STR analysis of artificially degraded DNA-results of a collaborative European exercise

Degradation of human DNA extracted from forensic stains is, in most cases, the result of a natural process due to the exposure of the stain samples to the environment. Experiences with degraded DNA from casework samples show that every sample may exhibit different properties in this respect, and that it is difficult to systematically assess the performance of routinely used typing systems for the analysis of degraded DNA samples. Using a batch of artificially degraded DNA with an average fragment size of approx. 200 bp a collaborative exercise was carried out among 38 forensic laboratories from 17 European countries. The results were assessed according to correct allele detection, peak height and balance as well as the occurrence of artefacts. A number of common problems were identified based on these results such as strong peak imbalance in heterozygous genotypes for the larger short tandem repeat (STR) fragments after increased PCR cycle numbers, artefact signals and allelic drop-out. Based on the observations, strategies are discussed to overcome these problems. The strategies include careful balancing of the amount of template DNA and the PCR cycle numbers, the reaction volume and the amount of Taq polymerase. Furthermore, a careful evaluation of the results of the fragment analysis and of automated allele calling is necessary to identify the correct alleles and avoid artefacts.     

Results of Y-SNP typing in three different populations

Y-SNPs are additional markers which came in the focus of forensic scientists since several years. The presented study uses 29 of these polymorphisms to detect the distribution of the haplogroups. Samples of three different European populations (one population is located in Far East, the Primorye Region of Russia) previously typed with Y-STRs were investigated. The results show the high potential of these markers to get information about the geographic origin of the paternal line.     

Population data of Y-chromosomal STRs in Russian males of the Primorye region population

Data of eight Y-chromosomal STRs, the so called "minimal core set", were obtained from 152 unrelated males of the Primorye region of Russia. The allelic frequencies correspond to other European populations. The background is a settlement of males from the European part of Russia, Ukraine and other states which were included in the former western part of the Soviet Union. On the other hand the distribution of the most frequent haplotypes differs to the Ukraine and Russian population. The most frequent haplotype was obtained five times in the population corresponding to 3.3%. The haplotype data reported here have been included into the Y-STR database maintained at the Institute of Legal Medicine, Humboldt-University, Berlin.     

ISFG: Recommendations on biostatistics in paternity testing

The Paternity Testing Commission (PTC) of the International Society for Forensic Genetics has taken up the task of establishing the biostatistical recommendations in accordance with the ISO 17025 standards and a previous set of ISFG recommendations specific to the genetic investigations in paternity cases. In the initial set, the PTC recommended that biostatistical evaluations of paternity are based on a likelihood ratio principle – yielding the paternity index, PI. Here, we have made five supplementary biostatistical recommendations. The first recommendation clarifies and defines basic concepts of genetic hypotheses and calculation concerns needed to produce valid PIs. The second and third recommendations address issues associated with population genetics (allele probabilities, Y-chromosome markers, mtDNA, and population substructuring) and special circumstances (deficiency/reconstruction and immigration cases), respectively. The fourth recommendation considers strategies regarding genetic evidence against paternity. The fifth recommendation covers necessary documentation, reporting details and assumptions underlying calculations. The PTC strongly suggests that these recommendations should be adopted by all laboratories involved in paternity testing as the basis for their biostatistical analysis.     

Population genetics of Y-chromosomal microsatellites in Baltic males

Y-chromosomal microsatellites (STRs) are potentially useful in forensic practice but, in contrast to autosomal systems, large and diverse population databases are required in order to facilitate the statistical evaluation of donor-stain matches. Since appropriate data from the Baltic region have so far been lacking, blood samples were obtained from 430 males originating from one of the three Baltic states and these samples were genotyped using a previously described "extended core set" of nine Y-STR marker systems. Allele frequency distributions and discrimination indices were calculated, and the three populations were tested for genetic differences by means of analysis of molecular variance (AMOVA). A larger genetic difference became apparent between Estonian and both Lithuanian and Latvian males than between the latter two, non-Finno-Ugric speaking populations. The haplotype data reported here have been included into the Y-STR database maintained at the Institute of Legal Medicine, Humboldt University, Berlin.     

Population data of Y-chromosomal STRs in lithuanian, latvian and estonian males

Data of nine Y-chromosomal STRs, the so called “extended core set”, were obtained from 152 unrelated males from Lithuania, 145 from Latvia and 133 from Estonia. The haplotype data reported here have been included into the Y-STR database maintained at the Institute of Legal Medicine, Humboldt-University, Berlin.     

A new method for the evaluation of matches in non-recombining genomes: application to Y-chromosomal short tandem repeat (STR) haplotypes in European males

A 9-locus microsatellite framework (minimal haplotype), previously developed for forensic purposes so as to facilitate stain analysis, personal identification and kinship testing, has been adopted for the establishment of a large reference database of male European Y-chromosomal haplotypes. The extent of population stratification pertaining to this database, an issue crucial for its practical forensic application, was assessed through analysis of molecular variance (AMOVA) of the 20 regional samples included. Despite the notion of some significant haplotype frequency differences, which were found to correlate with known demographic and historic features of Europeans, AMOVA generally revealed a high level of genetic homogeneity among the populations analyzed. Owing to their high diversity, however, accurate frequency estimation is difficult for Y-STR haplotypes when realistic (i.e. moderately sized) datasets are being used. As expected, strong pair-wise and higher order allelic associations were found to exist between all markers studied, implying that haplotype frequencies cannot be estimated as products of allele frequencies. A new extrapolation method was therefore developed which treats haplotype frequencies as random variables and generates estimates of the underlying distribution functions on the basis of closely related haplotypes. This approach, termed frequency 'surveying', is based upon standard population genetics theory and can in principle be applied to any combination of markers located on the Y-chromosome or in the mitochondrial genome. Application of the method to the quality assured reference Y-STR haplotype database described herein will prove very useful for the evaluation of positive trace-donor matches in forensic casework.     

Population genetics of ACTBP2 (SE33) in Western Saxony (Germany)

In order to apply a useful STR system we performed a population study in Western Saxony (Germany). The allele distribution was investigated in a sample of 431 unrelated adults. In addition, 170 families from routine paternity cases were examined for the presence of meiotic mutations, and two mutations were observed.     

Fatal trauma to the neck: immunohistochemical study of local injuries

As fatal trauma to the neck is often associated with short survival times, proof of vitality may often be difficult using standard histochemical techniques. Soft tissue neck injuries resulting from strangulation by ligature or manual strangulation were examined immunohistochemically using antibodies to myoglobin, fibronectin, C5b-9 and MRP14, and compared to controls consisting of accidental soft tissue neck injuries as well as undamaged neck soft tissue. Although survival times in the study and control groups were unknown and certainly some individual variation may be expected in the time course of normal wound development, both the study and control groups demonstrated similar time courses in the immunohistochemical detection of antigen. Myoglobin was always found in those samples in which only one antigen was shown to be involved in an injury-specific pattern; myoglobin and fibronectin were found in samples with dual antigen involvement. Samples involving three antigens always included C5b-9 in addition to myoglobin and fibronectin. The single positive MRP14 sample in the study and control groups was simultaneously positive for the other markers used. Myoglobin, fibronectin, C5b-9 and MRP14 are therefore suitable for immunohistochemical detection of vital reactions and estimation of temporal relationships in the early posttraumatic period after neck trauma.