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This paper describes the development and validation of a novel 31-locus, 6-dye STR multiplex system which is designed to meet the needs of the rapidly growing Chinese forensic database. This new assay combines 20 extended-CODIS core loci (D3S1358, D5S818, TPOX, CSF1PO, TH01, vWA, D7S820, D21S11, D8S1179, D18S51, D16S539, D13S317, FGA, D1S1656, D2S441, D2S1338, D10S1248, D12S391, D19S433 and D22S1045), 9 highly polymorphic loci in Chinese Han population (D3S3045, D6S1043, D6S477, D8S1132, D10S1435, D15S659, D19S253, Penta D and Penta E) and two gender determining markers, amelogenin and Y-Indel, which could amplify DNA from extracts, as well as direct amplification from substrates. To demonstrate the suitability for forensic applications, this system was validated by precision and accuracy evaluation, concordance tests, case sample tests, sensitivity, species specificity, stability, stutter calculation and DNA mixtures, according to the guidelines described by the Scientific Working Group on DNA Analysis Methods (SWGDAM) and regulations published by the China Ministry of Public Security. The validation results indicate the robustness and reliability of this new system, and it could be a potentially helpful tool for human identification and paternity testing in the Chinese population, as well as facilitating global forensic DNA data sharing. Additional supporting information may be found in the online version of this article at the publisher's web-site. Color online: See article online to view Figs. 1-5 and 7 in color. This article is protected by copyright. All rights reserved.

This study assessed the usefulness of DNA quantification to predict the success of historical samples when analyzing SNPs, mtDNA, and STR targets. Thirty burials from six historical contexts were utilized, ranging in age from 80 to 800 years postmortem. Samples underwent library preparation and hybridization capture with two bait panels (FORCE and mitogenome), and STR typing (autosomal STR and Y-STR). All 30 samples generated small (~80 bp) autosomal DNA target qPCR results, despite mean mappable fragments ranging from 55–125 bp. The qPCR results were positively correlated with DNA profiling success. Samples with human DNA inputs as low as 100 pg resulted in ≥80% FORCE SNPs at 10X coverage. All 30 samples resulted in mitogenome coverage ≥100X despite low human DNA input (as low as 1 pg). With PowerPlex Fusion, ≥30 pg human DNA input resulted in >40% of auSTR loci. At least 59% of Y-STR loci were recovered with Y-target qPCR-based inputs of ≥24 pg. The results also indicate that human DNA quantity is a better predictor of success than the ratio of human to exogenous DNA. Accurate quantification with qPCR is feasible for historical bone samples, allowing for the screening of extracts to predict the success of DNA profiling.

The extremely high nature of transmissibility and severity of infection due to the novel Corona virus is a serious threat to mankind. This letter delivers a cau-tion for forensic DNA experts in the era of COVID-19 infection from the Indian perspective. Samples are routinely transported to laboratories without any spe-cific guidelines. Therefore, this is high time to formu-late clear guidelines for the handling of biological ma-terial, from receiving to processing in the laboratory during such a pandemic. © 2021. AJFSFM.

Deoxyribonucleic acid (DNA) profiling, has had a tremendous impact on forensic genetics. Before DNA profiling, all forensic genetic casework (e.g., Paternity testing, criminal casework, individual identification) was performed using classical serological genetic markers. Blood groups, human leukocyte antigen (HLA), and polymorphic protein and enzymes were used for solving forensic genetic casework using immunological and electrophoretic methodologies. These genetic markers were nevertheless limited when it was necessary to analyze minimal or degraded material, which is commonly involved in forensic cases. An STR is a region of human DNA containing an array of tandem repeats. Arrays range from only a 10 to about a hundred repeated units. This essay confers the basic concepts of operating of DNA in the criminal investigation. This review primarily summarizes the major tandem repeat markers used in forensic DNA profiling, that assist criminal’s conviction, exonerate the inferring individuals, and recognize victims of violence, catastrophes, and armed conflict.

Current analysis of forensic DNA stains relies on the probabilistic interpretation of bulk-processed samples that represent mixed profiles consisting of an unknown number of potentially partial representations of each contributor. Single-cell methods, in contrast, offer a solution to the forensic DNA mixture problem by incorporating a step that separates cells before extraction. A forensically relevant single-cell pipeline relies on efficient direct-to-PCR extractions that are compatible with standard down- stream forensic reagents. Here we demonstrate the feasibility of implementing single-cell pipelines into the forensic process by exploring four metrics of electropherogram (EPG) signal quality—i.e., allele detection rates, peak heights, peak height ratios, and peak height balance across low- to high-molecular-weight short tandem repeat (STR) markers—obtained with four direct-to-PCR extraction treatments and a common post-PCR laboratory procedure. Each treatment was used to extract DNA from 102 single buccal cells, whereupon the amplification reagents were immediately added to the tube and the DNA was amplified/injected using post-PCR conditions known to elicit a limit of detection (LoD) of one DNA molecule. The results show that most cells, regardless of extraction treatment, rendered EPGs with at least a 50% true positive allele detection rate and that allele drop-out was not cell independent. Statistical tests demonstrated that extraction treatments significantly impacted all metrics of EPG quality, where the Arcturus® PicoPure™ extraction method resulted in the lowest median allele drop-out rate, highest median average peak height, highest median average peak height ratio, and least negative median values of EPG sloping for GlobalFiler™ STR loci amplified at half volume. We, therefore, conclude the feasibility of implementing single-cell pipelines for casework purposes and demonstrate that inferential systems assuming cell independence will not be appropriate in the probabilistic interpretation of a collection of single-cell EPGs.

The assignment of the number of contributors (N) to a forensic DNA profile is undertaken as part of the interpretation process. There is no requirement for N to be the same for both propositions wi...

The knowledge of the ancestry of an unknown individual from forensic DNA may provide important information for law enforcement investigation. This information can be acquired from biological traces by means of massive parallel sequencing (MPS) technologies and developed arrays of ancestry informative DNA markers (AIM panels). Development of forensic AIM panels takes two steps: (1) obtainment of information on DNA polymorphism of individuals from different populations; (2) selection of polymorphic DNA markers (autosomal, Y-, X-chromosomal) and their validation. The necessity of keeping medical secrecy for the protection of human rights may limit the usage of ancestry informative markers suitable for forensic needs if they are also associated with genetic diseases and physical traits. The present article deals with the analysis of published strategies for selecting markers capable of differentiating populations to design a panel of ancestry informative markers and applying it in forensic analysis in the Republic of Belarus.

The samples used for forensic DNA analysis in living individuals are usually blood and buccal swabs, however, blood collection requires an invasive method that can cause discomfort, thus a buccal swab can be a good choice for individuals examined, especially children. This study aimed to determine the effect of temperature and storage time of buccal swabs on the quantity of DNA as material for DNA examination in the forensic field. This study was a laboratory experiment to determine the effect after treatment. Buccal swab samples were 48 and divided into 2 temperature groups, namely room temperature (RT) and 4℃. The division of the temperature groups was also observed with time differences, namely 1, 3, 5, 7 days. EDNA extraction used the DNAzol method and DNA quantification used a Spectrophotometer. The PCR process was carried out with STR primers FGA and D13S317 loci. The visualization stage used acrylamide gel and silver staining. The results of this study prove that there is an effect of temperature and storage time of buccal swab samples. The longer the treatment time, the lower the DNA level. With statistical analysis, it is obtained p-value of

Pieces of evidence have continued to emerge, demonstrating the extensive efficiency and effectiveness of the DNA database in assisting criminal investigations around the world. Therefore, the present study aimed to determine the awareness level on the prominent role of Forensic DNA Database on Crime Investigation in Nigeria: a case study of Benin City. In conducting this research, a total of 458 questionnaires were distributed around Benin City between the periods of 12th January 2020 to 21st March 2020, with a particular focus on security agents and students. The questionnaire comprised of three main categories: Socio-demographic characteristics, Information about the National Forensic DNA Database, and Information about DNA evidence, and Nigeria Criminal Justice system. For the analysis of data collected; the statistical tool used was also Statistical Package for Social Sciences, version 22 for windows. Responses were compared using chi-square and presented as counts and percentages. In determining the level of awareness, the following responses were obtained. Of the total population: 53.28% had no idea about forensics, 19.21% were uncertain and 27.54% knew about forensics. The same trend was observed with Forensic DNA profiling, 42.14% did not know, 22.27% were uncertain and 35.59% demonstrated good knowledge of Forensic DNA profiling. On the knowledge about the National Forensic DNA Database, 48.47% had no knowledge, 22.27% were uncertain and 29.26% were knowledgeable about it. The result of the present study revealed that the awareness level of the forensic DNA Database was found to be inadequate.

DNA profiling is just one of many tools available to police in a criminal investigation. However, unlike any other criminal investigative tool, DNA profiling has captured the public imagination. It...

Contamination and degradation are known challenges for reliable genotyping, since they can cause, among other problems, false microsatellite profiles. In this study we described a method to decrease the proportion of false microsatellite profiles from fish scale samples of endangered allis shads (Alosa alosa) from a reintroduction program, where cross-contamination with DNA from other individuals and potentially degradation of samples occurred. To maximize the portion of reliably measurable results, we modified and combined two known approaches—thresholds used in forensic DNA analyses and a multiple-tubes approach. This combined approach increased reliable microsatellite profiles compared with single approaches. The forensic thresholds and the multiple-tubes approach increased the measurable results from 55 to 67% and 75%, respectively, whereas the combined approach accomplished an increase to 90%. This illustrates the potential of the combined approach for other studies with comparable problems or sample material.

Forensic DNA phenotyping (FDP) based on massive parallel sequencing (MPS) is an emerging technique within forensic genetics that enables the prediction of an individual's externally visible characteristics (EVCs) from DNA. Because of its achievements, FDP has become one of the most useful additional tools for aiding police investigations to narrow down the investigative pool in different types of forensic cases. Eye, hair and skin colour can now be predicted reliably and with practically useful accuracy. However, FDP has not yet been routinely implemented in the forensic science field due to, principally, the lack of complete genetic knowledge of pigmentation and facial traits and the lower predictability of intermediate phenotypes. Furthermore, in some countries its application has given rise to a number of ethical, social and legal issues, the latter being the most restrictive barrier to the implementation of FDP.

A impunidade tem despontado como um dos principais motivos do aumento da criminalidade no Brasil, principalmente devido à dificuldade dos processos investigativos em apontar a autoria de delitos. A indisponibilidade e a falta de transparência de dados da segurança pública e a alta taxa de subnotificação nos registros oficiais são alguns entraves que dificultam a avaliação da eficiência das políticas de segurança pública. Para o agravamento da situação, amostras coletadas em locais de crime pela Perícia Criminal podem não ser prontamente armazenadas nos bancos de DNA forense, ou quando o são, na maioria das vezes, não se obtém perfil genético compatível com os já cadastrados, diminuindo, assim, os índices nacionais de elucidação de autoria dos crimes. Dessa forma, o objetivo do artigo é apresentar um histórico sobre a implementação e evolução dos bancos de dados de perfis genéticos no Brasil. O processo de construção do banco de dados de DNA forense demonstra um avanço na qualidade das investigações policiais, encaixando-se perfeitamente como meio de prova no processo penal, somando-se às demais evidências necessárias à persecução penal. As técnicas de identificação baseadas na análise do DNA estão cada vez mais sendo empregadas no Brasil em amostras relacionadas a vestígios biológicos coletados em locais de crime, porém longe do patamar alcançado em países como os EUA e Reino Unido, que possuem milhões de perfis genéticos em seus bancos de dados, o que reflete nos seus altos índices de elucidação de crimes.

Massively parallel sequencing (MPS) of mitochondrial (mt) DNA allows practitioners the ability to fully resolve heteroplasmic sites. In forensic DNA analysis, identifying heteroplasmy (a naturally occurring mixture of two mtDNA profiles) can provide additional mtDNA profile information which can lead to an increase in the discrimination potential of an mtDNA match between an evidentiary sample and reference source. Forensic samples such as hair and skeletal remains, especially older, more compromised samples, can often exhibit DNA damage. Because both damage and heteroplasmy can manifest as a mixture of two nucleotides, it is important to differentiate between the two conditions when interpreting mtDNA MPS data. In this study, DNA damage was applied under controlled conditions to samples containing a range of template concentrations, including some with identified heteroplasmy. Damage was applied via storage in water at room temperature on samples diluted before or after storage to mimic low template scenarios. Damage was assessed with respect to the following areas: mtDNA quantification and degradation ratios, MPS read depth, MPS profile results, overall damage rates, and the interpretation of heteroplasmy. Datasets were generated to assess and compare two different amplification and library preparation strategies: the Promega PowerSeq™ CRM Nested System kit and a 1.16 kb target amplicon of the entire mtDNA control region followed by a Nextera® XT library preparation. The results of this study provide an evaluation of the Promega 10-plex MPS procedure as an improved process to mitigate the impact of mtDNA damage on low template samples. Some of the negative effects of damage observed in this study were a decrease in mtDNA yield by 20–30% and lower quality MPS sequencing results. These effects were observed more frequently when samples were diluted prior to inducing damage, illustrating that low template samples are more susceptible to damage. The findings of this study will assist forensic laboratories in differentiating between damage and heteroplasmy, which is essential when developing robust mtDNA MPS interpretation guidelines such as setting appropriate reporting thresholds.

Background Forensic DNA phenotyping has gained momentum in the recent past due to the prediction of externally visible characters (EVCs) from the biological sample. The most common phenotypes like eye, hair, and skin color are predicted from the biological samples using a web-based system called IrisPlex. Based on six genetic SNPs, the IrisPlex system is developed and validated for its prediction accuracy in diverse ethnic groups worldwide. In previous studies, this system proved to have significant prediction accuracy. The EVCs vary substantially based on different geographical locations. Hence, the objective of this study was to validate the accuracy of the IrisPlex system in predicting the eye colors in the Iraqi population. Methods Six genetic single-nucleotide polymorphisms SNPs (HERC2-rs12913832, OCA2- rs1800407, SLC24A4-rs12896399, SLC45A2- rs16891982, TYR-rs1393350, and IRF4- rs12203592) in 58 Iraqi subjects were performed using Sequenom MassARRAY Genotyping. According to Liu et al., a predicted probability of 0.7 was considered as the threshold. Results Participants in this study of brown color were observed in 44.83%, intermediate in 43.1%, and blue in 12.07%. Completely predictive accuracy is obtained in 1; we observed the AUC at threshold 0.7 was 0.91 for brown, 0.79 for blue, and 0.60 for intermediate eye color. The sensitivity was 42.85% for blue, 0% for intermediate eye color, and 100% for brown-colored eye. Specificity was 100% for blue, 100% for intermediate, and 78.13% for brown eye color. Conclusion Hence, it was concluded that the prediction accuracy of the IrisPlex system for blue and brown color eye in the Iraqi population is significant in the studied population size. However, a pivotal study with larger sample size is required to represent the prediction accuracy of the IrisPlex system in the whole Iraqi population.

Increasingly in courts, the source of forensic DNA trace evidence is not challenged by the defence. Instead, the issue relates to how the suspect’s DNA got to the crime scene, suggesting it arrived...

Forensic DNA phenotyping refers to an emerging field of forensic sciences aimed at the prediction of externally visible characteristics of unknown sample donors directly from biological materials. The aging process significantly affects most of the above characteristics making the development of a reliable method of age prediction very important. Today, the so-called "epigenetic clocks" represent the most accurate models for age prediction. Since they are technically not achievable in a typical forensic laboratory, forensic DNA technology has triggered efforts toward the simplification of these models. The present study aimed to build an epigenetic clock using a set of methylation markers of five different genes in a sample of the Italian population of different ages covering the whole span of adult life. In a sample of 330 subjects, 42 selected markers were analyzed with a machine learning approach for building a prediction model for age prediction. A ridge linear regression model including eight of the proposed markers was identified as the best performing model across a plethora of candidates. This model was tested on an independent sample of 83 subjects providing a median error of 4.5 years. In the present study, an epigenetic model for age prediction was validated in a sample of the Italian population. However, its applicability to advanced ages still represents the main limitation in forensic caseworks.

Understanding the relative success of recovering DNA profiles from different touched evidentiary items/substrates, and between different methods of collection, is critical for optimal targeting of ...

Missing data is pertinent to criminal networks due to the hidden nature of crime. Generally, researchers evaluate the impact of incomplete network data by extracting or adding nodes and/or edges from a known network. Statistics on this reduced or completed network are then compared with statistics from the known network. In this study, we integrate police data on known offenders with DNA data on unknown offenders. Statistics from the integrated dataset (‘known network’) are compared with statistics from the police data (‘reduced network’). Networks with both known and unknown offenders are bigger but also have a different structure to networks with only known offenders.

Forensic DNA Phenotyping (FDP) encompasses an emerging set of technologies aimed at predicting physical characteristics of unknown suspects from crime scene DNA traces. In its application FDP invol...

Forensic DNA testing can play a critical role in homicide investigations. Selecting the appropriate evidence on which to perform DNA testing requires foresight and reasoning based on experience and science. Although successful DNA testing can occur using many substrates, including blood, hair, and sweat/epithelial cells, positive results can also result from testing various unorthodox samples. The authors report on a triple-murder investigation where DNA testing of dog feces at the crime scene matched DNA testing of feces found on the shoe of a suspect resulting in successful prosecution of the case.

Mitochondrial DNA (mtDNA) is a useful tool in forensic investigation as it provides information about the matrilineal ancestry of individuals. In addition, mtDNA can be analyzed when the analysis of other nuclear markers is underperforming. Recently, we developed a minisequencing panel for the simultaneous analysis of 52 mtDNA SNPs to classify maternal lineages into the main haplogroups and their phylogeographic origin. In order to make this panel suitable for forensic genetics laboratories, a validation study has been performed in accordance with the Scientific Working Group on DNA Analysis Methods (SWGDAM) guidelines, including species specificity, reproducibility, sensitivity, and stability tests. The results demonstrate that the panel of 52 mtDNA SNPs is highly sensitive, since it enables to obtain complete genetic profiles of samples containing minimal amounts of DNA (1 pg). Furthermore, it provides sufficient genetic information to detect the matrilineal biogeographical origin of highly degraded samples, i.e., ancient dating skeletal remains, and samples with the presence of inhibitors, such as hematin and humic acid. In addition, this panel can detect mixtures in samples whose mtDNA haplogroups of contributors are different. Overall, the results of this study demonstrate the suitability of this minisequencing panel of 52 mtDNA SNPs to be used in forensic cases, with samples of low amount or degraded DNA.

This review paper covers the forensic-relevant literature in biological sciences from 2016 to 2019 as a part of the 19th Interpol International Forensic Science Managers Symposium. The review papers are also available at the Interpol website at: https://www.interpol.int/content/download/14458/file/Interpol%20Review%20Papers%202019.pdf.

Due to recent improvements in forensic DNA testing kit sensitivity, there has been an increased demand in the criminal justice community to revisit past convictions or cold cases. Some of these cases have little biological evidence other than touch DNA in the form of archived latent fingerprint lift cards. In this study, a previously developed optimised workflow for this sample type was tested on aged fingerprints to determine if improved short tandem repeat (STR) profiles could be obtained. Two-year-old samples processed with the optimised workflow produced an average of approximately five more STR alleles per profile over the traditional method. The optimised workflow also produced detectable alleles in samples aged out to 28 years. Of the methods tested, the optimised workflow resulted in the most informative profiles from evidence samples more representative of the forensic need. This workflow is recommended for use with archived latent fingerprint samples, regardless of the archival time.Key points The use of the optimised workflow on aged archived latent fingerprint (ALFP) lift card samples (aged 2–28 years) improves the number of STR alleles recovered, providing more discriminatory STR profiles than those processed using the traditional workflow. Interpretable STR alleles can be detected from ALFP lift card samples stored as long as 28 years when the optimised procedures are followed. The use of individual laboratory-sterilised tools for sample preparation and the addition of a re-purification step with Centri-Sep columns in the recommended optimised workflow seem to limit the ability to detect low-level secondary DNA sources.

The estimation of chronological age from biological fluids has been an important quest for forensic scientists worldwide, with recent approaches exploiting the variability of DNA methylation patterns with age in order to develop the next generation of forensic ‘DNA intelligence’ tools for this application. Drawing from the conclusions of previous work utilising massively parallel sequencing (MPS) for this analysis, this work introduces a DNA methylation-based age estimation method for blood that exhibits the best combination of prediction accuracy and sensitivity reported to date. Statistical evaluation of markers from 51 studies using microarray data from over 4000 individuals, followed by validation using in-house generated MPS data, revealed a final set of 11 markers with the greatest potential for accurate age estimation from minimal DNA material. Utilising an algorithm based on support vector machines, the proposed model achieved an average error (MAE) of 3.3 years, with this level of accuracy retained down to 5 ng of starting DNA input (~ 1 ng PCR input). The accuracy of the model was retained (MAE = 3.8 years) in a separate test set of 88 samples of Spanish origin, while predictions for donors of greater forensic interest (< 55 years of age) displayed even higher accuracy (MAE = 2.6 years). Finally, no sex-related bias was observed for this model, while there were also no signs of variation observed between control and disease-associated populations for schizophrenia, rheumatoid arthritis, frontal temporal dementia and progressive supranuclear palsy in microarray data relating to the 11 markers.