Snedeker Jennifer L, Peck Michelle A, Russell David A, Holmes Amy S, Neal Christina M, Reedy Carmen R, Hughes Sheree R, Houston Rachel M
Department of Forensic Science, College of Criminal Justice, Sam Houston State University, Huntsville, TX, USA.
Signature Science, LLC, Charlottesville, VA, USA.
Forensic Sci Int Genet. 2025 Mar;76:103209. doi: 10.1016/j.fsigen.2024.103209. Epub 2024 Dec 14.
While skeletal remains are known for their resilience and often serve as the final source of information for unidentified human remains (UHRs), the traditional downstream processing of these samples is challenging due to their low template nature, DNA degradation, and the presence of PCR inhibitors, typically resulting in limited probative information. To address this issue, advanced genotyping methods can be explored to retrieve additional genetic information from these challenging samples to maximize investigative leads. Therefore, this study investigated the effectiveness of three advanced genotyping methods and assessed their suitability with compromised skeletal samples: 1) targeted next generation sequencing (NGS) of both STRs and SNPs using the ForenSeq® DNA Signature Prep chemistry, 2) targeted NGS of SNPs using the ForenSeq® Kintelligence kit, and 3) SNP genotyping using a microarray via the Infinium Global Screening Array. The genotype recovery and added investigative leads were compared across all methods. All three approaches demonstrated success with the challenging skeletal samples used in this study. Specifically, the ForenSeq® DNA Signature Prep chemistry outperformed traditional STR typing by improving the recovery of CODIS core loci. Additionally, the ForenSeq® Kintelligence kit and Infinium Global Screening Array provided eligible results for forensic investigative genetic genealogy (FIGG) searching. Based on these successes, we have developed a proposed workflow for downstream processing of challenging skeletal samples. Following the guidelines of the US Department of Justice, the recovery of the CODIS core loci should be attempted through traditional CE-based methods or a NDIS-approved NGS chemistry, such as ForenSeq® DNA Signature Prep. Alternatively, a mitochondrial DNA profile may be uploaded to CODIS for comparisons in UHR cases. However, if no probative information is developed from the forensic profile uploaded to CODIS, then FIGG methods can be implemented using the Infinium Global Screening Array for high-quality skeletal samples (DNA concentrations ≥ 0.5 ng/µL) or the ForenSeq® Kintelligence chemistry for low-template skeletal remains (DNA concentration ≤ 0.5 ng/µL). These findings provide valuable insight into the suitability and efficacy of advanced genotyping methods, offering promising opportunities for enhancing the investigation of cases involving UHRs.
虽然骨骼遗骸以其韧性著称，并且常常作为身份不明人类遗骸（UHRs）的最终信息来源，但由于这些样本的模板量低、DNA降解以及存在PCR抑制剂，其传统的下游处理具有挑战性，通常导致可用于调查的信息有限。为了解决这个问题，可以探索先进的基因分型方法，从这些具有挑战性的样本中获取更多遗传信息，以最大限度地增加调查线索。因此，本研究调查了三种先进基因分型方法的有效性，并评估了它们对受损骨骼样本的适用性：1）使用ForenSeq® DNA Signature Prep化学方法对STR和SNP进行靶向新一代测序（NGS），2）使用ForenSeq® Kintelligence试剂盒对SNP进行靶向NGS，3）通过Infinium Global Screening Array微阵列进行SNP基因分型。对所有方法的基因型回收率和增加的调查线索进行了比较。所有这三种方法在本研究中使用的具有挑战性的骨骼样本上均取得了成功。具体而言，ForenSeq® DNA Signature Prep化学方法通过提高CODIS核心位点的回收率，优于传统的STR分型。此外，ForenSeq® Kintelligence试剂盒和Infinium Global Screening Array为法医调查遗传谱系（FIGG）搜索提供了合格的结果。基于这些成功经验，我们制定了一个针对具有挑战性的骨骼样本的下游处理建议工作流程。遵循美国司法部的指导方针，应尝试通过基于传统毛细管电泳（CE）的方法或经国家DNA索引系统（NDIS）批准的NGS化学方法（如ForenSeq® DNA Signature Prep）来恢复CODIS核心位点。或者，可以将线粒体DNA图谱上传到CODIS，用于UHR案件的比对。然而，如果上传到CODIS的法医图谱没有产生可用于调查的信息，那么对于高质量骨骼样本（DNA浓度≥0.5 ng/µL），可以使用Infinium Global Screening Array实施FIGG方法；对于低模板量的骨骼遗骸（DNA浓度≤0.5 ng/µL），则可以使用ForenSeq® Kintelligence化学方法。这些发现为先进基因分型方法的适用性和有效性提供了有价值的见解，为加强涉及UHRs案件的调查提供了有前景的机会。
