mitogenome analysis and facilitate the transition to NGS in forensic laboratories. Including such false variants in mtDNA haplotypes can lead to erroneous conclusions based on misinterpreted data. Furthermore, appropriate quality control (QC) measures must be established as analysis can be complicated by nuclear mtDNA segments (NUMTs), misalignment of homopolymer regions, sequencing errors, and other artefacts. A better understanding of NGS methods and data analysis is also necessary to ensure the generation of reliable mitogenome data. The primary implementation challenge of mitogenome analysis is a lack of forensic-quality reference data, which are required to determine the evidentiary weight of a match. Though the discriminatory power of mtDNA is limited by common haplotypes, next generation sequencing (NGS) offers feasible access to entire mitochondrial genome (mitogenome) data that can provide increased resolution of common haplotypes to unique sequences. The high copy number and uniparental inheritance pattern of mtDNA are advantageous in cases involving shed hairs and aged skeletal elements, especially decades-old missing persons cases. Mitochondrial DNA (mtDNA) analysis plays a specialized role in forensic applications, overcoming certain limitations of autosomal DNA markers. Genetics Evolutionary Biology Forensic Science Research subject Biology with Specialisation in Human Evolution and Genetics Biology with specialization in Molecular Biology Identifiers URN: urn:nbn:se:uu:diva-470689 OAI: oai::uu-470689 DiVA, id: diva2:1647794Ģ022 (English) Doctoral thesis, comprehensive summary (Other academic) Abstract Mitochondrial DNA, haplogroup, quality control, artificial recombination National Category All three tools provided valuable phylogenetic information to enable QC of mtDNA data, but critical review of the predicted haplogroup may be required for certain applications. An important consideration when using any haplogrouping tool is that the haplogroup identified is only a prediction, particularly when based on smaller regions. Overall, the tools performed similarly, but EMPOP’s SAM2 produced more precise haplogroup predictions than AQME and HaploGrep2 across all haplogroups and regions. As an important QC aspect, the comparison between the haplogroup predictions of authentic and artificial haplotypes showed that it is possible to identify recombinant mitogenome haplotypes from two large (~8500-bp) amplicons, but the differentiation is more difficult with HVS1-2 “artificial” haplotypes. Haplogroup assignments for the CR and HVS1-2 were similar though less precise haplogroups resulted for these HVS1-2 haplotypes compared to the CR due to haplogroup-diagnostic mutations outside the queried region. There were only two differences (out of 92) between all three tools when using the mitogenome, and in these instances the haplogroups were less precise by one or two nodes. Haplogroups were determined for 92 diverse mtDNA haplotypes by the three tools based on four regions: the entire mitochondrial genome (mitogenome), control region (CR), hypervariable segment 1 and 2 (HVS1-2) regions, and HVS1 only. ![]() The Mitochondrial Haplogrouper tool of the AFDIL-QIAGEN mtDNA Expert (AQME), a plug-in for the CLC Genomics Workbench, and the web-based tools HaploGrep2 and EMPOP were included in the evaluation. This study compared three haplogrouping tools and their ability to provide proper guidance based on the predicted phylogeny. Additionally, the phylogeny guides mtDNA nomenclature, which is crucial for forensic searches. Phylogenetic information can be used to infer maternal biogeographic ancestry and provide a valuable quality control (QC) check of mitochondrial DNA (mtDNA) data, identifying errors in the data such as artificial recombination and phantom mutations. Show others and affiliations (English) Manuscript (preprint) (Other academic) Abstract
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