@article{scheible_stinson_breen_callahan_thomas_meiklejohn_2024, title={The development of non-destructive sampling methods of parchment skins for genetic species identification}, volume={19}, ISSN={["1932-6203"]}, url={https://doi.org/10.1371/journal.pone.0299524}, DOI={10.1371/journal.pone.0299524}, abstractNote={Parchment, the skins of animals prepared for use as writing surfaces, offers a valuable source of genetic information. Many have clearly defined provenance, allowing for the genetic findings to be evaluated in temporal and spatial context. While these documents can yield evidence of the animal sources, the DNA contained within these aged skins is often damaged and fragmented. Previously, genetic studies targeting parchment have used destructive sampling techniques and so the development and validation of non-destructive sampling methods would expand opportunities and facilitate testing of more precious documents, especially those with historical significance. Here we present genetic data obtained by non-destructive sampling of eight parchments spanning the 15th century to the modern day. We define a workflow for enriching the mitochondrial genome (mtGenome), generating next-generation sequencing reads to permit species identification, and providing interpretation guidance. Using sample replication, comparisons to destructively sampled controls, and by establishing authentication criteria, we were able to confidently assign full/near full mtGenome sequences to 56.3% of non-destructively sampled parchments, each with greater than 90% of the mtGenome reference covered. Six of eight parchments passed all four established thresholds with at least one non-destructive sample, highlighting promise for future studies.}, number={3}, journal={PLOS ONE}, author={Scheible, Melissa and Stinson, Timothy L. and Breen, Matthew and Callahan, Benjamin J. and Thomas, Rachael and Meiklejohn, Kelly A.}, editor={Shakoori, Abdul RaufEditor}, year={2024}, month={Mar} } @article{meiklejohn_scheible_boggs_dunn_ricke_2023, title={Using FastID to analyze complex SNP mixtures from indoor dust}, volume={4}, ISSN={["1556-4029"]}, url={https://doi.org/10.1111/1556-4029.15246}, DOI={10.1111/1556-4029.15246}, abstractNote={AbstractForensically relevant single nucleotide polymorphisms (SNPs) can provide valuable supplemental information to short tandem repeats (STRs) for investigative leads, and genotyping can now be streamlined using massively parallel sequencing (MPS). Dust is an attractive evidence source, as it accumulates on undisturbed surfaces, often is overlooked by perpetrators, and contains sufficient human DNA for analysis. To assess whether SNPs genotyped from indoor dust using MPS could be used to detect known household occupants, 13 households were recruited and provided buccal samples from each occupant and dust from five predefined indoor locations. Thermo Fisher Scientific Precision ID Identity and Ancestry Panels were utilized for SNP genotyping, and sequencing was completed using Illumina® chemistry. FastID, a software developed to permit mixture analysis and identity searching, was used to assess whether known occupants could be detected from associated household dust samples. A modified “subtraction” method was also used in FastID to estimate the percentage of alleles in each dust sample contributed by known and unknown occupants. On average, 72% of autosomal SNPs were recovered from dust samples. When using FastID, (a) 93% of known occupants were detected in at least one indoor dust sample and could not be excluded as contributors to the mixture, and (b) non‐contributor alleles were detected in 54% of dust samples (29 ± 11 alleles per dust sample). Overall, this study highlights the potential of analyzing human DNA present in indoor dust to detect known household occupants, which could be valuable for investigative leads.}, journal={JOURNAL OF FORENSIC SCIENCES}, author={Meiklejohn, Kelly A. and Scheible, Melissa K. R. and Boggs, Laura M. and Dunn, Robert R. and Ricke, Darrell O.}, year={2023}, month={Apr} } @article{scheible_timpano_boggs_meiklejohn_2021, title={An alternate workflow for preparing Precision ID Ancestry and Identity Panel libraries for Illumina sequencing}, volume={135}, ISSN={["1437-1596"]}, url={https://doi.org/10.1007/s00414-021-02549-4}, DOI={10.1007/s00414-021-02549-4}, abstractNote={Single nucleotide polymorphisms (SNPs) are well-established for forensic applications. Although they are not compatible with existing criminal databases, they offer some advantages over short tandem repeat (STR) markers including smaller amplicons, no stutter artifacts, and biogeographic ancestry and phenotype predictions. The Precision ID NGS System, a commercial workflow by Thermo Fisher Scientific, offers a streamlined solution for genotyping forensically relevant SNPs using next-generation sequencing. The Precision ID Ancestry and Identity Panels combined target 289 SNPs, and their sensitivity, reproducibility, and accuracy have been evaluated by the forensic community. The aim of this study was to develop an alternative workflow to genotype these SNP panels using Illumina chemistry. Commercial genomic DNAs (gDNAs) (n, 3) were amplified using three uracil-tolerant polymerase master mixes. Resulting amplicons were prepared into libraries using the KAPA Hyper Prep Kit (KAPA Biosystems) and sequenced via Illumina's MiniSeq. Reads were analyzed using a published analysis pipeline to compile final genotypes with read depth information. Phusion U Multiplex PCR Master Mix (Thermo Fisher Scientific) statistically outperformed the other master mixes tested (P <0.0001), with respect to the number of SNPs genotyped. To ensure a workflow using Phusion U would be compatible across diverse samples, we optimized PCR cycle number using the same commercial gDNAs (n, 3), reference buccal swabs (n, 3), and environmental (household dust) samples (n, 6). Using the developed workflow, 93.9% of all SNPs were successfully genotyped across sample types. Implementation of the developed workflow should be straightforward for forensic laboratories and suitable for processing reference and casework samples.}, number={5}, journal={INTERNATIONAL JOURNAL OF LEGAL MEDICINE}, publisher={Springer Science and Business Media LLC}, author={Scheible, Melissa K. R. and Timpano, Emma K. and Boggs, Laura M. and Meiklejohn, Kelly A.}, year={2021}, month={Sep}, pages={1717–1726} } @article{timpano_scheible_meiklejohn_2020, title={Optimization of the second internal transcribed spacer (ITS2) for characterizing land plants from soil}, volume={15}, ISSN={["1932-6203"]}, url={https://doi.org/10.1371/journal.pone.0231436}, DOI={10.1371/journal.pone.0231436}, abstractNote={Molecular-based taxonomy, specifically DNA barcoding, has streamlined organism identification. For land plants, the recommended 2-locus barcode of rbcL and matK is not suitable for all groups, thus the second subunit of the nuclear internal transcribed spacer (ITS2) has received attention as a possible alternative. To date, evaluations of ITS2 have mostly been limited in scope to specific plant orders/families and single source material. Prior to using ITS2 to routinely characterize land plants present in environmental samples (i.e., DNA metabarcoding), a wet lab protocol optimized for bulk sample types is needed. To address this gap, in this study we determined the broad recoverability across land plants when using published ITS2 primer pairs, and subsequently optimized the PCR reaction constituents and cycling conditions for the best two performing primer pairs (ITS2F/ITSp4 and ITSp3/ITSu4). Using these conditions, both primer pairs were used to characterize land plants present in 17 diverse soils collected from across the US. The resulting PCR amplicons were prepared into libraries and pooled for sequencing on an Illumina® MiniSeq. Our existing bioinformatics workflow was used to process raw sequencing data and taxonomically assign unique ITS2 plant sequences by comparison to GenBank. Given strict quality criteria were imposed on sequences for inclusion in data analysis, only 43.6% and 7.5% of sequences from ITS2F/ITSp4 and ITSp3/ITSu4 respectively remained for taxonomic comparisons; ~7–11% of sequences originated from fungal co-amplification. The number of orders and families recovered did differ between primer pairs, with ITS2F/ITSp4 consistently outperforming ITSp3/ITSu4 by >15%. Primer pair bias was observed in the recovery of certain taxonomic groups; ITS2F/ITSp4 preferentially recovered flowering plants and grasses, whereas ITSp3/ITSu4 recovered more moss taxa. To maximize data recovery and reduce potential bias, we advocate that studies using ITS2 to characterize land plants from environmental samples such as soil use a multiple primer pair approach.}, number={4}, journal={PLOS ONE}, publisher={Public Library of Science (PLoS)}, author={Timpano, Emma K. and Scheible, Melissa K. R. and Meiklejohn, Kelly A.}, editor={Kalendar, RuslanEditor}, year={2020}, month={Apr} } @article{silva_sawitzki_scheible_bailey_alho_faith_2018, title={Paternity testing using massively parallel sequencing and the PowerSeq (TM) AUTO/Y system for short tandem repeat sequencing}, volume={39}, ISSN={["1522-2683"]}, DOI={10.1002/elps.201800072}, abstractNote={AbstractMassively parallel sequencing (MPS) is gaining attention as a new technology for routine forensic casework, including paternity testing. Recently released MPS multiplex panels provide many more loci compared to CE methods, plus provide sequence‐based alleles that together improve the statistical power of the genetic testing. Here, an MPS system (PowerSeq™ AUTO/Y) was applied for STR sequencing in the study of first‐degree STR sequence allele inheritance from families in Southern Brazil. In 29 trios (mother‐child‐father) analyzed, the paternity index values generally increased when data from sequence‐based analysis were used in comparison to length‐based data. Further, allele inconsistencies (e.g., single repeat mutation events) between child and parents could be resolved with MPS by assessing the core repeat and flanking region sequences. Lastly, the sequence information allowed for identification of isoalleles (alleles of the same size, but different sequence) to determine specific paternal and maternal inheritances. The results from this study showed advantages of implementing sequence‐based analysis, MPS, in paternity testing with improved statistical calculations and a greater resolution for the trios/families tested.}, number={21}, journal={ELECTROPHORESIS}, author={Silva, Deborah S. B. S. and Sawitzki, Fernanda R. and Scheible, Melissa K. R. and Bailey, Sarah F. and Alho, Clarice S. and Faith, Seth A.}, year={2018}, month={Nov}, pages={2669–2673} } @article{allwood_scheible_faith_2018, title={Demonstration of a basic mitochondrial enrichment method to produce the complete mitochondrial genome sequence of the endangered North Atlantic right whale (Eubalaena glacialis)}, volume={10}, ISSN={["1877-7260"]}, DOI={10.1007/s12686-017-0854-6}, number={3}, journal={CONSERVATION GENETICS RESOURCES}, author={Allwood, J. S. and Scheible, M. K. and Faith, S. A.}, year={2018}, month={Sep}, pages={483–486} } @article{silva_sawitzki_scheible_bailey_alho_faith_2018, title={Genetic analysis of Southern Brazil subjects using the PowerSeq (TM) AUTO/Y system for short tandem repeat sequencing}, volume={33}, ISSN={["1878-0326"]}, DOI={10.1016/j.fsigen.2017.12.008}, abstractNote={With the advent of Next-Generation Sequencing technology, sequencing of short tandem repeats (STRs) allows for a more detailed analysis when compared to size-based fragment methods (capillary electrophoresis-CE). The implementation of high-throughput sequencing can help uncover deeper genetic diversities of different populations. Subjects from the South region of Brazil present a particular and more homogeneous ancestry background when compared to other regions of the country. Both autosomal and Y- STRs have been analyzed in these individuals; however, all analyses published to date encompass data from CE-based fragment analysis. In this study, a genetic analysis of 59 individuals from Southern Brazil was performed on STR sequences. Forensically relevant STRs were PCR-enriched using a prototype of the PowerSeq™ AUTO/Y system (Promega Corp.). Next-generation sequencing was performed on an Illumina MiSeq instrument. The raw data (FASTQ files) were processed using a custom designed sequence processing tool, Altius. Isoalleles, which are sequence-based allelic variants that do not differ in length, were observed in nine autosomal and in six Y- STRs from the core global forensic marker set. The number of distinctive alleles based on sequence was higher when compared to those based on length, 37.3% higher in autosomal STRs and 13.8% higher in Y-STRs. The most polymorphic autosomal locus was D12S391, which presented 38 different sequence-based alleles. Among the loci in the Y chromosome, DYS389II presented the highest number of isoalleles. In comparison to CE analysis, Observed and Expected Heterozygosity, Polymorphic Information Content (PIC) and Genetic Diversity also presented higher values when the alleles were analyzed based on their sequence. For autosomal loci, Polymorphic Information Content (PIC) was 2.6% higher for sequence-based data. Diversity was 9.3% and 6.5% higher for autosomal and Y markers, respectively. In the analysis of the repeat structures for the STR loci, a new allele variant was found for allele 18 in the vWA locus. The STR flanking regions were also further investigated and sixteen variations were observed at nine autosomal STR loci and one Y-STR locus. The results obtained in this study demonstrate the importance of genetic analysis based on sequencing and highlight the diversity of the South Brazilian population when characterized by STR sequencing.}, journal={FORENSIC SCIENCE INTERNATIONAL-GENETICS}, author={Silva, Deborah S. B. S. and Sawitzki, Fernanda R. and Scheible, Melissa K. R. and Bailey, Sarah F. and Alho, Clarice S. and Faith, Seth A.}, year={2018}, month={Mar}, pages={129–135} } @article{scheible_just_sturk-andreaggi_saunier_parson_parsons_coble_irwin_2016, title={The mitochondrial landscape of African Americans: An examination of more than 2500 control region haplotypes from 22 US locations}, volume={22}, ISSN={["1878-0326"]}, DOI={10.1016/j.fsigen.2016.01.002}, abstractNote={The mitochondrial DNA (mtDNA) control region (16024-576) was Sanger-sequenced for a total of 2563 self-identified African Americans, using automated processing techniques and data review standards exceeding guidelines for forensic applications. Genetic diversity ranged from 0.9952 to 0.9998 in 22 population samples from 20 different states. Haplogroups of African ancestry, found in 82.48% of individuals overall, were most concentrated in the Southeast U.S. and decreased to the north and west. West African and West Central African haplotypes were well-represented in the population samples, especially in the southern U.S. states, while East African haplogroups were observed in low-frequency clusters in a handful of locations across the country. East Asian, Native American, and West Eurasian admixture was present in 3.16%, 2.93%, and 11.43% of samples, respectively. While some geographic substructure was detected across the population samples as clines in admixture frequencies, 20 of the 22 population samples were found to be statistically indistinguishable by pairwise comparisons and AMOVA calculations. Datasets from Hawaii and Idaho, however, were clear outliers. Overall, these more than 2500 control region sequences represent the most comprehensive regional sampling of African American mtDNA diversity to date, and are suitable for use in a forensic mtDNA database. The population data are made available via EMPOP (www.empop.org) and GenBank.}, journal={FORENSIC SCIENCE INTERNATIONAL-GENETICS}, author={Scheible, M. and Just, R. and Sturk-Andreaggi, K. and Saunier, J. and Parson, W. and Parsons, T. and Coble, M. and Irwin, J.}, year={2016}, month={May}, pages={139–148} }