An assessment of scientific and technical aspects of closed investigations of canine forensics DNA – case series from the University of California, Davis, USA

Aim To describe and assess the scientific and technical aspects of animal forensic testing at the University of California, Davis. The findings and recommendations contained in this report are designed to assess the past, evaluate the present, and recommend reforms that will assist the animal forensic science community in providing the best possible services that comply with court standards and bear judicial scrutiny. Methods A batch of 32 closed files of domestic dog DNA cases processed at the University of California, Davis, between August 2003 and July 2005 were reviewed in this study. The case files comprised copies of all original paperwork, copies of the cover letter or final report, laboratory notes, notes on analyses, submission forms, internal chains of custody, printed images and photocopies of evidence, as well as the administrative and technical reviews of those cases. Results While the fundamental aspects of animal DNA testing may be reliable and acceptable, the scientific basis for forensic testing animal DNA needs to be improved substantially. In addition to a lack of standardized and validated genetic testing protocols, improvements are needed in a wide range of topics including quality assurance and quality control measures, sample handling, evidence testing, statistical analysis, and reporting. Conclusion This review implies that although a standardized panel of short tandem repeat and mitochondrial DNA markers and publicly accessible genetic databases for canine forensic DNA analysis are already available, the persistent lack of supporting resources, including standardized quality assurance and quality control programs, still plagues the animal forensic community. This report focuses on closed cases from the period 2003-2005, but extends its scope more widely to include other animal DNA forensic testing services

Identifying rhesus macaque gene orthologs using heterospecific human CNV probes

AbstractWe used the Affymetrix® Genome-Wide Human SNP Array 6.0 to identify heterospecific markers and compare copy number and structural genomic variation between humans and rhesus macaques. Over 200,000 human copy number variation (CNV) probes were mapped to a Chinese and an Indian rhesus macaque sample. Observed genomic rearrangements and synteny were in agreement with the results of a previously published genomic comparison between humans and rhesus macaques. Comparisons between each of the two rhesus macaques and humans yielded 206 regions with copy numbers that differed by at least two fold in the Indian rhesus macaque and human, 32 in the Chinese rhesus macaque and human, and 147 in both rhesus macaques. The detailed genomic map and preliminary CNV data are useful for better understanding genetic variation in rhesus macaques, identifying derived changes in human CNVs that may have evolved by selection, and determining the suitability of rhesus macaques as human models for particular biomedical studies

Developmental Validation of Short Tandem Repeat Reagent Kit for Forensic DNA Profiling of Canine Biological Material

Aim To develop a reagent kit that enables multiplex polymerase chain reaction (PCR) amplification of 18 short tandem repeats (STR) and the canine sex-determining Zinc Finger marker. Methods Validation studies to determine the robustness and reliability in forensic DNA typing of this multiplex assay included sensitivity testing, reproducibility studies, intra- and inter-locus color balance studies, annealing temperature and cycle number studies, peak height ratio determination, characterization of artifacts such as stutter percentages and dye blobs, mixture analyses, species- specificity, case type samples analyses and population studies. Results The kit robustly amplified domesticated dog samples and consistently generated full 19-locus profiles from as little as 125 pg of dog DNA. In addition, wolf DNA samples could be analyzed with the kit. Conclusion The kit, which produces robust, reliable, and reproducible results, will be made available for the forensic research community after modifications based on this study’s evaluation to comply with the quality standards expected for forensic casework

Developmental Validation of Short Tandem Repeat Reagent Kit for Forensic DNA Profiling of Canine Biological Material

Aim To develop a reagent kit that enables multiplex polymerase chain reaction (PCR) amplification of 18 short tandem repeats (STR) and the canine sex-determining Zinc Finger marker. Methods Validation studies to determine the robustness and reliability in forensic DNA typing of this multiplex assay included sensitivity testing, reproducibility studies, intra- and inter-locus color balance studies, annealing temperature and cycle number studies, peak height ratio determination, characterization of artifacts such as stutter percentages and dye blobs, mixture analyses, species- specificity, case type samples analyses and population studies. Results The kit robustly amplified domesticated dog samples and consistently generated full 19-locus profiles from as little as 125 pg of dog DNA. In addition, wolf DNA samples could be analyzed with the kit. Conclusion The kit, which produces robust, reliable, and reproducible results, will be made available for the forensic research community after modifications based on this study’s evaluation to comply with the quality standards expected for forensic casework

Genetic diversity and population structure of long-tailed macaque (Macaca fascicularis) populations in Peninsular Malaysia

Background: The genetic diversity and structure of long-tailed macaques (Macaca fascicularis) in Peninsular Malaysia, a widely used non-human primate species in biomedical research, have not been thoroughly characterized. Methods: Thirteen sites of wild populations of long-tailed macaques representing six states were sampled and analyzed with 18 STR markers. Results: The Sunggala and Penang Island populations showed the highest genetic diversity estimates, while the Jerejak Island population was the most genetically discrete due to isolation from the mainland shelf. Concordant with pairwise Fst estimates, STRUCTURE analyses of the seven PCA-correlated clusters revealed low to moderate differentiation among the sampling sites. No association between geographic and genetic distances exists, suggesting that the study sites, including island study sites, are genetically if not geographically contiguous. Conclusions: The status of the genetic structure and composition of long-tailed macaque populations require further scrutiny to develop this species as an important animal model in biomedical research

Identification of constrained sequence elements across 239 primate genomes

Noncoding DNA is central to our understanding of human gene regulation and complex diseases1,2, and measuring the evolutionary sequence constraint can establish the functional relevance of putative regulatory elements in the human genome3–9. Identifying the genomic elements that have become constrained specifically in primates has been hampered by the faster evolution of noncoding DNA compared to protein-coding DNA10, the relatively short timescales separating primate species11, and the previously limited availability of whole-genome sequences12. Here we construct a whole-genome alignment of 239 species, representing nearly half of all extant species in the primate order. Using this resource, we identified human regulatory elements that are under selective constraint across primates and other mammals at a 5% false discovery rate. We detected 111,318 DNase I hypersensitivity sites and 267,410 transcription factor binding sites that are constrained specifically in primates but not across other placental mammals and validate their cis-regulatory effects on gene expression. These regulatory elements are enriched for human genetic variants that affect gene expression and complex traits and diseases. Our results highlight the important role of recent evolution in regulatory sequence elements differentiating primates, including humans, from other placental mammals

A global catalog of whole-genome diversity from 233 primate species.

The rich diversity of morphology and behavior displayed across primate species provides an informative context in which to study the impact of genomic diversity on fundamental biological processes. Analysis of that diversity provides insight into long-standing questions in evolutionary and conservation biology and is urgent given severe threats these species are facing. Here, we present high-coverage whole-genome data from 233 primate species representing 86% of genera and all 16 families. This dataset was used, together with fossil calibration, to create a nuclear DNA phylogeny and to reassess evolutionary divergence times among primate clades. We found within-species genetic diversity across families and geographic regions to be associated with climate and sociality, but not with extinction risk. Furthermore, mutation rates differ across species, potentially influenced by effective population sizes. Lastly, we identified extensive recurrence of missense mutations previously thought to be human specific. This study will open a wide range of research avenues for future primate genomic research

The landscape of tolerated genetic variation in humans and primates.

Personalized genome sequencing has revealed millions of genetic differences between individuals, but our understanding of their clinical relevance remains largely incomplete. To systematically decipher the effects of human genetic variants, we obtained whole-genome sequencing data for 809 individuals from 233 primate species and identified 4.3 million common protein-altering variants with orthologs in humans. We show that these variants can be inferred to have nondeleterious effects in humans based on their presence at high allele frequencies in other primate populations. We use this resource to classify 6% of all possible human protein-altering variants as likely benign and impute the pathogenicity of the remaining 94% of variants with deep learning, achieving state-of-the-art accuracy for diagnosing pathogenic variants in patients with genetic diseases