New Insights Into Mitochondrial DNA Reconstruction and Variant Detection in Ancient Samples
Ancient DNA (aDNA) studies are frequently focused on the analysis of the mitochondrial DNA (mtDNA), which is much more abundant than the nuclear genome, hence can be better retrieved from ancient remains. However, postmortem DNA damage and contamination make the data analysis difficult because of DN...
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| Format: | Article |
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Frontiers Media S.A.
2021-02-01
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| Series: | Frontiers in Genetics |
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| Online Access: | https://www.frontiersin.org/articles/10.3389/fgene.2021.619950/full |
| _version_ | 1819054138660487168 |
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| author | Maria Angela Diroma Alessandra Modi Martina Lari Luca Sineo David Caramelli Stefania Vai |
| author_facet | Maria Angela Diroma Alessandra Modi Martina Lari Luca Sineo David Caramelli Stefania Vai |
| author_sort | Maria Angela Diroma |
| collection | DOAJ |
| description | Ancient DNA (aDNA) studies are frequently focused on the analysis of the mitochondrial DNA (mtDNA), which is much more abundant than the nuclear genome, hence can be better retrieved from ancient remains. However, postmortem DNA damage and contamination make the data analysis difficult because of DNA fragmentation and nucleotide alterations. In this regard, the assessment of the heteroplasmic fraction in ancient mtDNA has always been considered an unachievable goal due to the complexity in distinguishing true endogenous variants from artifacts. We implemented and applied a computational pipeline for mtDNA analysis to a dataset of 30 ancient human samples from an Iron Age necropolis in Polizzello (Sicily, Italy). The pipeline includes several modules from well-established tools for aDNA analysis and a recently released variant caller, which was specifically conceived for mtDNA, applied for the first time to aDNA data. Through a fine-tuned filtering on variant allele sequencing features, we were able to accurately reconstruct nearly complete (>88%) mtDNA genome for almost all the analyzed samples (27 out of 30), depending on the degree of preservation and the sequencing throughput, and to get a reliable set of variants allowing haplogroup prediction. Additionally, we provide guidelines to deal with possible artifact sources, including nuclear mitochondrial sequence (NumtS) contamination, an often-neglected issue in ancient mtDNA surveys. Potential heteroplasmy levels were also estimated, although most variants were likely homoplasmic, and validated by data simulations, proving that new sequencing technologies and software are sensitive enough to detect partially mutated sites in ancient genomes and discriminate true variants from artifacts. A thorough functional annotation of detected and filtered mtDNA variants was also performed for a comprehensive evaluation of these ancient samples. |
| first_indexed | 2024-12-21T12:46:52Z |
| format | Article |
| id | doaj.art-419a1284e2de45d8b492483bcf2ef86a |
| institution | Directory Open Access Journal |
| issn | 1664-8021 |
| language | English |
| last_indexed | 2024-12-21T12:46:52Z |
| publishDate | 2021-02-01 |
| publisher | Frontiers Media S.A. |
| record_format | Article |
| series | Frontiers in Genetics |
| spelling | doaj.art-419a1284e2de45d8b492483bcf2ef86a2022-12-21T19:03:36ZengFrontiers Media S.A.Frontiers in Genetics1664-80212021-02-011210.3389/fgene.2021.619950619950New Insights Into Mitochondrial DNA Reconstruction and Variant Detection in Ancient SamplesMaria Angela Diroma0Alessandra Modi1Martina Lari2Luca Sineo3David Caramelli4Stefania Vai5Dipartimento di Biologia, Università degli Studi di Firenze, Florence, ItalyDipartimento di Biologia, Università degli Studi di Firenze, Florence, ItalyDipartimento di Biologia, Università degli Studi di Firenze, Florence, ItalyDipartimento di Scienze e Tecnologie Biologiche, Chimiche e Farmaceutiche, Università degli Studi di Palermo, Palermo, ItalyDipartimento di Biologia, Università degli Studi di Firenze, Florence, ItalyDipartimento di Biologia, Università degli Studi di Firenze, Florence, ItalyAncient DNA (aDNA) studies are frequently focused on the analysis of the mitochondrial DNA (mtDNA), which is much more abundant than the nuclear genome, hence can be better retrieved from ancient remains. However, postmortem DNA damage and contamination make the data analysis difficult because of DNA fragmentation and nucleotide alterations. In this regard, the assessment of the heteroplasmic fraction in ancient mtDNA has always been considered an unachievable goal due to the complexity in distinguishing true endogenous variants from artifacts. We implemented and applied a computational pipeline for mtDNA analysis to a dataset of 30 ancient human samples from an Iron Age necropolis in Polizzello (Sicily, Italy). The pipeline includes several modules from well-established tools for aDNA analysis and a recently released variant caller, which was specifically conceived for mtDNA, applied for the first time to aDNA data. Through a fine-tuned filtering on variant allele sequencing features, we were able to accurately reconstruct nearly complete (>88%) mtDNA genome for almost all the analyzed samples (27 out of 30), depending on the degree of preservation and the sequencing throughput, and to get a reliable set of variants allowing haplogroup prediction. Additionally, we provide guidelines to deal with possible artifact sources, including nuclear mitochondrial sequence (NumtS) contamination, an often-neglected issue in ancient mtDNA surveys. Potential heteroplasmy levels were also estimated, although most variants were likely homoplasmic, and validated by data simulations, proving that new sequencing technologies and software are sensitive enough to detect partially mutated sites in ancient genomes and discriminate true variants from artifacts. A thorough functional annotation of detected and filtered mtDNA variants was also performed for a comprehensive evaluation of these ancient samples.https://www.frontiersin.org/articles/10.3389/fgene.2021.619950/fullancient DNAmitochondrial DNANUMTsheteroplasmyvariant detection |
| spellingShingle | Maria Angela Diroma Alessandra Modi Martina Lari Luca Sineo David Caramelli Stefania Vai New Insights Into Mitochondrial DNA Reconstruction and Variant Detection in Ancient Samples Frontiers in Genetics ancient DNA mitochondrial DNA NUMTs heteroplasmy variant detection |
| title | New Insights Into Mitochondrial DNA Reconstruction and Variant Detection in Ancient Samples |
| title_full | New Insights Into Mitochondrial DNA Reconstruction and Variant Detection in Ancient Samples |
| title_fullStr | New Insights Into Mitochondrial DNA Reconstruction and Variant Detection in Ancient Samples |
| title_full_unstemmed | New Insights Into Mitochondrial DNA Reconstruction and Variant Detection in Ancient Samples |
| title_short | New Insights Into Mitochondrial DNA Reconstruction and Variant Detection in Ancient Samples |
| title_sort | new insights into mitochondrial dna reconstruction and variant detection in ancient samples |
| topic | ancient DNA mitochondrial DNA NUMTs heteroplasmy variant detection |
| url | https://www.frontiersin.org/articles/10.3389/fgene.2021.619950/full |
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