Abstract
Massively parallel sequencing (MPS) has emerged as a promising technology for targeting multiple genetic loci simultaneously in forensic genetics. Here, a novel 193-plex panel was designed to target 28 A-STRs, 41 Y-STRs, 21 X-STRs, 3 sex-identified loci, and 100 A-SNPs by employing a single-end 400 bp sequencing strategy on the MGISEQ-2000™ platform. In the present study, a series of validations and sequencing of 1642 population samples were performed to evaluate the overall performance of the MPS-based panel and its practicality in forensic application according to the SWGDAM guidelines. In general, the 193-plex markers in our panel showed good performance in terms of species specificity, stability, and repeatability. Compared to commercial kits, this panel achieved 100% concordance for standard gDNA and 99.87% concordance for 14,560 population genotypes. Moreover, this panel detected 100% of the loci from 0.5 ng of DNA template and all unique alleles at a 1:4 DNA mixture ratio (0.2 ng minor contributor), and the applicability of the proposed approach for tracing and degrading DNA was further supported by case samples. In addition, several forensic parameters of STRs and SNPs were calculated in a population study. High CPE and CPD values greater than 0.9999999 were clearly demonstrated and these results could be useful references for the application of this panel in individual identification and paternity testing. Overall, this 193-plex MPS panel has been shown to be a reliable, repeatable, robust, inexpensive, and powerful tool sufficient for forensic practice.
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Data availability
The datasets analyzed during the current study are available from the corresponding author upon reasonable request.
References
Ballard D, Winkler-Galicki J, Wesoly J (2020) Massive parallel sequencing in forensics: advantages, issues, technicalities, and prospects. Int J Legal Med 134(4):1291–1303. https://doi.org/10.1007/s00414-020-02294-0
Børsting C, Morling N (2015) Next generation sequencing and its applications in forensic genetics. Forensic Sci Int Genet 18:78–89. https://doi.org/10.1016/j.fsigen.2015.02.002
Børsting C, Mogensen HS, Morling N (2013) Forensic genetic SNP typing of low-template DNA and highly degraded DNA from crime case samples. Forensic Sci Int Genet 7(3):345–352. https://doi.org/10.1016/j.fsigen.2013.02.004
Butler JM (2006) Genetics and genomics of core short tandem repeat loci used in human identity testing. J Forensic Sci 51(2):253–265. https://doi.org/10.1111/j.1556-4029.2006.00046.x
Butler JM, Forensic DNA (2005) Typing: biology, technology, and genetics of STR markers, 2nd edn. Elsevier Academic Press, USA
Clayton TM, Whitaker JP, Maguire CN (1995) Identification of bodies from the scene of a mass disaster using DNA amplification of short tandem repeat (STR) loci. Forensic Sci Int 76(1):7–15. https://doi.org/10.1016/0379-0738(95)01787-9
de Knijf P (2019) From next generation sequencing to now generation sequencing in forensics. Forensic Sci Int Genet 38:175–180. https://doi.org/10.1016/j.fsigen.2018.10.017
Deng C, Song F, Li J et al (2017) Forensic parameters of 19 X-STR polymorphisms in two Chinese populations. Int J Legal Med 131(4):975–977. https://doi.org/10.1007/s00414-017-1538-1
Desmarais D, Zhong Y, Chakraborty R et al (1998) Development of a highly polymorphic STR marker for identity testing purposes at the human androgen receptor gene (HUMARA). J Forensic Sci 43(5):1046–1049
Elwick K, Zeng X, King J et al (2018) Comparative tolerance of two massively parallel sequencing systems to common PCR inhibitors. Int J Legal Med 132(4):983–995. https://doi.org/10.1007/s00414-017-1693-4
Fan H, Zeng Y, Wu W, Liu H, Xu Q, Du W, Hao H, Liu C, Ren W, Wu W, Chen L, Liu C (2021) The Y-STR landscape of coastal southeastern Han: forensic characteristics, haplotype analyses, mutation rates, and population genetics. Electrophoresis 42(16):1578–1593. https://doi.org/10.1002/elps.202100037
Fan H, Wang L, Liu C, Lu X, Xu X, Ru K, Qiu P, Liu C, Wen SQ (2022) Development and validation of a novel 133-plex forensic STR panel (52 STRs and 81 Y-STRs) using single-end 400 bp massive parallel sequencing. Int J Legal Med 136(2):447–464. https://doi.org/10.1007/s00414-021-02738-1
Fondevila M, Phillips C, Naveran N et al (2008) Case report: Identification of skeletal remains using short-amplicon marker analysis of severely degraded DNA extracted from a decomposed and charred femur. Forensic Sci Int Genet 2(3):212–218. https://doi.org/10.1016/j.fsigen.2008.02.005
Gettings KB, Kiesler KM, Faith SA, Montano E, Baker CH, Young BA, Guerrieri RA, Vallone PM (2016) Sequence variation of 22 autosomal STR loci detected by next generation sequencing. Forensic Sci Int Genet 21:15–21. https://doi.org/10.1016/j.fsigen.2015.11.005
Gill P, Jeffreys AJ, Werrett DJ (1985) Forensic application of DNA ‘fingerprints.’ Nature 318(6046):577–579. https://doi.org/10.1038/318577a0
Goodwin S, McPherson JD, McCombie WR (2016) Coming of age: ten years of next-generation sequencing technologies. Nat Rev Genet 17(6):333–351. https://doi.org/10.1038/nrg.2016.49
Gouy A, Zieger M (2017) STRAF—a convenient online tool for STR data evaluation in forensic genetics. Forensic Sci Int Genet 30:148–151. https://doi.org/10.1016/j.fsigen
Grandell I, Samara R, Tillmar AO (2016) A SNP panel for identity and kinship testing using massive parallel sequencing. Int J Legal Med 130(4):905–914. https://doi.org/10.1007/s00414-016-1341-4
Guo F, Yu J, Zhang L, Li J (2017) Massively parallel sequencing of forensic STRs and SNPs using the Illumina® ForenSeq™ DNA signature prep kit on the MiSeq FGx™ forensic genomics system. Forensic Sci Int Genet 31:135–148. https://doi.org/10.1016/j.fsigen.2017.09.003
Hampikian G, West E, Akselrod O (2011) The genetics of innocence: analysis of 194 U.S. DNA exonerations. Annu Rev Genomics Hum Genet 12:97–120. https://doi.org/10.1146/annurev-genom-082509-141715
Hares DR (2015) Selection and implementation of expanded CODIS core loci in the United States. Forensic Sci Int Genet 17:33–34. https://doi.org/10.1016/j.fsigen.2015.03.006
Hosoi E (2008) Biological and clinical aspects of ABO blood group system. J Med Investig 55(3–4):174–182. https://doi.org/10.2152/jmi.55.174
Huston KA (1998) Statistical analysis of STR data. Profiles DNA 1:14–15
Imam J, Reyaz R, Singh RS et al (2018) Genomic portrait of population of Jharkhand, India, drawn with 15 autosomal STRs and 17 Y-STRs. Int J Legal Med 132(1):139–140. https://doi.org/10.1007/s00414-017-1610-x
Jeffreys AJ, Wilson V, Thein SL (1985) Hypervariable‘minisatellite’regions in human DNA. Nature 314(6006):67–73. https://doi.org/10.1038/314067a0
Jobling MA, Gill P (2004) Encoded evidence: DNA in forensic analysis. Nat Rev Genet 5(10):739–751. https://doi.org/10.1038/nrg1455
Keerti A, Ninave S (2022) DNA fingerprinting: use of autosomal short tandem repeats in forensic DNA typing. Cureus 14(10):e30210. https://doi.org/10.7759/cureus.30210
Li R, Shen X, Chen H, Peng D, Wu R, Sun H (2021) Developmental validation of the MGIEasy signature identification library prep kit, an all-in-one multiplex system for forensic applications. Int J Legal Med 135:739–753. https://doi.org/10.1007/s00414-021-02507-0
Lin H, Ye Q, Tang P, Mo T, Yu X, Tang J (2020) Analyzing genetic polymorphism and mutation of 44 Y-STRs in a Chinese Han population of Southern China. Leg Med (Tokyo) 42:101643. https://doi.org/10.1016/j.legalmed.2019.101643
Liu QL, Li ZD, Li CT, Zhao H, Wu YD, Li Q, Lu DJ (2013) X chromosomal recombination—a family study analyzing 26 X-STR Loci in Chinese Han three-generation pedigrees. Electrophoresis 34(20–21):3016–3022. https://doi.org/10.1002/elps.201300204
Liu J, Wang Z, He G et al (2018) Massively parallel sequencing of 124 SNPs included in the precision ID identity panel in three East Asian minority ethnicities. Forensic Sci Int Genet 35:141–148. https://doi.org/10.1016/j.fsigen.2018.05.002
Miao X, Shen Y, Gong X et al (2021) A novel forensic panel of 186-plex SNPs and 123-plex STR loci based on massively parallel sequencing. Int J Legal Med 135(3):709–718. https://doi.org/10.1007/s00414-020-02403-z
Mujibi FDN, Rao J, Agaba M et al (2019) Performance evaluation of highly admixed Tanzanian smallholder dairy cattle using SNP derived kinship matrix. Front Genet 10:375. https://doi.org/10.3389/fgene.2019.00375
Phillips C, García-Magariños M, Salas A, Carracedo A, Lareu MV (2012) SNPs as supplements in simple kinship analysis or as core markers in distant pairwise relationship tests: when do SNPs add value or replace well-established and powerful STR tests? Transfus Med Hemother 39(3):202–210. https://doi.org/10.1159/000338857
Pontes L, Sousa JC, Medeiros R (2017) SNPs and STRs in forensic medicine. A strategy for kinship evaluation. Arch Med Sadowej Kryminol 67(3):226–240. https://doi.org/10.5114/amsik.2017.73194
Ralf A, Kayser M (2021) Investigative DNA analysis of two-person mixed crime scene trace in a murder case. Forensic Sci Int Genet 54:102557. https://doi.org/10.1016/j.fsigen.2021.102557
Ren ZL, Zhang JR, Zhang XM, Liu X, Lin YF, Bai H, Wang MC, Cheng F, Liu JD, Li P, Kong L, Bo XC, Wang SQ, Ni M, Yan JW (2021) Forensic nanopore sequencing of STRs and SNPs using Verogen’s ForenSeq DNA signature prep kit and MinION. Int J Legal Med 135(5):1685–1693. https://doi.org/10.1007/s00414-021-02604-0
Shewale JG, Qi L, Calandro LM (2012) Principles, practice, and evolution of capillary electrophoresis as a tool for forensic DNA analysis. Forensic Sci Rev 24(2):79–100. https://doi.org/10.1201/b15361-11
Shimada I, Rand S, Brinkmann B et al (2002) Kurdish population data for 11 STR loci (ACTBP2, CSF1PO, FGA, TH01, TPOX, vWA, D3S1358, D5S818, D7S820, D13S317 and D21S11). Int J Legal Med 116(5):301–303. https://doi.org/10.1007/s00414-002-0294-y
Song F, Wei X, Zhou C, Wang S, Deng C, Liao M, Luo H (2022) Resolving the recombination pattern of 38 X-STRs from Chinese Han three-generation pedigrees. Leg Med (Tokyo) 59:102135. https://doi.org/10.1016/j.legalmed.2022.102135
Sun S, Liu Y, Li J et al (2020) Development and application of a nonbinary SNP-based microhaplotype panel for paternity testing involving close relatives. Forensic Sci Int Genet 46:102255. https://doi.org/10.1016/j.fsigen.2020.102255
Tao R, Qi W, Chen C, Zhang J, Yang Z, Song W, Zhang S, Li C (2019) Pilot study for forensic evaluations of the precision ID GlobalFiler™ NGS STR panel v2 with the ion S5™ system. Forensic Sci Int Genet 43:102147. https://doi.org/10.1016/j.fsigen.2019.102147
The Relationship Identification and Implementation Specification for Biological Full Sibling (SF/T 0117-2021). http://www.ssfjd.com/Info/Info2.aspx?D=3563
Tillmar A, Grandell I, Montelius K (2018) DNA identification of compromised samples with massive parallel sequencing. Forensic Sci Res 4(4):331–336. https://doi.org/10.1080/20961790.2018.1509186
Urquhart A, Kimpton CP, Downes TJ et al (1994) Variation in short tandem repeat sequences—a survey of twelve microsatellite loci for use as forensic identification markers. Int J Legal Med 107(1):13–20. https://doi.org/10.1007/BF01247268
Wang Z, Zhou D, Wang H et al (2017) Massively parallel sequencing of 32 forensic markers using the precision id GlobalFiler™ NGS STR panel and the ion PGM™ system. Forensic Sci Int Genet 31:126–134. https://doi.org/10.1016/j.fsigen.2017.09.004
Wang J, Wu J, Sun Q et al (2023) Extensive genetic admixture between Tai-Kadai-speaking people and their neighbours in the northeastern region of the Yungui Plateau inferred from genome-wide variations. BMC Genomics 24(1):317. https://doi.org/10.1186/s12864-023-09412-3
Waples RK, Hauptmann AL, Seiding I et al (2021) The genetic history of Greenlandic-European contact. Curr Biol 31(10):2214-2219.e4. https://doi.org/10.1016/j.cub.2021.02.041
Xavier C, Parson W (2017) Evaluation of the Illumina ForenSeq™ DNA Signature Prep Kit—MPS forensic application for the MiSeq FGx™ benchtop sequencer. Forensic Sci Int Genet 28:188–194. https://doi.org/10.1016/j.fsigen.2017.02.018
Xu H, Wang CC, Shrestha R et al (2015) Inferring population structure and demographic history using Y-STR data from worldwide populations. Mol Gen Genomics 290(1):141–150. https://doi.org/10.1007/s00438-014-0903-8
Zhang J, Zhang J, Tao R et al (2020) A newly devised multiplex assay of novel polymorphic non-CODIS STRs as a valuable tool for forensic application. Forensic Sci Int Genet 48:102341. https://doi.org/10.1016/j.fsigen.2020.102341
Zhou Z, Shao C, Xie J et al (2020) Genetic polymorphism and phylogenetic analyses of 21 non-CODIS STR loci in a Chinese Han population from Shanghai. Mol Genet Genomic Med. 8(2):e1083. https://doi.org/10.1002/mgg3.1083
Acknowledgements
The authors gratefully acknowledge the following organization for their help in obtaining animal samples: Guangzhou Zoo & Guangzhou Wildlife Research Center, Guangzhou, Guangdong, China. The authors would also like to thank the reviewers for their helpful contributions toward this paper.
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This work was supported by the Science and Technology Program of Guangzhou, China (No. 2019030016) and Open Project of Key Laboratory of Forensic Genetics, Ministry of Public Security (2020FGKFKT05).
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All the authors contributed to the study conception and design. Material preparation, data collection and analysis were performed by Xueyuan Liu, Chengliang Yang and Xiaohui Chen. The first draft of the manuscript was written by Xueyuan Liu and Chengliang Yang. All the authors commented on previous versions of the manuscript. All the authors read and approved the final manuscript.
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Liu, X., Yang, C., Chen, X. et al. A novel 193-plex MPS panel integrating STRs and SNPs highlights the application value of forensic genetics in individual identification and paternity testing. Hum. Genet. 143, 371–383 (2024). https://doi.org/10.1007/s00439-024-02658-1
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DOI: https://doi.org/10.1007/s00439-024-02658-1