Autosomal STR loci for improving the kinship discrimination power for missing children and forensic application

Short tandem repeats (STRs) loci are very useful genomic markers with high power of individual discrimination, thus, they have been used for population genetics, forensic application and complex kinship analyses. In this study, we examined allele frequencies and forensic parameters for a total of 23 STR loci, that is, 17 established STRs with 13 CODIS core STR loci and D2S1338, D19S433, Penta E, and Penta D loci, and 6 new STRs (D10S1248, D22S1045, D2S441, D1S1656, D12S391, and SE33) in a sample of 545 unrelated individuals in South Korea. All loci were highly polymorphic and no significant departure from Hardy–Weinberg equilibrium was observed. The addition of 6 new loci to the 17 established STRs increased their power of discrimination by almost eight orders of magnitude (2.52 × 10^?20–4.44 × 10^?28) and improved the specificity of missing children database searches. Furthermore, we found several microvariant alleles at D2S441 and SE33 loci that have not been reported in the Korean population. We believe that this analysis will be useful for forensic application, deficiency paternity testing and expanding previously established Korean DNA databases.     

Polymorphic distribution and forensic effectiveness study of eight miniSTR in Chinese Uyghur ethnic group

We obtained the allelic frequencies and forensic efficiency data for eight mini short tandem repeat loci including Penta E, D12S391, D6S1043, D2S1338, D19S433, CSF1PO, Penta D and D19S253 loci from a sample of 128 unrelated Uyghur individuals from China. The amplification products of the eight STR loci are <240 bp in size. A total of 94 alleles were observed and the corresponding allelic frequencies ranged from 0.0039 to 0.3438 in the present study. Observed genotype distributions for each locus do not show deviations from Hardy–Weinberg equilibrium expectations. The combined power of discrimination, combined power of exclusion and combined matching probability of the eight STR loci equaled to 0.999999999963373, 0.9997770 and 3.6627 × 10^?11, respectively. Because of the small fragment length of PCR products and the high degree of polymorphisms, the eight STR loci are highly beneficial for the forensic analysis of degraded DNA samples which are commonly observed in forensic cases. The STR data of the Uyghur group were compared with the previously published population STR data of other groups from different ethnic or areas, and significant differences were observed among these groups at some loci.     

Forensic and population genetic analyses of the GlobalFiler STR loci in the Mongolian population

We have analyzed 24 loci including autosomal and Y-chromosomal short tandem repeats (STRs), Y-indel, and sex-determining marker in a sample of 267 unrelated individuals from the Mongolian population using the GlobalFiler? PCR Amplification Kit to provide an expanded and more reliable forensic database. Khalkh among 15 Mongolian minor-groups accounts for about 80% of the entire Mongolian population. A total of 267 different DNA profiles were found in this work. The highest gene diversity was observed in the SE33 (0.9376) locus, and the lowest value was found in the TPOX (0.6142) locus. Although individual power of discrimination estimates varied at the studied loci, combined probability of match from the 21 STR loci was estimated to be 1.139?×?10^?24, which is highly informative. Based on the results of pairwise F _ST genetic distances and multi-dimensional scaling plot showed that Mongolians were clustered into Europeans and Asians, although Mongolia is geographically located in Northeastern Asia. Thus, the present survey of the Mongolian population may help establish a comprehensive reference database for forensic and population genetic analyses.     

Direct estimate of the spontaneous germ line mutation rate in African green monkeys

Here, I provide the first direct estimate of the spontaneous mutation rate in an Old World monkey, using a seven individual, three‐generation pedigree of African green monkeys. Eight de novo mutations were identified within ～1.5 Gbp of accessible genome, corresponding to an estimated point mutation rate of 0.94 × 10^?8 per site per generation, suggesting an effective population size of ～12000 for the species. This estimation represents a significant improvement in our knowledge of the population genetics of the African green monkey, one of the most important nonhuman primate models in biomedical research. Furthermore, by comparing mutation rates in Old World monkeys with the only other direct estimates in primates to date–humans and chimpanzees–it is possible to uniquely address how mutation rates have evolved over longer time scales. While the estimated spontaneous mutation rate for African green monkeys is slightly lower than the rate of 1.2 × 10^?8 per base pair per generation reported in chimpanzees, it is similar to the lower range of rates of 0.96 × 10^?8–1.28 × 10^?8 per base pair per generation recently estimated from whole genome pedigrees in humans. This result suggests a long‐term constraint on mutation rate that is quite different from similar evidence pertaining to recombination rate evolution in primates.     

Long-term population genetic structure of an invasive urochordate: the ascidian Botryllus schlosseri

The accelerated pace of marine biological invasions raises questions pertaining to genetic traits and dynamics underlying the successful establishment of invasive species. Current research stresses the importance of multiple introductions and prolonged gene flow as the main sources for genetic diversity, which, along with genetic drift, affect invasive species success. We here attempt to determine the relative contribution of gene flow and mutation rates as sources of genetic variability using the invasive tunicate Botryllus schlosseri as a model. The study was performed over a 13-year period in the Santa Cruz Harbor, California. With a characteristic life history of five generations/year, the Santa Cruz Botryllus population has already experienced approximately 155 generations since the onset of its invasion. The results (278 specimens, 127 scored alleles, five microsatellite loci) support limited gene flow rate (2.89?×?10^?3) and relative genetic isolation. Furthermore, the study population was found to be influenced by both, genetic drift and a high mutation rate (2.47?×?10^?2). These findings were supported by high fluctuations in the frequencies of microsatellite alleles, the appearance of new alleles and the loss of others. The balance between genetic drift and a high mutation rate is further elucidated by the high, stable level of genetic variation. We suggest that rapid mutation rates at the microsatellite loci reflect genome-wide phenomena, helping to maintain high genetic variability in relatively isolated populations. The potential adaptability to new environments is discussed.     

Characterization of 11 polymorphic tetranucleotide microsatellites for forensic applications in California elk (Cervus elaphus canadensis)

Eleven polymorphic tetranucleotide microsatellite loci have been developed for forensic use in the protection of California elk. Based on a reference sample of elk taken from three races throughout California, the loci consist of 4–9 alleles (average 6.125). Probabilities of identity ( Paetkau et al. 1995 ) range from 0.079 to 0.288, with an overall probability of identity of 1.3 × 10^?9 (one in 7.8 × 10⁸).     

Genetic variation at five trimeric and tetrameric tandem repeat loci in four human population groups.

Trimeric and tetrameric short tandem repeats (STRs) represent a rich source of highly polymorphic markers in the human genome that may be studied with the polymerase chain reaction (PCR). We report the analysis of a multilocus genotype survey of 97-380 chromosomes in U.S. Black, White, Mexican-American, and Asian populations at five STR loci located on chromosomes 1, 4, 11, and X. The heterozygote frequencies of the loci ranged from 0.36 to 0.91 and the number of alleles from 6 to 20 for the 20 population and locus combinations. Relative allele frequencies exhibited differences between populations and unimodal, bimodal, and complex distributions. Although deviations were noted at some locus-population test combinations, genotype data from the loci were consistent overall with Hardy-Weinberg equilibrium by three tests. Population subheterogeneity within each ethnic group was not detected by two additional tests. No mutations were detected in a total of 860 meioses for two loci studied in the CEPH kindreds and five loci studied in other families. An indirect estimate of the mutation rates gave values from 2.3 x 10(-5) to 15.9 x 10(-5) for the five loci. Higher mutation rates appear to be associated with greater numbers of tandem repeats in the core motif. The most frequent genotype for all five loci combined appears to have a frequency of 7.59 x 10(-4). Together, these results suggest that trimeric and tetrameric STR loci are useful markers for the study of new mutations and genetic linkage analysis and for application to personal identification in the medical and forensic sciences.     

A large population genetic study of 15 autosomal short tandem repeat loci for establishment of Korean DNA Profile Database

Genotyping of highly polymorphic short tandem repeat (STR) markers is widely used for the genetic identification of individuals in forensic DNA analyses and in paternity disputes. The National DNA Profile Databank recently established by the DNA Identification Act in Korea contains the computerized STR DNA profiles of individuals convicted of crimes. For the establishment of a large autosomal STR loci population database, 1805 samples were obtained at random from Korean individuals and 15 autosomal STR markers were analyzed using the AmpFlSTR Identifiler PCR Amplification kit. For the 15 autosomal STR markers, no deviations from the Hardy-Weinberg equilibrium were observed. The most informative locus in our data set was the D2S1338 with a discrimination power of 0.9699. The combined matching probability was 1.521 × 10⁻¹⁷. This large STR profile dataset including atypical alleles will be important for the establishment of the Korean DNA database and for forensic applications.     

Mutation rate in human microsatellites: influence of the structure and length of the tandem repeat.

In 10,844 parent/child allelic transfers at nine short-tandem-repeat (STR) loci, 23 isolated STR mismatches were observed. The parenthood in each of these cases was highly validated (probability >99.97%). The event was always repeat related, owing to either a single-step mutation (n=22) or a double-step mutation (n=1). The mutation rate was between 0 and 7 x 10(-3) per locus per gamete per generation. No mutations were observed in three of the nine loci. Mutation events in the male germ line were five to six times more frequent than in the female germ line. A positive exponential correlation between the geometric mean of the number of uninterrupted repeats and the mutation rate was observed. Our data demonstrate that mutation rates of different loci can differ by several orders of magnitude and that different alleles at one locus exhibit different mutation rates.     

High genome-wide mutation rates in vegetatively propagated bermudagrass

A cascade DNA amplification strategy that generates arbitrary signatures from amplification profiles (ASAP) was used to measure genome-wide mutation rates in bermudagrass (Cynodon). ASAP quantified nucleotide changes that were induced by irradiation, genetic instabilities and normal vegetative growth of cultivars and accessions of sterile interspecific hybrids. DNA sequence divergence between cultivar ‘Tifway’ and its γ radiation-induced mutant ‘Tifway II’ (0.70 ± 0.66%) was comparable to estimates in radiation-induced mutants and spontaneous sports of chrysanthemum (Chrysanthemum morifolium Ramat.). A similar divergence in sequence (0.95 ± 0.20%) was observed in the pairwise comparison of 17 nondisjunctive ‘Tifgreen’ and ‘Tifdwarf’ accessions. Mutation during normal Tifdwarf vegetative growth was evaluated by planting sprigs and sampling their offspring. Somatic sequence divergence levels (0.004 ± 0.007%) resulted in a mutation rate of 1.05 × 10^?8 per nucleotide per generation, assuming that a bermudagrass sprig constitutes a generation of growth. These rates were comparable to those found in germinal cells and individuals of either human or Drosophila melanogaster, supporting the notion that eukaryotic evolution is generation rather than time dependent. The high accumulation of somatic mutations (10 per triploid genome) is consistent with a model whereby mutation load in a population exhibiting obligate vegetative reproduction is substantially higher than in a population under sexual or asexual reproduction. These constraints could be the cause of reported genetic instabilities in the Tifgreen–Tifdwarf complex. Finally, a long-term rate measured across accessions and indicative of the accumulation of mutations in 17 Tifgreen–Tifdwarf populations (µ = 1.02 × 10^?8 per nucleotide per generation) was strikingly congruent with the bermudagrass vegetative mutation rate, suggesting absence of evolutionary constraints in the sampled genomic regions. Mutation rates calculated from across-accesions divergence estimates (5.18 ± 0.53%) indicated that plant material was evolving 100 times faster (3.8 × 10^?7 changes per nucleotide per year) than a molecular clock rate estimate for grasses, probably resulting from the compound effect of clonal growth and life span of the hybrid plant material.     

Recurrent simple tandem repeat mutations during human Y-chromosome radiation in Caucasian subpopulations

The haplotypes at four polymorphic loci of theY chromosome were determined in 245 Caucasian males from 12 subpopulations. The data show that haplotype radiation occurred among Caucasians. Haplotype radiation was accompanied by recurrent mutations at STR loci that caused partial randomization of haplotype structure. The present distribution of alleles at short tandem repeats (STRs) can be explained by a mutation pattern similar to those described for autosomal STRs. The degree of variation among groups of subpopulations was assayed by using the Analysis of Molecular Variance. The results confirm a faster divergence of the Y chromosome as compared to the rest of the genome. Correspondence to: A. Novelletto     

Application of Short Tandem Repeat markers in diagnosis of chromosomal aneuploidies and forensic DNA investigation in Pakistan

Purpose

Short Tandem Repeat (STR) genetic markers hold great potential in forensic investigations, molecular diagnostics and molecular genetics research. AmpFlSTR® Identifiler™ PCR amplification kit is a multiplex system for co-amplification of 15 STR markers used worldwide in forensic investigations. This study attempts to assess forensic validity of these STRs in Pakistani population and to investigate its applicability in quick and simultaneous diagnosis and tracing parental source of common chromosomal aneuploidies.

Methodology

Samples from 554 healthy Pakistani individuals from 5 different ethnicities were analyzed for forensic parameters using Identifiler STRs and 74 patients' samples with different aneuploidies were evaluated for diagnostic strengths of these markers.

Results

All STRs hold sufficient forensic applicability in Pakistani population with paternity index between 1.5 and 3.5, polymorphic information content from 0.63 to 0.87 and discrimination power ≥ 0.9 (except TPOX locus). Variation from Hardy–Weinberg equilibrium was observed at some loci reflecting selective breeding and intermarriages trend in Pakistan. Among aneuploidic samples, all trisomies were precisely detectable while aneuploidies involving sex chromosomes or missing chromosomes were not clearly detectable using Identifiler STRs. Parental origin of aneuploidy was traceable in 92.54% patients.

Conclusion

The studied STR markers are valuable tools for forensic application in Pakistan and utilizable for quick and simultaneous identification of some common trisomic conditions. Adding more sex chromosome specific STR markers can immensely increase the diagnostic and forensic potential of this system.     

Determining microsatellite genotyping reliability and mutation detection ability: an approach using small-pool PCR from sperm DNA

Microsatellite genotyping from trace DNA is now common in fields as diverse as medicine, forensics and wildlife genetics. Conversely, small-pool PCR (SP-PCR) has been used to investigate microsatellite mutation mechanisms in human DNA, but has had only limited application to non-human species. Trace DNA and SP-PCR studies share many challenges, including problems associated with allelic drop-out, false alleles and other PCR artefacts, and the need to reliably identify genuine alleles and/or mutations. We provide a framework for the validation of such studies without a multiple tube approach and demonstrate the utility of that approach with an analysis of microsatellite mutations in the tammar wallaby (Macropus eugenii). Specifically, we amplified three autosomal microsatellites from somatic DNA to characterise efficiency and reliability of PCR from low-template DNA. Reconstruction experiments determined our ability to discriminate mutations from parental alleles. We then developed rules to guide data interpretation. We estimated mutation rates in sperm DNA to range from 1.5 × 10⁻² to 2.2 × 10⁻³ mutations per locus per generation. Large multi-step mutations were observed, providing evidence for complex mutation processes at microsatellites and potentially violating key assumptions in the stepwise mutation model. Our data demonstrate the necessity of actively searching for large mutation events when investigating microsatellite evolution and highlight the need for a thorough understanding of microsatellite amplification characteristics before embarking on SP-PCR or trace DNA studies.     

Population data for 17 short tandem repeat loci on Y chromosome in northern Croatia

Human Y-short tandem repeats (STRs) are tandem repeat arrays of two to seven base pair units on non-recombining region (NRY) of the human Y chromosome. Studies on Y-STR are interesting in both population genetics and forensics. The aim of this study was to investigate the population genetic properties of 17 STR loci on Y chromosome in the northern Croatia region. We carried out a statistical analysis of the data from previously performed genetic analysis collected during routine forensic work by the Forensic Science Centre “Ivan Vu?eti?”. A total of 220 unrelated healthy men from northern Croatia were selected for the purpose of this study. Genomic DNA was extracted using Chelex procedure from FTA^® cards. Y-chromosomal STRs were determined using the AmpFISTR Yfiler PCR amplification kit. The haplotype frequencies were determined by direct counting and analyzed using Arlequin 3.1 and analysis of molecular variance calculated with the Y chromosome haplotype reference database online analysis tool. A total of 210 haplotypes were identified, 200 of which were unique. Total haplotype diversity was 0.995. Locus diversity varied from 0.331 for DYS392 to 0.783 for DYS385 locus. Allele frequencies diversity was 0.662. Discrimination capacity was 95.7%. The use of European minimal haplotype set indicated the most resemblance of this population to the Croatian capital of Zagreb, with modest resemblance to Bosnia and Herzegovina, Serbia and Hungary. This article provides the first overview of the Y chromosome STR variability in northern Croatia, thus providing the referent point for any future forensic and genetic epidemiology efforts in this region.     

Thirty polymorphic microsatellite loci from the critically endangered kakapo (Strigops habroptilus)

Thirty polymorphic microsatellite loci were developed from the critically endangered kakapo (Strigops habroptilus), using an enriched genomic library. Characterization of loci using 90 kakapo revealed an average of 3.3 alleles per locus (range: 2–5) and an average expected heterozygosity of 0.47 (range: 0.17–0.70). The probability of identity (7.2 × 10^?15) and probability of exclusion (0.999999) demonstrate that these loci are a highly informative marker set that can aid the genetic management of the kakapo.     

Mutation and selection processes regulating short tandem repeats give rise to genetic and phenotypic diversity across species

Short tandem repeats (STRs) are units of 1–6 bp that repeat in a tandem fashion in DNA. Along with single nucleotide polymorphisms and large structural variations, they are among the major genomic variants underlying genetic, and likely phenotypic, divergence. STRs experience mutation rates that are orders of magnitude higher than other well-studied genotypic variants. Frequent copy number changes result in a wide range of alleles, and provide unique opportunities for modulating complex phenotypes through variation in repeat length. While classical studies have identified key roles of individual STR loci, the advent of improved sequencing technology, high-quality genome assemblies for diverse species, and bioinformatics methods for genome-wide STR analysis now enable more systematic study of STR variation across wide evolutionary ranges. In this review, we explore mutation and selection processes that affect STR copy number evolution, and how these processes give rise to varying STR patterns both within and across species. Finally, we review recent examples of functional and adaptive changes linked to STRs.     

Mexican mestizo population sub-structure: effects on genetic and forensic statistical parameters

Since Mexican mestizos are an admixed population, it is necessary to determine the effects that the substructure of the population has on genetic and forensic parameters. With this aim, a study was performed with 15 STR loci (CODIS plus D2S1338 and D19S433) on 1,640 unrelated Mexican mestizos. We determine allele and genotypic frequencies observing departure from Hardy–Weinberg expectation (12 out of 15 loci, with an excess of homozygotes, Fis?>?0), as well as pairs of loci in an apparent linkage disequilibrium (13 of 92 loci). We conducted a test for genetic population stratification, the results show that the Mexican mestizo population is substructured into three subgroups, which are in HW and linkage equilibrium. The combination of the 15 loci in the whole population has high forensic efficiency with the capacity to genetically discriminate one individual in one quintillion (1/10¹⁸). Our data potentially validates the use of these 15 STR loci to establish forensic identity and parentage testing for legal purposes, and offers a powerful tool for genetic variation analysis. However, given that the population is stratified, we highly recommend applying a correction with the inbreeding coefficient in calculations of paternity and forensic studies to avoid erroneous assumptions.     

The migratory history of anadromous and non-anadromous tapertail anchovy Coilia nasus in the Yangtze River Estuary revealed by the otolith Sr:Ca ratio

The migratory history of tapertail anchovy Coilia nasus in the Yangtze River Estuary, China was investigated using otolith Sr:Ca ratios and two-dimensional images of the Sr level from an X-ray electron probe microanalyzer (EPMA). The results showed that 17 of the 22 young-of-the-year (YOY) specimens had low Sr:Ca ratios (1.2–2.4?×?10^?3;1.5?±?0.3?×?10^?3) at the central otolith area, indicating their riverine origin and initial freshwater residence. In addition, 11 of the 14 adult specimens had low Sr:Ca ratios (1.3–2.2?×?10^?3; 1.7?±?0.4?×?10^?3) at the central otolith area but showed alternating changes between high (>4.0?×?10^?3) and low (<2.5?×?10^?3) values outside of this region, reflecting their riverine origin and the migration between freshwater and estuarine habitats. These 28 specimens represented the anadromous population in this region. The other 5 YOY specimens had high Sr:Ca ratios (3.6–5.9?×?10^?3; 4.8?±?0.8?×?10^?3) throughout the life history. Similarly, the other 3 adult specimens had high Sr:Ca ratios (4.0–5.7?×?10^?3; 4.8?±?0.7?×?10^?3) at the central otolith area but showed alternating changes between low and high values outside this region, suggesting that estuarine-origin non-anadromous individuals occurred in this region. The average of the otolith Sr:Ca ratios and Sr level mapping along the life-history transects could be used as a scalar for charting the migratory history of the tapertail anchovy in the Yangtze River Estuary: <2.0?×?10^?3 for freshwater residence and 3.5–6.0?×?10^?3 for estuarine residence.     

Human health risk from exposure to metals through fish and shellfish ingestion in Saudi Arabia

Abstract

In this study, human exposure and risks of metals through fish ingestion were predicted. Concentrations of 10 metals (Cd, As, Cr, Cu, Hg, Mn, Ni, Pb, V, and Zn) in 17 commonly eaten fish species in Saudi Arabia were characterized. Using the fish ingestion patterns, chronic daily intakes of metals among the Saudi and expatriate populations were predicted to be in the ranges of 8.89?×?10^?06–1.08?×?10^?02 and 6.59?×?10^?06–7.97?×?10^?03?mg/kg/day, respectively. The average cancer risks from inorganic arsenic were 2.76?×?10^?05 and 2.09?×?10^?05 for Saudi and expatriate, and the ranges were 1.61?×?10^?09–1.58?×?10^?03 and 2.64?×?10^?09–1.27?×?10^?03, respectively. The predicted risks were much lower than the previously reported risks. There were 47.4% and 42.4% chances that cancer risks would be higher than 1.0?×?10^?05 (10 per million) among Saudi and expatriate, respectively. The average cumulative hazard index (HI) for Saudi and expatriate were 0.324 and 0.239 with the ranges of 0.0142–7.26 and 0.017–6.43, respectively. Approximately 3.06% and 1.56% cases among Saudi and expatriate had HI greater than unity, indicating possible health concern, respectively. Through comprehensive understanding of exposure and risks, strategies can be adopted to protect human health.     

pH effects on growth and lipid accumulation of the biofuel microalgae Nannochloropsis salina and invading organisms

Biofuels derived from non-crop sources, such as microalgae, offer their own advantages and limitations. Despite high growth rates and lipid accumulation, microalgae cultivation still requires more energy than it produces. Furthermore, invading organisms can lower efficiency of algae production. Simple environmental changes might be able to increase algae productivity while minimizing undesired organisms like competitive algae or predatory algae grazers. Microalgae are susceptible to pH changes. In many production systems, pH is kept below 8 by CO₂ addition. Here, we uncouple the effects of pH and CO₂ input, by using chemical pH buffers and investigate how pH influences Nannochloropsis salina growth and lipid accumulation as well as invading organisms. We used a wide range of pH levels (5, 6, 7, 8, 9, and 10). N. salina showed highest growth rates at pH 8 and 9 (0.19?±?0.008 and 0.19?±?0.011, respectively; mean ± SD). Maximum cell densities in these treatments were reached around 21 days into the experiment (95.6?×?10⁶?±?9?×?10⁶ cells mL^?1 for pH 8 and 92.8?×?10⁶?±?24?×?10⁶ cells mL^?1 for pH 9). Lipid accumulation of unbuffered controls were 21.8?±?5.8 % fatty acid methyl esters content by mass, and we were unable to trigger additional significant lipid accumulation by manipulating pH levels at the beginning of stationary phase. Ciliates (grazing predators) occurred in significant higher densities at pH 6 (56.9?±?39.6?×?10⁴ organisms mL^?1) than higher pH treatments (0.1–6.8?×?10⁴ organisms mL^?1). Furthermore, the addition of buffers themselves seemed to negatively impact diatoms (algal competitors). They were more abundant in an unbuffered control (12.7?±?5.1?×?10⁴ organisms mL^?1) than any of the pH treatments (3.6–4.7?×?10⁴ organisms mL^?1). In general, pH values of 8 to 9 might be most conducive to increasing algae production and minimizing invading organisms. CO₂ addition seems more valuable to algae as an inorganic carbon source and not as an essential mechanism to reduce pH.