Results of a collaborative study of the EDNAP group regarding mitochondrial DNA heteroplasmy and segregation in hair shafts

A collaborative exercise was carried out by the European DNA Profiling Group (EDNAP) in order to evaluate the distribution of mitochondrial DNA (mtDNA) heteroplasmy amongst the hairs of an individual who displays point heteroplasmy in blood and buccal cells. A second aim of the exercise was to study reproducibility of mtDNA sequencing of hairs between laboratories using differing chemistries, further to the first mtDNA reproducibility study carried out by the EDNAP group. Laboratories were asked to type 2 sections from each of 10 hairs, such that each hair was typed by at least two laboratories. Ten laboratories participated in the study, and a total of 55 hairs were typed. The results showed that the C/T point heteroplasmy observed in blood and buccal cells at position 16234 segregated differentially between hairs, such that some hairs showed only C, others only T and the remainder, C/T heteroplasmy at varying ratios. Additionally, differential segregation of heteroplasmic variants was confirmed in independent extracts at positions 16093 and the poly(C) tract at 302-309, whilst a complete A-G transition was confirmed at position 16129 in one hair. Heteroplasmy was observed at position 16195 on both strands of a single extract from one hair segment, but was not observed in the extracts from any other segment of the same hair. Similarly, heteroplasmy at position 16304 was observed on both strands of a single extract from one hair. Additional variants at positions 73, 249 and the HVII poly(C) region were reported by one laboratory; as these were not confirmed in independent extracts, the possibility of contamination cannot be excluded. Additionally, the electrophoresis and detection equipment used by this laboratory was different to those of the other laboratories, and the discrepancies at position 249 and the HVII poly(C) region appear to be due to reading errors that may be associated with this technology. The results, and their implications for forensic mtDNA typing, are discussed in the light of the biology of hair formation.     

Results of collaborative study regarding the standardization of the Y-linked STR system DYS385 by the European DNA Profiling (EDNAP) group.

Y-chromosome linked short tandem repeat (STR) loci are inherited as a closely linked haplotype, which appears to remain stable in a given paternal lineage over many generations. In forensic cases, Y-linked STRs are particularly useful for the identification of human remains as well as in rape cases with mixed male/female stain samples. DYS385 is derived from tandemly duplicated segments of the Y chromosome thus giving rise to two fragments of variable length which do not behave like alleles but genotypes. The European DNA Profiling (EDNAP) group has carried out a collaborative exercise among 14 participating laboratories using DYS385 for typing of five unknown bloodstains and a control sample. Furthermore, population data from eight different European countries with samples sizes between 91 and 150 male individuals were collected. The results confirm previous observations that DYS385 is one of the most informative Y-linked STR loci. It could also be demonstrated that reproducible results can be obtained independently from the electrophoretic separation and detection methods used. Thus DYS385 may serve as a useful complementation to the routinely used autosomal STR systems in special cases.     

Results of a collaborative study of the EDNAP group regarding the reproducibility and robustness of the Y-chromosome STRs DYS19, DYS389 I and II, DYS390 and DYS393 in a PCR pentaplex format

A collaborative exercise was carried out by the European DNA Profiling Group (EDNAP) in the frame work of the STADNAP program, i.e. standardization of DNA profiling in Europe, in order to evaluate the performance of a Y-chromosome STR pentaplex, which includes the loci DYS19, DYS389 I and II, DYS390 and DYS393 and to determine whether uniformity of results could be achieved among different European laboratories.Laboratories were asked to analyze the five Y-STRs using singleplex and multiplex conditions in three bloodstains and one mixed stain (95% female and 5% male).All the laboratories reported the same results even for the mixed stain included in the exercise. This demonstrates the reproducibility and robustness of Y-chromosome STR typing even with multiplex formats and proves the usefulness of Y-STR systems for analyzing mixed stains with a male component.A total of 930 male samples from 10 different populations from Europe were also analysed for all the loci included in the pentaplex. Eight of these ten populations also included haplotype data.As for single gene analysis, haplotype diversity was higher in Germany and Italy and lower in Western European countries and Finland.Pairwise haplotype analysis shows the Finnish departure from the rest of the populations and a relatively homogeneity in the other European populations with F(ST) estimates lower than 0.05.UPGMA analysis shows an association of Western European population (Ireland, UK, Portugal and Galicia) on the one hand and central European populations on the other.     

Y-chromosome haplotypes in Italy: the GEFI collaborative database.

A sample of 1176 males from 10 Italian regions have been typed for DYS19, DYS389-I, DYS389-II, DYS390, DYS391, DYS392, DYS393, and DYS385. Individual haplotype data are available on line. A low degree of variation is present among regions. Use of this database is specifically recommended for forensic applications in Italy.     

RNA/DNA co-analysis from blood stains—Results of a second collaborative EDNAP exercise

A second collaborative exercise on RNA/DNA co-analysis for body fluid identification and STR profiling was organized by the European DNA Profiling Group (EDNAP). Six human blood stains, two blood dilution series (5-0.001 μl blood) and, optionally, bona fide or mock casework samples of human or non-human origin were analyzed by the participating laboratories using a RNA/DNA co-extraction or solely RNA extraction method. Two novel mRNA multiplexes were used for the identification of blood: a highly sensitive duplex (HBA, HBB) and a moderately sensitive pentaplex (ALAS2, CD3G, ANK1, SPTB and PBGD). The laboratories used different chemistries and instrumentation. All of the 18 participating laboratories were able to successfully isolate and detect mRNA in dried blood stains. Thirteen laboratories simultaneously extracted RNA and DNA from individual stains and were able to utilize mRNA profiling to confirm the presence of blood and to obtain autosomal STR profiles from the blood stain donors. The positive identification of blood and good quality DNA profiles were also obtained from old and compromised casework samples. The method proved to be reproducible and sensitive using different analysis strategies. The results of this collaborative exercise involving a RNA/DNA co-extraction strategy support the potential use of an mRNA based system for the identification of blood in forensic casework that is compatible with current DNA analysis methodology.     

Strategy of multiplex analysis of STR-loci polymorphism of Y-chromosome on the basis of "minimal haplotype"

A molecular-genetic analysis of Y chromosome is a convenient tool of paternal affinity determination. This method is now widely introduced in expert practice. Previous experience in typing of Y chromosome is outlined. Strategy of development and experience in operating a multiplex system for a simultaneous analysis of seven STR loci of Y chromosome is discussed.     

Forensic typing of autosomal SNPs with a 29 SNP-multiplex—Results of a collaborative EDNAP exercise

We report the results of an inter-laboratory exercise on typing of autosomal single nucleotide polymorphisms (SNP) for forensic genetic investigations in crime cases. The European DNA Profiling Group (EDNAP), a working group under the International Society for Forensic Genetics (ISFG), organised the exercise. A total of 11 European and one US forensic genetic laboratories tested a subset of a 52 SNP-multiplex PCR kit developed by the SNPforID consortium. The 52 SNP-multiplex kit amplifies 52 DNA fragments with 52 autosomal SNP loci in one multiplex PCR. The 52 SNPs are detected in two separate single base extension (SBE) multiplex reactions with 29 and 23 SNPs, respectively, using SNaPshot kit, capillary electrophoresis and multicolour fluorescence detection. For practical reasons, only the 29 SBE multiplex reaction was carried out by the participating laboratories. A total of 11 bloodstains on FTA cards including a sample of poor quality and a negative control were sent to the laboratories together with the essential reagents for the initial multiplex PCR and the multiplex SBE reaction. The total SNP locus dropout rate was 2.8% and more than 50% of the dropouts were observed with the poor quality sample. The overall rate of discrepant SNP allele assignments was 2.0%. Two laboratories reported 60% of all the discrepancies. Two laboratories reported all 29 SNP alleles in all 10 positive samples correctly. The results of the collaborative exercise were surprisingly good and demonstrate that SNP typing with SBE, capillary electrophoresis and multicolour detection methods can be developed for forensic genetics.     

A Spanish population study of 17 Y-chromosome STR loci

Haplotype, allele frequencies and population data of 17 Y-chromosome STR loci DYS19, DYS385, DYS389I, DYS389II, DYS390, DYS391, DYS392, DYS393, DYS437, DYS438, DYS439, DYS460 (GATA A7.1), DYS461 (GATA A7.2), GATA A10, GATA C4 and GATA H4 were determined from a sample of 148 unrelated male individuals from Spain. A total of 144 haplotypes were identified by the 17 Y-STR markers, of which 141 were unique, two were found in two individuals and one was found in three individuals. The haplotype diversity (99.95%) and discrimination capacity (97.30%) were calculated. Comparisons were made with previously published haplotype data on other Iberian population samples and no significant differences were found.     

A Peruvian population study of eight Y-chromosome STR loci

We have analyzed the distribution of the allele frequencies and haplotypes at eight Y-chromosomal short tandem repeat (STR) loci (DYS437, DYS438, DYS439, DYS460, DYS461, GATA A10, GATA C4 and GATA H4) in a sample population of 87 unrelated individuals from Perú.     

Analysis of artificially degraded DNA using STRs and SNPs--results of a collaborative European (EDNAP) exercise

Recently, there has been much debate about what kinds of genetic markers should be implemented as new core loci that constitute national DNA databases. The choices lie between conventional STRs, ranging in size from 100 to 450 bp; mini-STRs, with amplicon sizes less than 200 bp; and single nucleotide polymorphisms (SNPs). There is general agreement by the European DNA Profiling Group (EDNAP) and the European Network of Forensic Science Institutes (ENFSI) that the reason to implement new markers is to increase the chance of amplifying highly degraded DNA rather than to increase the discriminating power of the current techniques. A collaborative study between nine European and US laboratories was organised under the auspices of EDNAP. Each laboratory was supplied with a SNP multiplex kit (Foren-SNPs) provided by the Forensic Science Service, two mini-STR kits provided by the National Institute of Standards and Technology (NIST) and a set of degraded DNA stains (blood and saliva). Laboratories tested all three multiplex kits, along with their own existing DNA profiling technique, on the same sets of degraded samples. Results were collated and analysed and, in general, mini-STR systems were shown to be the most effective. Accordingly, the EDNAP and ENFSI working groups have recommended that existing STR loci are reengineered to provide smaller amplicons, and the adoption of three new European core loci has been agreed.     

A parameter study regarding the IBIS correlator

The electronic system IBIS has been used by numerous agencies worldwide as the standard tool to compare firearm markings on bullets and fired cartridges. There is a general interest among users concerning the likelihood with which the IBIS correlator may locate hits in its databases. Test results of the performance under different test conditions have been published in various papers. Experience has also been gained with the IBIS system from years of practical usage. All of these findings are difficult to compare with each other. No systematic presentation exists that actually shows the parameters upon which the success rate of the IBIS correlator depends. There has also been no mention of what values these parameters take on during each test. This paper first generally defines the success and error rates of the IBIS correlator. The parameters used will be discussed. Results of previously published tests will be re-examined based on this methodology. An illustrative form of presentation for the success rate of an electronic comparison system will also be suggested. It will be shown that the success rate of the IBIS correlator highly depends on the quality of the firearms-generated markings. It increases with the number of considered mark types, the number of available signatures per firearm, and the number of items inspected in the hit list. The success rate decreases with the database size. The paper will conclude with a series of practical recommendations for the setting up and successful operation of an electronic collection of ballistic evidence.     

Evaluation of Raman spectroscopy for the analysis of colored fibers: a collaborative study

A collaborative study on Raman spectroscopy was carried out by members of the ENFSI (European Network of Forensic Science Institutes) European Fibres Group (EFG) on three dyed fibers: two red acrylics and one red wool. Raman instruments from six different manufacturers were tested as well as nine different laser wavelengths ranging from blue (lambda = 458 nm) to near infrared-NIR (lambda = 1064 nm). This represents the largest comparison study of Raman analytical parameters carried out on identical fiber samples. For the chosen fiber and dye samples, red lasers (lambda = 633 and 685 nm) gave the poorest spectral quality whereas blue (458 nm), green (514 nm) and near infrared lasers (785, 830 and 1064 nm) provided average results. Blue (488 nm) and green lasers (532 nm) globally gave the best quality spectra. Fluorescence problems were often encountered with some of the excitation wavelengths and therefore a flexible Raman instrument equipped with different lasers can be recommended to measure forensic fiber samples. The instrument should also be equipped with a Raman microscope in order to be able to focus on a single fiber. This study shows that Raman spectroscopy usually enables the identification of the main dye present in a colored fiber; however, minor dye components are much more difficult to detect. SERRS (Surface Enhanced Resonance Raman Scattering) techniques give an improvement of the dye's spectral intensity but no spectral improvement was observed for the two red acrylic and red wool fibers tested.     

Y-chromosome STR haplotypes of Han ethnic group in Xi'an (NW China)

Allele and haplotype frequencies of six Y-chromosomal STR loci, namely DYS393, DYS19, DYS389II, DYS390, DYS391, and DYS385, were determined from 114 healthy unrelated males of Han ethnic group in Xi'an (NW China).     

浙江汉族人群5个Y-STR基因座的遗传多态性调查

目的　完善浙江省汉族人群Y STR基因座遗传多态性的频率调查 ,为法医学应用提供基础数据。方法　应用Y PLEXTM5荧光标记复合扩增系统 ,对浙江汉族 2 0 0名无关男性个体进行 5个STR基因座的复合扩增 ,用ABI310 0型基因分析仪对扩增产物进行检测 ,统计 5个Y STR基因座的群体遗传学参数。　结果DYS389Ⅰ、DYS389Ⅱ、DYS4 39、DYS4 38和DYS392基因座分别检出 4、7、5、4、6个等位基因 ,其GD值分别为0 5 86 8、0 7779、0 6 734、0 5 0 6 2和 0 6 32 8。观察到 5个Y STR基因座共同构成的单倍型 95种 ,其中有 5 4种单倍型只出现 1次 ,15种出现 2次 ,12种出现 3次 ,5种出现 4次 ,4种出现 5次 ,1种出现 6次 ,1种出现 7次 ,2种出现 8次 ,1种出现 11次 ,累计GD值为 0 992 0。结论　 5个Y STR基因座多态性分布较好 ,适合浙江法庭科学应用。     

唾液酯酶多态性在个体识别中的应用研究

目的 探讨唾液酯酶（ Set）多态性在法医学亲权鉴定和个体识别方面的应用价值。方法 应用聚丙烯酰胺凝胶盘状电泳及固蓝 RR染色方法,调查了 114名中国人 Set的表型分布及基因频率,用χ 2检验进行统计学分析。结果 中国人酯酶表型频率 Set F 22.81％, Set FS 50.88％, Set S 26.31％ ;基因频率为 SetF 0.482 5, SetS 0.517 5;非父排除机率为 0.187 5,个体识别率为 0.619 9。结论 Set有较高的父权排除率和个体识别率,可作为法医学亲权鉴定和个体识别的重要标记系统之一。     

DNA Commission of the International Society of Forensic Genetics: recommendations on forensic analysis using Y-chromosome STRs

During the past few years, the DNA Commission of the International Society of Forensic Genetics has published a series of documents providing guidelines and recommendations concerning the application of DNA polymorphisms to the problems of human identification. This latest report addresses a relatively new area — namely, Y-chromosome polymorphisms, with particular emphasis on short tandem repeats (STRs). This report addresses nomenclature, use of allelic ladders, population genetics and reporting methods.