Forensic and Investigative Genetics

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    The Validation of the RapidHIT ID System for Human Identification
    (2020) King, Jonathan; Budowle, Bruce; Cihlar, Jennifer; Campos, Gemma R.
    Forensic scientists have routinely employed short tandem repeat (STR) typing as a means to identify or exclude individuals that may have left biological evidence at crime scenes. Despite developments in automation of STR typing workflow, the work is time consuming and requires the expertise and resources of a dedicated forensic genetics laboratory. Rapid DNA technology offers a simple, 90-minute swab-to-profile approach to generate STR profiles, requiring minimal training and expertise to operate the system. Use of Rapid DNA technology in the field has been met with some success in identifying victims of natural disasters and determining familial relationships. As Rapid DNA continues to be developed and implemented, validation studies are necessary to ensure the quality of analyses performed by Rapid DNA instruments. A series of studies in accordance with SWGDAM guidelines were designed to test different performance measures of the RapidHIT ID System for Human Identification (Thermo Fisher Scientific). Blood and buccal samples at varying amounts of DNA were collected and subsequently analyzed using the RapidHIT instrument to identify optimal DNA input amounts. Concordance between RapidHIT and capillary electrophoresis-generated profiles served to demonstrate comparability to the current standard in STR typing. Sample retrieval and re-analysis through Rapid DNA technology or other typing methods were tested as this workflow can be useful in both a practical and laboratory setting. The results from this study provide opportunities for protocol and user guide revisions, software improvements, and highlighting parts of instrumentation that can be improved for future models.
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    Variation in Mitochondrial DNA Heteroplasmy from Blood, Buccal, and Hair Samples
    (2020) King, Jonathan; Budowle, Bruce; Churchill Cihlar, Jennifer; Colon, Natalie
    Forensic samples may experience suboptimal environmental conditions wherein nuclear DNA is susceptible to degradation. The relatively higher copy number per cell typical of mitochondrial DNA (mtDNA) makes it more persistent under these conditions and, thus, forensically relevant. Furthermore, mtDNA has been found to express point heteroplasmy (PHP), or single base substitutions, that impact lineage identification. The aim of the current research is to determine if variation in expression of PHP exists when comparing blood, buccal, and head hair samples from five subjects previously determined to have PHPs. Five hairs were collected from five discrete scalp locations (25 total) per subject. One scalp site from a single subject was used to further study variation down the length of the hair shaft. Extracted mtDNA was processed and sequenced using the Ion Chef and Ion S5 platform, followed by data analysis using the Torrent Suite and Converge Software. Results were evaluated using performance metrics such as concordance across sampling sites, strand balance, noise, and read depth. Current SWGDAM guidelines state a sequence comparison results in a judgement of exclusion if the samples differ at two or more nucleotide positions and is inconclusive if the samples differ at one nucleotide position. As it is possible for individuals to express two heteroplasmic sites or present as homoplasmic in one tissue type and heteroplasmic in another, the results of this study may suggest revision of sample collection methodology from known individuals when interpreting hair evidence.
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    Selecting informative skin microbiome single nucleotide polymorphisms (SNPs) for human identification
    (2020) Budowle, Bruce; Woerner, August E.; Sherier, Allison
    The human microbiome is considered the second human genome, contributing to a large portion of the genetic content of the human body. The microbiome of the skin adds additional genetic information that can be used to supplement traditional forensic DNA evidence derived from human sources. Based on past research, a massively parallel sequencing (MPS) panel called hidSkinPlex was designed as a targeted clade-specific multiplex which contains 282 bacterial and 4 phage markers. The hidSkinPlex was developed under the hypothesis that genes from stable, universal microbial species can differentiate skin microbiomes of individuals and be applied towards forensic human identification (HID) purposes. Further testing of hidSkinPlex has produced extensive sequence data from the microbiomes of 51 unrelated individuals from 3 forensically relevant body-sites in triplicate. The focus of this research study is to improve upon classification accuracies by using genetic distance to identify individuals instead of taxonomic differences like past research. Selecting SNPs that differentiate individuals based on their microbial populations will allow for a novel approach to HID using the skin microbiome, similar to past approaches for selecting SNPs that differentiate individuals based on their human bioancestry. Wright's fixation index (FST) was used as a novel genetic approach to select SNPs of interest in conjunction with supervised machine learning techniques. This work supports the hypothesis that the skin microbiome may be a viable source for HID and potentially improve analysis of samples currently yielding low level human DNA, such as touch samples