Enhancing the Potential of Microhaplotypes for Forensic Applications: Insights from Afghan and Somali Populations
Abstract
:1. Introduction
2. Materials and Methods
2.1. Population Samples and Extraction
2.2. Amplification
2.3. Library Building and Sequencing
2.4. Population Data Analysis and Forensic Statistical Parameters
2.5. Kinship Analyses
3. Results and Discussion
3.1. Sequencing Results
3.2. Reported Variants and Rare Alleles
3.3. Population and Forensic Parameters
3.4. Kinship Analyses
4. Conclusions
Supplementary Materials
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Acknowledgments
Conflicts of Interest
References
- Butler, J.M. Short Tandem Repeat Typing Technologies Used in Human Identity Testing. BioTechniques 2007, 43, Sii–Sv. [Google Scholar] [CrossRef]
- Thompson, R.; Zoppis, S.; McCord, B. An Overview of DNA Typing Methods for Human Identification: Past, Present, and Future. In DNA Electrophoresis Protocols for Forensic Genetics; Alonso, A., Ed.; Humana Press: Totowa, NJ, USA, 2012; pp. 3–16. ISBN 978-1-61779-461-2. [Google Scholar]
- Haddrill, P.R. Developments in Forensic DNA Analysis. Emerg. Top. Life Sci. 2021, 5, 381–393. [Google Scholar] [CrossRef]
- Oldoni, F.; Podini, D. Forensic Molecular Biomarkers for Mixture Analysis. Forensic Sci. Int. Genet. 2019, 41, 107–119. [Google Scholar] [CrossRef] [PubMed]
- Bennett, L.; Oldoni, F.; Long, K.; Cisana, S.; Madella, K.; Wootton, S.; Chang, J.; Hasegawa, R.; Lagacé, R.; Kidd, K.K.; et al. Mixture Deconvolution by Massively Parallel Sequencing of Microhaplotypes. Int. J. Legal Med. 2019, 133, 719–729. [Google Scholar] [CrossRef] [PubMed]
- Wu, R.; Li, H.; Li, R.; Peng, D.; Wang, N.; Shen, X.; Sun, H. Identification and Sequencing of 59 Highly Polymorphic Microhaplotypes for Analysis of DNA Mixtures. Int. J. Legal Med. 2021, 135, 1137–1149. [Google Scholar] [CrossRef]
- Pang, J.-B.; Rao, M.; Chen, Q.-F.; Ji, A.-Q.; Zhang, C.; Kang, K.-L.; Wu, H.; Ye, J.; Nie, S.-J.; Wang, L. A 124-Plex Microhaplotype Panel Based on Next-Generation Sequencing Developed for Forensic Applications. Sci. Rep. 2020, 10, 1945. [Google Scholar] [CrossRef]
- Staadig, A.; Tillmar, A. Evaluation of Microhaplotypes in Forensic Kinship Analysis from a Swedish Population Perspective. Int. J. Legal Med. 2021, 135, 1151–1160. [Google Scholar] [CrossRef] [PubMed]
- Kidd, K.K.; Pakstis, A.J.; Speed, W.C.; Lagace, R.; Chang, J.; Wootton, S.; Ihuegbu, N. Microhaplotype Loci Are a Powerful New Type of Forensic Marker. Forensic Sci. Int. Genet. Suppl. Ser. 2013, 4, e123–e124. [Google Scholar] [CrossRef]
- de la Puente, M.; Phillips, C.; Xavier, C.; Amigo, J.; Carracedo, A.; Parson, W.; Lareu, M.V. Building a Custom Large-Scale Panel of Novel Microhaplotypes for Forensic Identification Using MiSeq and Ion S5 Massively Parallel Sequencing Systems. Forensic Sci. Int. Genet. 2020, 45, 102213. [Google Scholar] [CrossRef]
- Turchi, C.; Melchionda, F.; Pesaresi, M.; Fattorini, P.; Tagliabracci, A. Performance of a Massive Parallel Sequencing Microhaplotypes Assay on Degraded DNA. Forensic Sci. Int. Genet. Suppl. Ser. 2019, 7, 782–783. [Google Scholar] [CrossRef]
- Chen, J.; Chen, A.; Tao, R.; Zhu, R.; Zhang, H.; You, X.; Li, C.; Zhang, S. Solution to a Case Involving the Interpretation of Trace Degraded DNA Mixtures. Int. J. Legal Med. 2024, 138, 2325–2330. [Google Scholar] [CrossRef] [PubMed]
- Chen, P.; Yin, C.; Li, Z.; Pu, Y.; Yu, Y.; Zhao, P.; Chen, D.; Liang, W.; Zhang, L.; Chen, F. Evaluation of the Microhaplotypes Panel for DNA Mixture Analyses. Forensic Sci. Int. Genet. 2018, 35, 149–155. [Google Scholar] [CrossRef] [PubMed]
- Oldoni, F.; Kidd, K.K.; Podini, D. Microhaplotypes in Forensic Genetics. Forensic Sci. Int. Genet. 2019, 38, 54–69. [Google Scholar] [CrossRef] [PubMed]
- Oldoni, F.; Bader, D.; Fantinato, C.; Wootton, S.C.; Lagacé, R.; Kidd, K.K.; Podini, D. A Sequence-Based 74plex Microhaplotype Assay for Analysis of Forensic DNA Mixtures. Forensic Sci. Int. Genet. 2020, 49, 102367. [Google Scholar] [CrossRef]
- Oldoni, F.; Yoon, L.; Wootton, S.C.; Lagacé, R.; Kidd, K.K.; Podini, D. Population Genetic Data of 74 Microhaplotypes in Four Major U.S. Population Groups. Forensic Sci. Int. Genet. 2020, 49, 102398. [Google Scholar] [CrossRef]
- Liu, J.; Su, Y.; Wen, Y.; Jiang, L.; Li, S.; Zhao, M.; Chen, X.; Wang, Z. Massively Parallel Sequencing of 74 Microhaplotypes and Forensic Characteristics in Three Chinese Sino-Tibetan Populations. Forensic Sci. Int. Genet. 2023, 66, 102905. [Google Scholar] [CrossRef]
- Tomas, C.; Rodrigues, P.; Jønck, C.G.; Barekzay, Z.; Simayijiang, H.; Pereira, V.; Børsting, C. Performance of a 74-Microhaplotype Assay in Kinship Analyses. Genes 2024, 15, 224. [Google Scholar] [CrossRef]
- Oldoni, F.; Della Rocca, C.; Podini, D. Investigation of 74 Microhaplotypes for Kinship Testing in US Populations. Forensic Sci. Int. Genet. Suppl. Ser. 2022, 8, 40–41. [Google Scholar] [CrossRef]
- Oldoni, F.; Bader, D.; Fantinato, C.; Wootton, S.C.; Lagacé, R.; Hasegawa, R.; Chang, J.; Kidd, K.; Podini, D. A Massively Parallel Sequencing Assay of Microhaplotypes for Mixture Deconvolution. Forensic Sci. Int. Genet. Suppl. Ser. 2019, 7, 522–524. [Google Scholar] [CrossRef]
- Jønck, C.G.; Børsting, C. Introduction of the Python Script MHinNGS for Analysis of Microhaplotypes. Forensic Sci. Int. Genet. Suppl. Ser. 2022, 8, 79–81. [Google Scholar] [CrossRef]
- Excoffier, L.; Lischer, H.E.L. Arlequin Suite Ver 3.5: A New Series of Programs to Perform Population Genetics Analyses under Linux and Windows. Mol. Ecol. Resour. 2010, 10, 564–567. [Google Scholar] [CrossRef] [PubMed]
- Gouy, A.; Zieger, M. STRAF-A Convenient Online Tool for STR Data Evaluation in Forensic Genetics. Forensic Sci. Int. Genet. 2017, 30, 148–151. [Google Scholar] [CrossRef]
- Crow, J.F.; Kimura, M. An Introduction To Population Genetics Theory; Harper & Row: New York, NY, USA, 1970. [Google Scholar]
- Goudet, J. Hierfstat, a package for R to compute and test hierarchical F-statistics. Mol. Ecol. Notes 2005, 5, 184–186. [Google Scholar] [CrossRef]
- Patterson, N.; Price, A.L.; Reich, D. Population Structure and Eigenanalysis. PLoS Genet. 2006, 2, e190. [Google Scholar] [CrossRef]
- Pritchard, J.K.; Stephens, M.; Donnelly, P. Inference of Population Structure Using Multilocus Genotype Data. Genetics 2000, 155, 945–959. [Google Scholar] [CrossRef]
- 1000 Genomes Project Consortium; Auton, A.; Brooks, L.D.; Durbin, R.M.; Garrison, E.P.; Kang, H.M.; Korbel, J.O.; Marchini, J.L.; McCarthy, S.; McVean, G.A.; et al. A Global Reference for Human Genetic Variation. Nature 2015, 526, 68–74. [Google Scholar] [CrossRef]
- Kling, D.; Tillmar, A.O.; Egeland, T. Familias 3—Extensions and New Functionality. Forensic Sci. Int. Genet. 2014, 13, 121–127. [Google Scholar] [CrossRef]
- Machado, P.; Gusmão, L.; Conde-Sousa, E.; Pinto, N. The Influence of the Different Mutation Models in Kinship Evaluation. Forensic Sci. Int. Genet. Suppl. Ser. 2017, 6, e255–e256. [Google Scholar] [CrossRef]
- Kitayama, T.; Kiesler, K.M.; Fukagawa, T.; Watahiki, H.; Mita, Y.; Fujii, K.; Sekiguchi, K.; Vallone, P.M.; Mizuno, N. Massively Parallel Sequencing Data of 31 Autosomal STR Loci Obtained Using the Precision ID GlobalFiler NGS STR Panel v2 for 82 Japanese Population Samples. Leg. Med. 2022, 58, 102082. [Google Scholar] [CrossRef]
- Aguilar-Velázquez, J.A.; Duran-Salazar, M.Á.; Córdoba-Mercado, M.F.; Coronado-Avila, C.E.; Salas-Salas, O.; Martinez-Cortés, G.; Casals, F.; Calafell, F.; Ramos-González, B.; Rangel-Villalobos, H. Characterization of 58 STRs and 94 SNPs with the ForenSeqTM DNA Signature Prep Kit in Mexican-Mestizos from the Monterrey City (Northeast, Mexico). Mol. Biol. Rep. 2022, 49, 7601–7609. [Google Scholar] [CrossRef]
- Kidd, K.K.; Speed, W.C. Criteria for Selecting Microhaplotypes: Mixture Detection and Deconvolution. Investig. Genet. 2015, 6, 1. [Google Scholar] [CrossRef] [PubMed]
- Moltke, I.; Fumagalli, M.; Korneliussen, T.S.; Crawford, J.E.; Bjerregaard, P.; Jørgensen, M.E.; Grarup, N.; Gulløv, H.C.; Linneberg, A.; Pedersen, O.; et al. Uncovering the Genetic History of the Present-Day Greenlandic Population. Am. J. Hum. Genet. 2015, 96, 54–69. [Google Scholar] [CrossRef] [PubMed]
- Kidd, K.K.; Speed, W.C.; Pakstis, A.J.; Podini, D.S.; Lagacé, R.; Chang, J.; Wootton, S.; Haigh, E.; Soundararajan, U. Evaluating 130 Microhaplotypes across a Global Set of 83 Populations. Forensic Sci. Int. Genet. 2017, 29, 29–37. [Google Scholar] [CrossRef]
- Kidd, K.K.; Pakstis, A.J.; Speed, W.C.; Lagace, R.; Wootton, S.; Chang, J. Selecting Microhaplotypes Optimized for Different Purposes. Electrophoresis 2018, 39, 2815–2823. [Google Scholar] [CrossRef] [PubMed]
- Qu, S.; Schneider, P.M.; Lagacé, R.; Neis, M.; Liang, W.; Zhang, L. The Population Genetics Characteristics of Ion AmpliSeqTM MH-74 Plex Microhaplotype Research Panel. Forensic Sci. Int. Genet. Suppl. Ser. 2022, 8, 297–299. [Google Scholar] [CrossRef]
- Pereira, V.; Santangelo, R.; Børsting, C.; Tvedebrink, T.; Almeida, A.P.F.; Carvalho, E.F.; Morling, N.; Gusmão, L. Evaluation of the Precision of Ancestry Inferences in South American Admixed Populations. Front. Genet. 2020, 11, 966. [Google Scholar] [CrossRef]
- Pereira, V.; Mogensen, H.S.; Børsting, C.; Morling, N. Evaluation of the Precision ID Ancestry Panel for Crime Case Work: A SNP Typing Assay Developed for Typing of 165 Ancestral Informative Markers. Forensic Sci. Int. Genet. 2017, 28, 138–145. [Google Scholar] [CrossRef]
- Martínez-Cruz, B.; Vitalis, R.; Ségurel, L.; Austerlitz, F.; Georges, M.; Théry, S.; Quintana-Murci, L.; Hegay, T.; Aldashev, A.; Nasyrova, F.; et al. In the Heartland of Eurasia: The Multilocus Genetic Landscape of Central Asian Populations. Eur. J. Hum. Genet. 2011, 19, 216–223. [Google Scholar] [CrossRef]
- Di Cristofaro, J.; Pennarun, E.; Mazières, S.; Myres, N.M.; Lin, A.A.; Temori, S.A.; Metspalu, M.; Metspalu, E.; Witzel, M.; King, R.J.; et al. Afghan Hindu Kush: Where Eurasian Sub-Continent Gene Flows Converge. PLoS ONE 2013, 8, e76748. [Google Scholar] [CrossRef]
- Zhabagin, M.; Balanovska, E.; Sabitov, Z.; Kuznetsova, M.; Agdzhoyan, A.; Balaganskaya, O.; Chukhryaeva, M.; Markina, N.; Romanov, A.; Skhalyakho, R.; et al. The Connection of the Genetic, Cultural and Geographic Landscapes of Transoxiana. Sci. Rep. 2017, 7, 3085. [Google Scholar] [CrossRef]
- Haber, M.; Platt, D.E.; Ashrafian Bonab, M.; Youhanna, S.C.; Soria-Hernanz, D.F.; Martínez-Cruz, B.; Douaihy, B.; Ghassibe-Sabbagh, M.; Rafatpanah, H.; Ghanbari, M.; et al. Afghanistan’s Ethnic Groups Share a Y-Chromosomal Heritage Structured by Historical Events. PLoS ONE 2012, 7, e34288. [Google Scholar] [CrossRef]
- Hodgson, J.A.; Mulligan, C.J.; Al-Meeri, A.; Raaum, R.L. Early Back-to-Africa Migration into the Horn of Africa. PLoS Genet. 2014, 10, e1004393. [Google Scholar] [CrossRef]
- Du, Q.; Ma, G.; Lu, C.; Wang, Q.; Fu, L.; Cong, B.; Li, S. Development and Evaluation of a Novel Panel Containing 188 Microhaplotypes for 2nd-Degree Kinship Testing in the Hebei Han Population. Forensic Sci. Int. Genet. 2023, 65, 102855. [Google Scholar] [CrossRef]
- Wu, R.; Chen, H.; Li, R.; Zang, Y.; Shen, X.; Hao, B.; Wang, Q.; Sun, H. Pairwise Kinship Testing with Microhaplotypes: Can Advancements Be Made in Kinship Inference with These Markers? Forensic Sci. Int. 2021, 325, 110875. [Google Scholar] [CrossRef]
- Xue, J.; Tan, M.; Zhang, R.; Chen, D.; Liu, G.; Zheng, Y.; Wu, Q.; Xiao, Y.; Liao, M.; Qu, S.; et al. Evaluation of Microhaplotype Panels for Complex Kinship Analysis Using Massively Parallel Sequencing. Forensic Sci. Int. Genet. 2023, 65, 102887. [Google Scholar] [CrossRef]
- Qu, N.; Lin, S.; Gao, Y.; Liang, H.; Zhao, H.; Ou, X. A Microhap Panel for Kinship Analysis through Massively Parallel Sequencing Technology. Electrophoresis 2020, 41, 246–253. [Google Scholar] [CrossRef]
- Børsting, C.; Sanchez, J.J.; Hansen, H.E.; Hansen, A.J.; Bruun, H.Q.; Morling, N. Performance of the SNPforID 52 SNP-Plex Assay in Paternity Testing. Forensic Sci. Int. Genet. 2008, 2, 292–300. [Google Scholar] [CrossRef]
- Zhang, Q.; Wang, X.; Cheng, P.; Yang, S.; Li, W.; Zhou, Z.; Wang, S. Complex Kinship Analysis with a Combination of STRs, SNPs, and Indels. Forensic Sci. Int. Genet. 2022, 61, 102749. [Google Scholar] [CrossRef]
- Hussing, C.; Kampmann, M.-L.; Mogensen, H.S.; Børsting, C.; Morling, N. Quantification of Massively Parallel Sequencing Libraries—A Comparative Study of Eight Methods. Sci. Rep. 2018, 8, 1110. [Google Scholar] [CrossRef]
- Li, R.; Li, H.; Peng, D.; Hao, B.; Wang, Z.; Huang, E.; Wu, R.; Sun, H. Improved Pairwise Kinship Analysis Using Massively Parallel Sequencing. Forensic Sci. Int. Genet. 2019, 38, 77–85. [Google Scholar] [CrossRef] [PubMed]
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Rodrigues, P.; Pinto, N.; Otterlund, T.; Jønck, C.G.; Prata, M.J.; Børsting, C.; Pereira, V. Enhancing the Potential of Microhaplotypes for Forensic Applications: Insights from Afghan and Somali Populations. Genes 2025, 16, 532. https://doi.org/10.3390/genes16050532
Rodrigues P, Pinto N, Otterlund T, Jønck CG, Prata MJ, Børsting C, Pereira V. Enhancing the Potential of Microhaplotypes for Forensic Applications: Insights from Afghan and Somali Populations. Genes. 2025; 16(5):532. https://doi.org/10.3390/genes16050532
Chicago/Turabian StyleRodrigues, Pedro, Nádia Pinto, Tess Otterlund, Carina G. Jønck, Maria João Prata, Claus Børsting, and Vania Pereira. 2025. "Enhancing the Potential of Microhaplotypes for Forensic Applications: Insights from Afghan and Somali Populations" Genes 16, no. 5: 532. https://doi.org/10.3390/genes16050532
APA StyleRodrigues, P., Pinto, N., Otterlund, T., Jønck, C. G., Prata, M. J., Børsting, C., & Pereira, V. (2025). Enhancing the Potential of Microhaplotypes for Forensic Applications: Insights from Afghan and Somali Populations. Genes, 16(5), 532. https://doi.org/10.3390/genes16050532