Mismatch distribution analysis of Y-STR haplotypes as a tool for the evaluation of identity-by-state proportions and significance of matches—the European picture

We suggest the use of the mismatch distribution methodology as an easy way to estimate the distance between all pairs of haplotypes present in a sample. This approach allows the evaluation of the proportion of pairs of Y-STR haplotypes that are prone to become identical by state (IBS), in one generation, by recurrent mutation, a statistic of major importance in the forensic field. The mismatch approach presents some advantages alternatively to the empirical one, since it is not necessary to have simultaneous information on STRs and SNPs, and it allows the evaluation of IBS also within-haplogroups. The estimation of IBS at an European scale showed that there is a high population substructuring for this parameter, increasing from southern-central European countries towards west and north, in accordance to what was found for Y-biallelic markers. This result seems to imply a more careful use of large databases for matching evaluation, even in the absence of population structure for general Y-STR diversity. Furthermore, mismatch distribution can be used to measure the distance between a particular haplotype and all the haplotypes in a sample. When applied to the most frequent haplotypes in Europe it revealed that the opportunity for IBS is not directly related to the frequency of a haplotype, but highly dependent on the proportion of neighbouring haplotypes—so, that reporting on the haplotype frequency for evaluating the significance of a match can be misleading.     

用Y-STR单倍型信息指导数据库采样分析

目的分析浙江省绍兴诸暨市（县级市）各镇、村和姓的Y-STR单倍型分布，为Y_STR数据库的采样与应用提供依据。方法采集诸暨市17个镇156村的55个姓氏，且各镇一村有同姓人员10人以上（含）的家族样本5903份男性个体血样。采用YfilerTM复合扩增试剂盒进行17个Y-STR分型，所得数据进行镇（乡／街道）／村／姓氏的组合和镇（乡／街道）／村／姓氏／单倍型组合分布情况统计分析。结果在5903份男性样本中，获得1987种Y-STR单倍型，它们分布于235种镇（乡／街道）／＋-t／姓氏组合，共构成2547种镇（乡／街道）／村／姓氏／单倍型组合。各单倍型对应的“镇／村／姓”组合次数出现从1到18次不等，其中有1686种单倍型对应1种镇一村一姓组合（84．9％），绝大部分的单倍型对应1～2种镇一村一姓组合（95．3％）。各镇／村中同姓人员出现的主流分型大部分为1～2种（90．7％）。在镇／村的同姓人员中出现次要分型的频率平均为18．22％。结论Y-STR数据库在诸暨的采样在家族调查的基础上，应针对次要分型较多的姓增加采样量，减少遗漏风险，获得高质量的YSTR数据库。     

Y染色体STR的银染复合扩增

目的建立一套Y染色体STR的复合扩增体系,检测中国藏族人群的单倍型分布。方法利用复合扩增的方法扩增DYS434、DYS443和DYS456三个基因座,利用聚丙烯酰胺凝胶电泳银染进行分型,检测西藏藏族101名无关男性个体单倍型分布。结果三个基因座在藏族样本中分别检测出4、4、6个等位基因,共检测出31种单倍型,其单倍型的变异度是0.9481,标准误为0.0049。结论Y-STR的复合扩增在法医学的亲权鉴定和个人识别中有重要的作用。     

Allele frequencies and haplotypes of 12 Y-STR loci for the local Chinese population in Hong Kong

Haplotype frequencies were established for 12 Y-chromosome STR loci, including all loci recommended by Scientific Working Group on DNA Analysis Methods Y-STR Subcommittee (DYS391, DYS389I/II, DYS439, DYS393, DYS390, DYS385a/b, DYS438, DYS19 and DYS392) plus DYS437, in the local Chinese population in Hong Kong. In a sample of 481 unrelated males, it was possible to define 424 different haplotypes of which 388 were unique, 26 was found in two individuals, 2 were shared in three individuals, 5 were shared in four individuals and 3 were shared in five individuals. The allele diversity values for each locus ranged from 0.4273 (DYS438) to 0.9555 (DYS385a/b). The observed haplotype diversity value and discrimination capacity were 0.9992 and 0.8815, respectively. In a genetic study of these unrelated males, triple alleles were found at the DYS358 locus in six individuals. The combined Y-chromosome STR polymorphisms provide a powerful discrimination tool for routine forensic applications.     

Allele sequences of six new Y-STR loci and haplotypes in the Chinese Han population

Human chromosome Y-specific short tandem repeat (Y-specific STR) markers have useful properties for forensic applications. However, there is a need to develop more Y-specific STR markers, because the discriminating power of each STR locus is limited. In the present study, we describe our results on six new Y-specific STR markers that were initially located using sequence database information by Ayub et al. and were named DYS434, DYS435, DYS436, DYS437, DYS438 and DYS439. Our studies focused on the analysis of the DNA sequence for each allele at all six Y-specific STR loci in order to understand their structures in the human genome and to construct human allelic ladders, which are necessary for forensic DNA typing. In addition, the haplotype distribution for all six analyzed loci was studied in a Chinese Han population sample. The results indicate that DYS434, DYS435, DYS436, DYS437, DYS438 and DYS439 are useful Y-specific STR markers for forensic sciences.     

Diversity of 26-locus Y-STR haplotypes in a Nepalese population sample: isolation and drift in the Himalayas

Twenty-six Y-chromosomal short tandem repeat (STR) loci were amplified in a sample of 769 unrelated males from Nepal, using two multiplex polymerase chain reaction (PCR) assays. The 26 loci gave a discriminating power of 0.997, with 59% unique haplotypes, and the highest frequency haplotype occurring 12 times. We identified novel alleles at four loci, microvariants at a further two, and nine examples of amelogenin-Y deletions (1.2%). Comparison with a similarly sized Bhutanese sample typed with the same markers suggested histories of isolation and drift, with drift having a greater effect in Bhutan. Extended (11-locus) haplotypes for the Nepalese samples have been submitted to the Y-STR Haplotype Reference Database (YHRD).     

中国汉族及日本群体7个Y—STR位点及单倍型的遗传多态性和群体差异

目的调查Y染色体7个STR位点及单倍型的遗传多态性并分析其群体差别。方法应用PCR、变性聚丙烯酰胺凝胶电泳结合银染显色分型技术,检测45例中国汉族及59例日本男性DNA样品。结果在DYS393、DYS389Ⅰ、DYS19、DYS390、DYS389Ⅱ、DYS392等6个位点中共检出33个等位基因,DYS385位点检出39个等位基因组,其频率分布在0.0169～0.6444之间,DP值分布在0.5406～0.9579之间,以DYS385位点最高。7个位点数据综合比较,二组群体间在遗传学上存在显著性差异P<0.05。由7个位点组成的单倍型有95种,中国汉族有41种,DP值为0.9960,日本群体有54种,DP值为0.9965,2群体间未发现相同的单倍型。结论上述7个STR位点属于高鉴别能力位点,单倍型具有很高的遗传多态性并显示出明显的民族特征。     

Development and evaluation of multiplex Y-STR assays for application in molecular genealogy

Three multiplex PCRs were developed for the analysis of 14 single-copy and 4 multi-copy Y chromosome Short Tandem Repeat (STR) loci routinely used by several public genealogical databases. These assays were used in addition to PowerPlex^® Y for the analysis of 245 DNA samples from a genealogical project. In total 244 different haplotypes composed of 37–40 alleles were identified with one haplotype identical between two males with the same surname. The multi-copy loci DYS464 and DYS724 were the most polymorphic with a gene diversity of at least 0.964. The use of DYS454 and DYS455 can be questioned as these loci had the lowest gene diversity (0.039 and 0.269, respectively).     

Blind field test evaluation of Raman spectroscopy as a forensic tool

Analytical instrumentation for Raman spectroscopy has advanced rapidly in recent years to the point where commercial field-portable instruments are available. Raman analysis with portable instrumentation is a new capability that can provide emergency response teams with on-site evaluation of hazardous materials. Before Raman analysis is accepted and implemented in the field, realistic studies applied to unknown samples need to be performed to define the reliability of this technique. Studies described herein provide a rigorous blind field test that utilizes two instruments and two operators to analyze a matrix that consists of 58 unknown samples. Samples were searched against a custom hazardous materials reference library (Hazardous Material Response Unit (HMRU) Spectral Library Database). Experimental design included a number of intentionally difficult situations including binary solvent mixtures and a variety of compounds that yield medium-quality spectra that were not contained in the HMRU library. Results showed that over 97% of the samples were correctly identified with no occurrences of false positive identifications (compounds that were not in the library were never identified as library constituents). Statistical analysis indicated equivalent performance for both the operators and instruments. These results indicate a high level of performance that should extrapolate to actual field situations. Implementation of Raman techniques to emergency field situations should proceed with a corresponding level of confidence.     

STR melting curve analysis as a genetic screening tool for crime scene samples

In this proof-of-concept study, high-resolution melt curve (HRMC) analysis was investigated as a postquantification screening tool to discriminate human CSF1PO and THO1 genotypes amplified with mini-STR primers in the presence of SYBR Green or LCGreen Plus dyes. A total of 12 CSF1PO and 11 HUMTHO1 genotypes were analyzed on the LightScanner HR96 and LS-32 systems and were correctly differentiated based upon their respective melt profiles. Short STR amplicon melt curves were affected by repeat number, and single-source and mixed DNA samples were additionally differentiated by the formation of heteroduplexes. Melting curves were shown to be unique and reproducible from DNA quantities ranging from 20 to 0.4 ng and distinguished identical from nonidentical genotypes from DNA derived from different biological fluids and compromised samples. Thus, a method is described which can assess both the quantity and the possible probative value of samples without full genotyping.     

The application of chemometrics on Infrared and Raman spectra as a tool for the forensic analysis of paints

The aim of this work is to evaluate the capabilities and limitations of chemometric methods and other mathematical treatments applied on spectroscopic data and more specifically on paint samples. The uniqueness of the spectroscopic data comes from the fact that they are multivariate - a few thousands variables - and highly correlated. Statistical methods are used to study and discriminate samples. A collection of 34 red paint samples was measured by Infrared and Raman spectroscopy. Data pretreatment and variable selection demonstrated that the use of Standard Normal Variate (SNV), together with removal of the noisy variables by a selection of the wavelengths from 650 to 1830 cm(-1) and 2730-3600 cm(-1), provided the optimal results for infrared analysis. Principal component analysis (PCA) and hierarchical clusters analysis (HCA) were then used as exploratory techniques to provide evidence of structure in the data, cluster, or detect outliers. With the FTIR spectra, the Principal Components (PCs) correspond to binder types and the presence/absence of calcium carbonate. 83% of the total variance is explained by the four first PCs. As for the Raman spectra, we observe six different clusters corresponding to the different pigment compositions when plotting the first two PCs, which account for 37% and 20% respectively of the total variance. In conclusion, the use of chemometrics for the forensic analysis of paints provides a valuable tool for objective decision-making, a reduction of the possible classification errors, and a better efficiency, having robust results with time saving data treatments.     

The distribution of natural stable isotopes of carbon as a possible tool for the differentiation of samples of TNT.

TNT samples from different countries are shown to have different 13C/12C ratios. The delta13C spread (against the PDB standard) is about 6.5%. 13C/12C ratios can therefore be utilized in order to correlate or discriminate between samples of TNT.     

The New Zealand DNA databank: its development and significance as a crime solving tool.

The New Zealand DNA Databank was established following the introduction of legislation in August 1996. Using the Second Generation Multiplex (SGM), DNA profiles from over 13,000 convicted offenders and volunteer donors have been completed to the National DNA Database. Since June 1998, DNA profiles from over 1,400 unsolved crimes have been entered onto the Crime Sample Database. Of all unsolved crimes analysed, 33% are linked to individuals and 21% are linked to other unsolved crimes. Several high profile types of case including homicides, sexual offenses and burglaries are amongst those regularly solved.     

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.