Short Tandem Repeat Data Research Articles

Genetic diversity and origin of Kazakh Tobet Dogs

The Kazakh Tobet is an indigenous Kazakh dog breed that has been used to guard livestock since ancient times. To understand the genetic structure and phylogenetic relationship of the Kazakh Tobet breed with other herding and livestock guarding dog breeds, we analysed short tandem repeat data of 107 Kazakh Tobet dogs from different regions of Kazakhstan and Mongolia, as well as whole genome sequencing data from two Kazakh Tobet dogs and 43 dogs from 24 working breeds. Our results indicate a high genetic diversity of the Kazakh Tobet, with the average number of alleles per locus ranging from 6.00 to 10.22 and observed heterozygosity ranging from 76 to 78%. The breed has a complex genetic structure characterised by seven different clusters. The neighbour-joining tree constructed based on 14,668,406 autosomal and the maximum likelihood tree based on mitochondrial D-loop sequences indicate a common genetic heritage between the Kazakh Tobet, the Central Asian Shepherd Dog and the Turkish Akbash. The presence of haplotype A18 in the Kazakh Tobets supports the hypothesis of the ancient origin of the breed, which was previously suggested by archaeological finds and written sources. These results provide an important genetic basis for the ongoing efforts to improve the Kazakh Tobet breed, to ensure its preservation as an independent genetic lineage and to recognise a breed on an international level.

Short Tandem Repeats in the era of next-generation sequencing: from historical loci to population databases.

In this study, we explore the landscape of short tandem repeats (STRs) within the human genome through the lens of evolving technologies to detect genomic variations. STRs, which encompass approximately 3% of our genomic DNA, are crucial for understanding human genetic diversity, disease mechanisms, and evolutionary biology. The advent of high-throughput sequencing methods has revolutionized our ability to accurately map and analyze STRs, highlighting their significance in genetic disorders, forensic science, and population genetics. We review the current available methodologies for STR analysis, the challenges in interpreting STR variations across different populations, and the implications of STRs in medical genetics. Our findings underscore the urgent need for comprehensive STR databases that reflect the genetic diversity of global populations, facilitating the interpretation of STR data in clinical diagnostics, genetic research, and forensic applications. This work sets the stage for future studies aimed at harnessing STR variations to elucidate complex genetic traits and diseases, reinforcing the importance of integrating STRs into genetic research and clinical practice.

Sequencing-induced artefacts in NGS STR data

Significant progress has been made in recent years in the development of techniques for Next Generation Sequencing (NGS), or Massively Parallel Sequencing (MPS), of forensically relevant short tandem repeat (STR) loci. However, as these technologies are investigated and adopted by forensic laboratories, new challenges unfold that require further scrutiny. In the analysis of DNA profiles generated using the MiSeq FGx sequencing system, we have observed noise sequences with relatively high readcounts that are challenging to distinguish from genuine alleles. These high read count noise sequences appear as allele sequences with one or a few substituted bases compared to a known allele sequence within the profile.An examination of ForenSeq DNA Signature Prep Kit STR noise sequences revealed that the substituted base of a parent allele can align to the same position on the sequence across noise sequences. This suggests that these substitution events occur at specific positions within the amplicon, resulting in multiple noise reads with substitutions at the same position. Mapping of the noise events onto the original raw read positions revealed a high number of events, or “noise spikes”, occurring atspecific positions within a given sequencing run. These noise spikes affected reads across the entire run, agnostic of locus or sample, while the position, occurrence, and amplitude of the spikes differed across runs. The majority of noise sequences with high read counts in a DNA profile were generated from base changes at these spike positions, and could be classified as “noise spike artefacts”.In this paper we present evidence of the noise spike artefacts and their genesis during the sequencing process in the sequencing-by-synthesis (SBS) cycles, as well as the methods developed to detect them. The information and methods will assist laboratories with detecting noise spikes in MiSeq FGx sequencing runs, differentiating authentic allele sequences from noise spike artefacts, and developing protocols for analyst review and handling of MiSeq FGx data.

Fighting passport fraudulence using DNA analyses

ABSTRACT Passports are the number one form of individual identification when crossing borders resulting in counterfeit or modified passports being used to allow illegal travel. Here we report on the use of Diamond Dye to detect cells deposited by touch onto parts of the photograph page of a passport. The passport page was divided into a grid of 11 rectangles of approximately equal dimensions. Thumbprints were deposited at 10 of the 11 squares, with the 11th left blank as a negative control. The location and number of deposited cells were determined and recorded. Cellular material was removed from each square separately, using a moistened swab or a tapelift. Tapelifting was better suited for the substrate resulting in an increase in DNA recovered. The highest total quantity of DNA isolated from the 10 samples post extraction was 0.162 ng. Short tandem repeat data were obtained from all 10 samples using the GlobalFilerTM PCR Amplification Kit. The DNA profiles were single source or two-person mixtures and the donor was always the major contributor. These data show the potential to visualize cellular material deposited by touch and then generate STR data from an item such as a passport after only brief physical contact.

Population genetic analyses of Eastern Chinese Han nationality using ForenSeq™ DNA Signature Prep Kit.

Currently, the most commonly used method for human identification and kinship analysis in forensic genetics is the detection of length polymorphism in short tandem repeats (STRs) using polymerase chain reaction (PCR) and capillary electrophoresis (CE). However, numerous studies have shown that considerable sequence variations exist in the repeat and flanking regions of the STR loci, which cannot be identified by CE detection. Comparatively, massively parallel sequencing (MPS) technology can capture these sequence differences, thereby enhancing the identification capability of certain STRs. In this study, we used the ForenSeq™ DNA Signature Prep Kit to sequence 58 STRs and 94 individual identification SNPs (iiSNPs) in a sample of 220 unrelated individuals from the Eastern Chinese Han population. Our aim is to obtain MPS-based STR and SNP data, providing further evidence for the study of population genetics and forensic applications. The results showed that the MPS method, utilizing sequence information, identified a total of 486 alleles on autosomal STRs (A-STRs), 97 alleles on X-chromosome STRs (X-STRs), and 218 alleles on Y-chromosome STRs (Y-STRs). Compared with length polymorphism, we observed an increase of 260 alleles (157, 31, and 72 alleles on A-STRs, X-STRs, and Y-STRs, respectively) across 36 STRs. The most substantial increments were observed in DYF387S1 and DYS389II, with increases of 287.5% and 250%, respectively. The most increment in the number of alleles was found at DYF387S1 and DYS389II (287.5% and 250%, respectively). The length-based (LB) and sequence-based (SB) combined random match probability (RMP) of 27 A-STRs were 6.05E-31 and 1.53E-34, respectively. Furthermore, other forensic parameters such as total discrimination power (TDP), cumulative probability of exclusion of trios (CPEtrio), and duos (CPEduo) were significantly improved when using the SB data, and informative data were obtained for the 94 iiSNPs. Collectively, these findings highlight the advantages of MPS technology in forensic genetics, and the Eastern Chinese Han genetic data generated in this study could be used as a valuable reference for future research in this field.

Modification of Huntington's disease by short tandem repeats.

Expansions of glutamine-coding CAG trinucleotide repeats cause a number of neurodegenerative diseases, including Huntington's disease and several of spinocerebellar ataxias. In general, age-at-onset of the polyglutamine diseases is inversely correlated with the size of the respective inherited expanded CAG repeat. Expanded CAG repeats are also somatically unstable in certain tissues, and age-at-onset of Huntington's disease corrected for individual HTT CAG repeat length (i.e. residual age-at-onset), is modified by repeat instability-related DNA maintenance/repair genes as demonstrated by recent genome-wide association studies. Modification of one polyglutamine disease (e.g. Huntington's disease) by the repeat length of another (e.g. ATXN3, CAG expansions in which cause spinocerebellar ataxia 3) has also been hypothesized. Consequently, we determined whether age-at-onset in Huntington's disease is modified by the CAG repeats of other polyglutamine disease genes. We found that the CAG measured repeat sizes of other polyglutamine disease genes that were polymorphic in Huntington's disease participants but did not influence Huntington's disease age-at-onset. Additional analysis focusing specifically on ATXN3 in a larger sample set (n = 1388) confirmed the lack of association between Huntington's disease residual age-at-onset and ATXN3 CAG repeat length. Additionally, neither our Huntington's disease onset modifier genome-wide association studies single nucleotide polymorphism data nor imputed short tandem repeat data supported the involvement of other polyglutamine disease genes in modifying Huntington's disease. By contrast, our genome-wide association studies based on imputed short tandem repeats revealed significant modification signals for other genomic regions. Together, our short tandem repeat genome-wide association studies show that modification of Huntington's disease is associated with short tandem repeats that do not involve other polyglutamine disease-causing genes, refining the landscape of Huntington's disease modification and highlighting the importance of rigorous data analysis, especially in genetic studies testing candidate modifiers.

STRategy: A support system for collecting and analyzing next-generation sequencing data of short tandem repeats for forensic science.

Short tandem repeats (STRs) are short repeated sequences commonly found in the human genome and valuable in forensic science, used for human identity and relatedness markers. Next-generation sequencing (NGS) technologies, e.g., ForenSeq Signature Prep, can sequence STRs, inferring length-based alleles and single nucleotide polymorphisms (SNPs) and providing valuable insights into population and sub-population structures. Despite the potential benefits of NGS for STRs, no open-source software platform integrates the collection, management, and analysis of STR data from NGS into one place. Users must use multiple programs to process their STR data and then collect the results into a separate database or a file system folder. Moreover, analyzing repeat structures (STR repeat motifs) may require learning multiple software tools, making the process inefficient and cumbersome. To address this gap, we introduce the STRategy, a standalone web-based application supporting essential STR data management and analysis capabilities. The STRategy allows users to collect their data into its database, automatically calculates forensic parameters, and visualizes the analyzed data in various forms. Users can search the database using different options, such as by profile, loci, and genotypes, with and without a specific test kit. Moreover, users can also find the nucleotide variants of a locus among the samples. We designed the STRategy for internal use in a laboratory or an organization. Hence, our system includes role-based access control that allows users to search for or access specific data based on their responsibilities. The administrator role can customize the system, for example, configure maps according to the samples' geographic data, and manage reference STR repeat motifs. A laboratory or an organization can download and install a copy of STRategy on their local system using Docker, as described in https://github.com/cucpbioinfo/STRategy. In summary, the STRategy is an end-to-end system that provides users with a database to collect the analyzed STR data from NGS, the dynamic analyses of forensic parameters, and the variants of STR patterns according to the newly added samples, which are then explorable via various search options and visualizations. The system is helpful for both forensic investigations and forensic genetics.

A genetic Study of the Ghanaian Population Using 15 Autosomal STR Loci.

Autosomal short tandem repeat (STR) population data collected from a well characterized population are needed to correctly assigning the weight of DNA profiles in the courtroom and widely used for ancestral analyses. In this study, allele frequencies for the 15 autosomal short tandem repeat (STR) loci included in the AmpFlSTR® Identifiler® plus kit (D8S1179, D21S11, D7S820, CSF1PO, D3S1358, TH01, D13S317, D16S539, D2S1338, D19S433, VWA, TPOX, D18S51, D5S818, FGA) were obtained by genotyping 332 unrelated individuals of Ghanaian origin. Statistical tests on STR genotype data showed no significant departure from Hardy-Weinberg equilibrium (HWE). The overall match probability, combined power of exclusion and combined power of discrimination for these loci were 1 in 3.85 × 1017, 0.99999893 and 0.99999998, respectively. Polymorphic information content (PIC) greater than 0.70 was observed for all loci except TH01 and D13S317. These statistical parameters confirm that this combination of loci is valuable for forensic identification and parentage analysis. Our results were also compared with those for 20 other human populations analyzed for the same set of markers. We observed that the Ghanaian population grouped with other African populations in two-dimensional principal coordinate (PCO) and a neighbor-joining (N-J) data mapping and placed closest to Nigerians. This observation reflects cultural similarities and geographical factors, coupled with the long history of migration and trading activities between Ghana and Nigeria. Our report provides what we believe to be the first published autosomal STR data for the general Ghanaian population using 15 loci genotyped using the AmpFlSTR® Identifiler® plus kit methodology. Our data show that the loci tested have sufficient power to be used reliably for DNA profiling in forensic casework and help to elucidate the genetic history of people living in the country.

Optimization and Validation of Candida auris Short Tandem Repeat Analysis.

ABSTRACTCandida auris is an easily transmissible yeast with resistance to different antifungal compounds. Outbreaks of C. auris are mostly observed in intensive care units. To take adequate measures during an outbreak, it is essential to understand the transmission route, which requires isolate genotyping. In 2019, a short tandem repeat (STR) genotyping analysis was developed for C. auris. To determine the discriminatory power of this method, we performed STR analysis of 171 isolates with known whole-genome sequencing (WGS) data using Illumina reads, and we compared their resolutions. We found that STR analysis separated the 171 isolates into four clades (clades I to IV), as was also seen with WGS analysis. Then, to improve the separation of isolates in clade IV, the STR assay was optimized by the addition of 2 STR markers. With this improved STR assay, a total of 32 different genotypes were identified, while all isolates with differences of >50 single-nucleotide polymorphisms (SNPs) were separated by at least 1 STR marker. Altogether, we optimized and validated the C. auris STR panel for clades I to IV and established its discriminatory power, compared to WGS SNP analysis using Illumina reads.IMPORTANCE The emerging fungal pathogen Candida auris poses a threat to public health, mainly causing outbreaks in intensive care units. Genotyping is essential for investigating potential outbreaks and preventing further spread. Previously, we developed a STR genotyping scheme for rapid and high-resolution genotyping, and WGS SNP outcomes for some isolates were compared to STR data. Here, we compared WGS SNP and STR outcomes for a larger sample cohort. Also, we optimized the resolution of this typing scheme with the addition of 2 STR markers. Altogether, we validated and optimized this rapid, reliable, and high-resolution typing scheme for C. auris.

ABCL-371 STR Loci Allele Imbalance at 9p24.1 in Primary Mediastinal B-Cell Lymphoma

Molecular factors that determine the prognosis of immunotherapy response in primary mediastinal B-cell lymphoma (PMBCL) are not yet described. PD-L1/L2 expression levels in tumor samples have demonstrated clinical utility for many solid tumors. PD-L1/L2 expression levels could be affected by chromosomal lesions on 9p24.1 (losses or gains). Short tandem repeat (STR) analysis of appropriate markers is the simplest way to detect these lesions. To analyze possible associations between STR loci allele imbalance (AI) on 9p24.1 and PD-L1 expression levels in tumor cells from PMBCL patients. Two STR Loci (gt) n and (ttat)m on 9p24.1 in tumor cells were analyzed for a cohort of 36 PMBCL patients admitted to the National Medical Research Center for Hematology (Moscow, Russia) from 2013 to 2022. For six patients, STR data were compared with PD-L1 expression assessed by immunohistochemistry. DNA was isolated from tumor biopsy samples taken at diagnosis. Control DNA samples were taken from the blood of patients in complete remission or from the buccal epithelium. STR analysis for AI was performed by PCR with original primers. The fragment analysis was performed on an ABI3130 Genetic Analyzer. Inclusion criteria: de novo diagnosed PMBCL patients. pretreatment. The heterozygosity level for the (gt)n locus was 31 of 36 patients (86%), and AI was found in 16 of 31 (51.6%). The heterozygosity level for the (ttat)m locus was 20 of 36 patients (56%), and AI was found in 9 of 20 (45%). One patient was homozygous for both loci. No patients were found with AI at one locus and a normal heterozygous variant at the second locus. PD-L1 expression in tumor cells was evaluated for six patients. In two patients with aberrant STR loci, PD-L1 expression was found in 75%-100% of tumor cells. None of the four patients with normal 9p24.1 STR markers had PD-L1-positive cells. The high incidence of AI in the 9p24.1 locus indicates its possible involvement in the pathogenesis of PMBCL. The association between AI and PD-L1 expression in tumor cells requires confirmation in an extended cohort of patients. This study was supported by Rakfond grant 2/2020.

Analysis of Genetic Diversity in the American Standardbred Horse Utilizing Short Tandem Repeats and Single Nucleotide Polymorphisms.

American Standardbreds were developed as a harness racing horse breed. The United States Trotting Association closed the studbook in 1973 and implemented a book size cap in 2009. This study aimed to investigate genetic diversity in the American Standardbred after the studbook cap was introduced using short tandem repeats (STRs) and single-nucleotide polymorphisms (SNPs). Sixteen STRs from horses foaled from 2010 to 2015 and their sires and dams (n = 50 621) were utilized to examine allelic richness (Ar), expected heterozygosity (HE), observed heterozygosity (HO), unbiased heterozygosity (HU), inbreeding coefficient (FIS), and fixation index (FST). These analyses found that trotting and pacing sires were less genetically diverse than dams (HEPBonferroni = 0.029 and 6.3 × 10−5, respectively) and their offspring (ArPBonferroni = 0.034 and 6.9 × 10-6, respectively), and pacing offspring were significantly less diverse than their dams (HEPBonferroni = 2 × 10-3). Inbreeding coefficients for trotters (FIS = −0.014) and pacers (FIS = −0.012) suggest that breeding practices have maintained diversity. Moderate levels of genetic differentiation (0.066 < FST < 0.11) were found between pacing and trotting groups. Additionally, 10 of the most prolific trotting sires and their male offspring (n = 84) were genotyped on the 670K Axiom Equine HD Array. HO values higher than HE (P < 0.001), low inbreeding coefficients (mean F = −0.064), and mean FROH = 21% indicate relatively high levels of diversity in this cohort, further supporting the STR data. However, in contrast, HO values were higher for trotting sires (0.41) than their offspring (0.36). This observation warrants further monitoring of diversity over time. These data provide an updated foundation of diversity indices for further, long-term analysis in the breed.

Explainable artificial intelligence in forensics: Realistic explanations for number of contributor predictions of DNA profiles

Machine learning obtains good accuracy in determining the number of contributors (NOC) in short tandem repeat (STR) mixture DNA profiles. However, the models used so far are not understandable to users as they only output a prediction without any reasoning for that conclusion. Therefore, we leverage techniques from the field of explainable artificial intelligence (XAI) to help users understand why specific predictions are made. Where previous attempts at explainability for NOC estimation have relied upon using simpler, more understandable models that achieve lower accuracy, we use techniques that can be applied to any machine learning model. Our explanations incorporate SHAP values and counterfactual examples for each prediction into a single visualization. Existing methods for generating counterfactuals focus on uncorrelated features. This makes them inappropriate for the highly correlated features derived from STR data for NOC estimation, as these techniques simulate combinations of features that could not have resulted from an STR profile. For this reason, we have constructed a new counterfactual method, Realistic Counterfactuals (ReCo), which generates realistic counterfactual explanations for correlated data. We show that ReCo outperforms state-of-the-art methods on traditional metrics, as well as on a novel realism score. A user evaluation of the visualization shows positive opinions of end-users, which is ultimately the most appropriate metric in assessing explanations for real-world settings.

Population data for 15 autosomal STR loci in Orang Asli subgroups of Peninsular Malaysia.

Short tandem repeats (STRs) data for the Orang Asli population in Peninsular Malaysia is still scanty, especially for specific Orang Asli subgroups. The Orang Asli is believed as the earliest population arrived in Peninsular Malaysia about 50,000years ago and currently makes up only 0.6% of the total population of Malaysia. This study reports the allele frequencies and several forensic statistical parameters for 15 autosomal STR loci for six Orang Asli subgroups. A total of 164 Orang Asli individuals representing the Semai, Che Wong, Orang Kanaq, Lanoh, Bateq, and Kensui subgroups were recruited for this study. This STR data will enrich the existing Malaysian autosomal STR database and will be useful for kinship testing and forensic applications.

The impact of motherless paternity testing in a South African population.

Paternity investigations play an important role in determining biological relatedness, and in South Africa, the outcome of these investigations impacts medical, judicial and home affairs decisions. Short Tandem Repeat (STR) analysis is utilised to perform paternity and kinship analysis, due to the polymorphic nature of STR loci. The cost associated with paternity testing is high, and there is a demand for motherless testing. This study aims to determine what the impact of motherless testing would have been by evaluating 6182 paternity trio cases. The AmpFLSTR™ Identifiler™ PCR Amplification kit was used to profile each of the trio cases. A scenario was created where the mother was eliminated from the test results to determine if the paternity outcome would change. Putative fathers were excluded in 27% of all cases, and in 2.5% of those cases, putative fathers would have been falsely included, had the mother not been tested. These false inclusions are attributed to coincidental STR loci that are shared between the mother and the putative father. The addition of loci to the STR profiling kit may resolve the issue; however, comparable STR data with more loci will have to be evaluated to ensure it overcomes the issue of coincidentally shared loci between unrelated individuals. We would recommend that within our setup and within similar setups, the mother always be included for testing, except in extreme scenarios such as death. False inclusion of putative fathers could have serious legal implications for testing laboratories.

Application of a probabilistic genotyping software to MPS mixture STR data is supported by similar trends in LRs compared with CE data

The interpretation of short tandem repeat (STR) profiles can be challenging when, for example, alleles are masked due to allele sharing among contributors and/or when they are subject to drop-out, for instance from sample degradation. Mixture interpretation can be improved by increasing the number of STRs and/or loci with a higher discriminatory power. Both capillary electrophoresis (CE, 6-dye) and massively parallel sequencing (MPS) provide a platform for analysing relatively large numbers of autosomal STRs. In addition, MPS enables distinguishing between sequence variants, resulting in enlarged discriminatory power. Also, MPS allows for small amplicon sizes for all loci as spacing is not an issue, which is beneficial with degraded DNA. Altogether, MPS has the potential to increase the weights of evidence for true contributors to (complex) DNA profiles. In this study, likelihood ratio (LR) calculations were performed using STR profiles obtained with two different MPS systems and analysed using different settings: 1) MPS PowerSeq™ Auto System profiles analysed using FDSTools equipped with optimized settings such as noise correction, 2) ForenSeq™ DNA Signature Prep Kit profiles analysed using the default settings in the Universal Analysis Software (UAS), and 3) ForenSeq™ DNA Signature Prep Kit profiles analysed using FDSTools empirically adapted to cope with one-directional reads and provisional, basic settings. The LR calculations used genotyping data for two- to four-person mixtures varying for mixture proportion, level of drop-out and allele sharing and were generated with the continuous model EuroForMix. The LR results for the over 2000 sets of propositions were affected by the variation for the number of markers and analysis settings used in the three approaches. Nevertheless, trends for true and non-contributors, effects of replicates, assigned number of contributors, and model validation results were comparable for the three MPS approaches and alike the trends known for CE data. Based on this analogy, we regard the probabilistic interpretation of MPS STR data fit for forensic DNA casework. In addition, guidelines were derived on when to apply LR calculations to MPS autosomal STR data and report the corresponding results.

Concordance and characterization of massively parallel sequencing at 58 STRs in a Tibetan population.

BackgroundMassively parallel sequencing (MPS) is a promising supplementary method for forensic casework in short tandem repeats (STRs) genotyping, owing to several advantageous features in comparison to traditional capillary electrophoresis (CE). However, the application of MPS in casework requires accessible datasets from the worldwide population to enrich the allele frequencies of sequence‐based STR genotypes.MethodsIn this study, we report the characterization of sequence‐based allele frequencies of 58 STRs from a Tibetan population comprising 120 unrelated individuals using the ForenSeq™ DNA Signature Prep Kit. A concordance study evaluating MPS and CE allele data was performed to ensure that MPS is compatible with current CE‐based forensic databases. The diversity of observed alleles, allele frequencies, and forensic parameters per locus by length (LB), sequence without flanking region (RSB), and sequence with flanking region (FSB) were analyzed and compared.ResultsThe concordance study demonstrated a concordance rate exceeding 99%. The combined random match probability (RMP) for the 26 A‐STRs was 2.04 × 10–29, 1.93 × 10–31, and 9.56 × 10–33 for LB, RSB, and FSB, respectively. Similar trends were observed in other forensic parameters resulting from the increase in the number of unique alleles available. A total of 111 and 113 unique haplotypes in the Y‐STR loci were observed when using length‐based and sequence‐based alleles, respectively. In addition, we identified 35 novel alleles at 25 loci and 25 polymorphisms in the flanking regions at 17 STRs.ConclusionsOur data suggest that MPS‐ and CE‐derived alleles are compatible. MPS‐based analysis of the STR data substantially increased the allele diversity and improved the forensic parameters, which clearly demonstrated the advantages of MPS in comparison to CE. With more pooled data and larger‐scale validation, MPS could play a valuable role in forensic genetics and might be an additional tool for routine casework.

Evaluation of a Custom SNP Panel for Identifying and Rectifying of Misjudged Paternity in Deficiency Cases.

Parentage testing is routinely performed by genotyping short tandem repeat (STR) through capillary electrophoresis in the present. However, ambiguous or even misjudged paternity based on STRs happens from time to time in cases where only one putative parent is available. We analyzed STR data of 7,818,969 unrelated pairs and 75 close-relative pairs and found that although the probability of a random false match between non-relatives was 4.22 × 10–6, the incidence of false or ambiguous paternity results between children and first-degree relatives of their true parent was as high as 18.67%. These results highlight the risk of false inclusion of a relative or even non-relatives in parentage testing with STRs. We then validated all ambiguous STR results by targeted sequencing with a custom panel containing 4,830 individual identification single nucleotide polymorphisms (IISNP), found that the ratio of mismatch loci to total SNPs was 1.78–6.95% in close relatives compared with 10.93–13.49% in unrelated pairs. Last, we reported three real cases with undetermined paternity by STRs and rectified them by dissecting with our IISNP panel. These results suggested that high-density IISNP panel can be used to identify and rectify misjudged cases effectively.

Allelic diversity and forensic estimations of the Beijing Hans: Comparative data on sequence-based and length-based STRs

Short tandem repeat (STR) profiling is routinely used in forensic genetics. At present, STR analysis is mainly performed by capillary electrophoresis (CE). However, due to limitations associated with the CE method, STR genotyping has been limited to length polymorphisms only. Because next generation sequencing (NGS) is capable of providing full resolution STR data at the sequence variation level, the individual identification capability of forensic STR loci could be significantly improved. Here we present sequence-based STR data for the Beijing Han population in which 291 individuals were screened for 23 commonly used forensic STRs using the SeqTypeR24 CASE kit on an Ion PGM platform. In total, 234 length-based alleles and 356 sequence-based alleles, which included 22 novel core repeat sequences, were observed. The sequence-based matching probability and power of discrimination were superior to the length-based numbers for 16 loci bearing micro-variant alleles. Combined matching probability reached 8.2 × 10−29 for 23 STR loci at the sequence level. This was two orders of magnitude higher than the parameters at length level and provides a data base for sequence-based STR casework applications.

Large-scale identification of human bone remains via SNP microarray analysis with reference SNP database

Single nucleotide polymorphisms (SNPs) are valuable markers complementary to conventional forensic short tandem repeat (STR) markers in genetic typing, with potential advantages in challenging forensic casework. With the advent of high-throughput technologies, such as microarrays and massively parallel sequencing, the use of SNP typing has now expanded to large-scale forensic applications. Herein, a forensic case is presented to demonstrate the usefulness of SNP typing in identifying large-scale human bone remains with reference database construction. A total of 402 bone remains were recovered from an island in the Jeju Province of Korea where a massive disaster occurred in 1948. The first phase of the identification process was accomplished via conventional DNA typing methods including autosomal and Y-chromosomal STR typing, and mitochondrial DNA sequencing, which resulted in the identification of 74 of 402 remains. The second phase of the identification involved the remaining 327 unidentified remains using SNP typing as a supplementary tool based on Affymetrix resequencing array. The SNP markers of 782 family members were also analyzed and a reference database was constructed for comparison. An additional 51 bone remains were identified in the second phase. SNP data obtained from the supplementary genotyping yielded additional genetic information as well as contributed to kinship testing to determine the second degrees of relationship. In addition SNPs are useful in discriminating ambiguous relationship when only STR data are available. A software program developed for SNP typing system enabled efficient kinship analysis for large-scale forensic identification. The results and the casework are described and discussed.

Expanded CODIS STR allele frequencies - Evidence for the irrelevance of race-based DNA databases.

The US Federal Bureau of Investigation's (FBI) core Combined DNA Index System (CODIS) short tandem repeat (STR) panel is required for the calculations of random match probabilities (RMPs) in forensic DNA analysis. Current practice dictates that RMPs should be generated across appropriate reference STR allele frequency databases, including African American, Asian, Caucasian, Hispanic, and Native American, when the suspect's race is unknown. Should the suspect declare their race, a specific reference database that pertains to that designation is used. This practice is based on the presumption that racial population group is relevant for calculating conservative RMPs that favor the defendant. The core CODIS panel has been expanded to 20 STRs, however, the relationship between RMP and race has not been re-evaluated. Genetic structure analyses and Bayesian-based population assignment of expanded CODIS profiles from one race-neutral and five race-specific reference databases revealed that STR data could not distinguish races as distinct biological clusters. For instance, while the average race-specific RMPs for Hispanic or Caucasian profiles were almost equally-conservative when calculated from either population's reference database, the Hispanic profiles closely affined with the Native American population. Race-neutral RMPs computed with a correction factor (θ) of 0.03 favor the defendant as much as race-specific RMPs based on a θ of 0.01. Insufficient genetic differentiation observed among the US racial populations as well as inconsequential differences between race-specific and race-neutral RMPs undermine the value of using "race" in the context of forensic DNA analysis and support the argument that forensic databases should be race-neutral.