Development and validation of a STR and SNP multiplex detection system (88 STRs and 348 SNPs) using massively parallel sequencing.
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| Titel: | Development and validation of a STR and SNP multiplex detection system (88 STRs and 348 SNPs) using massively parallel sequencing. |
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| Autoren: | Lu Y; Institute of Forensic Science, Fudan University, Shanghai, 200032, China., Yang F; Key Laboratory of Forensic Evidence and Science Technology, Institute of Forensic Science, Ministry of Public Security, Shanghai, 200042, China., Liu Y; Key Laboratory of Forensic Evidence and Science Technology, Institute of Forensic Science, Ministry of Public Security, Shanghai, 200042, China.; Ministry of Education's Key Laboratory of Contemporary Anthropology, School of Life Sciences, Fudan University, Shanghai, 200438, China., Xuan Y; Institute of Forensic Science, Fudan University, Shanghai, 200032, China., Zhao J; Institute of Forensic Science, Fudan University, Shanghai, 200032, China., Liao M; Institute of Forensic Science, Fudan University, Shanghai, 200032, China., Xing Y; Institute of Forensic Science, Fudan University, Shanghai, 200032, China., Liu S; Institute of Forensic Science, Fudan University, Shanghai, 200032, China., Zhang S; Institute of Forensic Science, Fudan University, Shanghai, 200032, China. zhang_suhua@fudan.edu.cn., Chen A; Institute of Forensic Science, Fudan University, Shanghai, 200032, China. anqi_chen@fudan.edu.cn. |
| Quelle: | International journal of legal medicine [Int J Legal Med] 2025 Dec 18. Date of Electronic Publication: 2025 Dec 18. |
| Publication Model: | Ahead of Print |
| Publikationsart: | Journal Article |
| Sprache: | English |
| Info zur Zeitschrift: | Publisher: Springer International Country of Publication: Germany NLM ID: 9101456 Publication Model: Print-Electronic Cited Medium: Internet ISSN: 1437-1596 (Electronic) Linking ISSN: 09379827 NLM ISO Abbreviation: Int J Legal Med Subsets: MEDLINE |
| Imprint Name(s): | Original Publication: Heidelberg, FRG : Springer International, c1990- |
| Abstract: | Short tandem repeats (STRs) have long been the gold standard in forensic DNA analysis, while single nucleotide polymorphisms (SNPs) have increasingly emerged as valuable complementary markers. The advent of massively parallel sequencing (MPS) technology has significantly enhanced the detection resolution of both STRs and SNPs, providing novel approaches for forensic human identification. In this study, we developed and validated a comprehensive multiplex detection system utilizing MPS technology, which enabled the concurrent analyses of 88 STRs and 348 SNPs. Following the SWGDAM guidelines, validation studies were conducted to assess the forensic applicability of the system. Sensitivity study revealed that 100% STR profiles could be obtained from ≥ 0.125 ng, and 100% SNP profiles were achieved with ≥ 0.0625 ng. The panel showed high accuracy, achieving a 99.78% concordance rate with conventional capillary electrophoresis (CE) DNA profiles. The system exhibited robust performance in the presence of four common PCR inhibitors (including humic acid, melanin, indigo, and hematin). Additionally, 99.42% of STR genotypes and 100% of SNP genotypes were successfully obtained from degraded DNA shorter than 500 bp. Furthermore, across gender combinations in mixed samples, both STR and SNP markers performed robustly at 1:1 and 1:3 ratios. At extreme proportions, male-female mixtures (99:1) enable the detection of scarce minor contributors at Y-STR loci. For SNP loci, the ACR value serves as a key evaluative metric for distinguishing mixtures. ROC analysis of these SNP data yielded an AUC of 0.909. Population genetics analyses revealed that the cumulative power of discrimination (CPD) and cumulative probability of exclusion (CPE) both exceeded 0.999999, highlighting its extensive applicability and practical value in forensic genetics. This comprehensive validation study confirmed that the multiplex panel provided high sensitivity, accuracy, and reliability, representing a significant advancement in forensic DNA analysis methodology. (© 2025. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.) |
| Competing Interests: | Declarations. Ethics approval: The study was approved by the Ethics Committee of the Academy of Forensic Science. Ministry of Justice, P.R. China (NO.2022-11). Conflict of interest: The authors declare no competing interests. |
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| Grant Information: | 2024YFC3306702 National Key R&D Program of China; 82302124 National Natural Science Foundation of China |
| Contributed Indexing: | Keywords: Forensic genetics; Massively parallel sequencing (MPS); SWGDAM validation; Short tandem repeat (STR); Single nucleotide polymorphism (SNP) |
| Entry Date(s): | Date Created: 20251217 Latest Revision: 20251217 |
| Update Code: | 20251218 |
| DOI: | 10.1007/s00414-025-03682-0 |
| PMID: | 41407971 |
| Datenbank: | MEDLINE |
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Nájsť tento článok vo Web of Science | Abstract: | Short tandem repeats (STRs) have long been the gold standard in forensic DNA analysis, while single nucleotide polymorphisms (SNPs) have increasingly emerged as valuable complementary markers. The advent of massively parallel sequencing (MPS) technology has significantly enhanced the detection resolution of both STRs and SNPs, providing novel approaches for forensic human identification. In this study, we developed and validated a comprehensive multiplex detection system utilizing MPS technology, which enabled the concurrent analyses of 88 STRs and 348 SNPs. Following the SWGDAM guidelines, validation studies were conducted to assess the forensic applicability of the system. Sensitivity study revealed that 100% STR profiles could be obtained from ≥ 0.125 ng, and 100% SNP profiles were achieved with ≥ 0.0625 ng. The panel showed high accuracy, achieving a 99.78% concordance rate with conventional capillary electrophoresis (CE) DNA profiles. The system exhibited robust performance in the presence of four common PCR inhibitors (including humic acid, melanin, indigo, and hematin). Additionally, 99.42% of STR genotypes and 100% of SNP genotypes were successfully obtained from degraded DNA shorter than 500 bp. Furthermore, across gender combinations in mixed samples, both STR and SNP markers performed robustly at 1:1 and 1:3 ratios. At extreme proportions, male-female mixtures (99:1) enable the detection of scarce minor contributors at Y-STR loci. For SNP loci, the ACR value serves as a key evaluative metric for distinguishing mixtures. ROC analysis of these SNP data yielded an AUC of 0.909. Population genetics analyses revealed that the cumulative power of discrimination (CPD) and cumulative probability of exclusion (CPE) both exceeded 0.999999, highlighting its extensive applicability and practical value in forensic genetics. This comprehensive validation study confirmed that the multiplex panel provided high sensitivity, accuracy, and reliability, representing a significant advancement in forensic DNA analysis methodology.<br /> (© 2025. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.) |
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| ISSN: | 1437-1596 |
| DOI: | 10.1007/s00414-025-03682-0 |