Marina Dobosz

Coordinating Y-chromosomal STR research for the courts

The growing interest of contemporary biology in the human Y chromosome (Hammer 1995; Jobling and TylerSmith 1995) has extended to forensic circles. This trend emerges from a consideration of the increasing number of papers published in forensic science journals concerned in qualifying polymorphisms of human Y chromosomes as tools for molecular identification. Jobling et al. (1997) list a number of forensic issues to which the male-specific profiles could be applied and discuss their appropriateness and suitability. In view of the inherent court application, we wish to discuss some critical aspects of this new class of DNA profiles and issue some guidelines for those researchers who are considering to prepare and submit forensic YSTR work to the International Journal of Legal Medicine.

Expansion to full mutation of a FMR1 intermediate allele over two generations

Fragile X syndrome is due to an expanded CGG repeat in the 5′ UTR of the FMR1 gene. According to repeat size, we distinguish four allele categories: normal (<40 CGG), intermediate (46–60 CGG), premutated (55–200 CGG) and full mutated (>200 CGG). However, the boundaries among these categories are unclear, making it difficult to classify unstable alleles and to estimate the risk of expansion. We report a family with a proband, carrying a methylated full mutation with an amplification of 1.2 kb. PCR analysis demonstrated two alleles of 29 and 61 CGGs in the mother. Sequencing of the 61 CGG allele showed no AGG interruptions. Both mothers sisters had two alleles of 31 and 44 CGGs, and the daughter of one of these had two alleles of 22 and 44 repeats, demonstrating stable transmission of the 44 CGG allele. The maternal grandfather was deceased, but haplotype reconstruction using markers DXS548 and FRAXAC1 demonstrated that he was carrier of the premutated allele. Furthermore, molecular analysis confirmed the same paternity with a probability of 99.79% for all the three sisters. According to these findings, it is likely that the maternal grandfather carried the 44 CGG allele, showing unstable transmission, given that it expanded first to 61 CGGs in one daughter, and then to full mutation in her child. Although we cannot exclude paternal mosaicism, it is likely that a rare event of progression from an intermediate to a premutated and on to a full mutated allele occurred in this family over two generations.

Increased forensic efficiency of a STR-based Y-specific haplotype by addition of the highly polymorphic DYS385 locus

Abstract The polymorphic short tandem repeat (STR) locus DYS385 mapping to the male-specific region of human Y chromosome, was used to reinvestigate 125 unrelated Italian males, from our data archive, who had been previously typed for 7 different Y-specific STRs (DYS19, DYS389 I and II, DYS390, DYS391, DYS392, DYS393), defining a haplotype now widely adopted in the forensic context. The aim of this study was to improve the information value of the original haplotype in view of its application to issues of personal identification and parental analysis. DYS385 proved to be highly polymorphic (94.5% gene diversity) and the overall individualization capacity of the 8-loci haplotype was raised to 93.6%, with 117 unique assets out of 125 tested samples.

Clinal Variation in the Nuclear DNA of Europeans

Abstract Allele frequencies are clinally distributed for many protein polymorphisms in Europe, suggesting that the current populations are derived from an ancestral group that expanded from the Near East. It is not yet fully established whether that expansion took place during the Neolithic or earlier or whether the detectable protein variation faithfully reflects the underlying molecular variation. In this study we address the latter question by describing geographic patterns of genetic diversity at seven highly polymorphic DNA markers. Two of these markers are minisatellites, four are microsatellites, and the seventh is a locus of the HLA system. By analyzing a database of 304 samples, with more than 130,000 chromosomes, we found evidence for a major clinal component of genetic variation. At most loci spatially close populations resemble each other genetically, and the degree of genetic similarity, as measured by spatial autocorrelation statistics, decreases at increasing distances. The observed patterns of molecular variation do not seem to differ qualitatively from those identified for protein polymorphisms. This suggests that low levels of population structuring, described in some mitochondrial DNA studies, may reflect different evolutionary histories for nuclear and maternally inherited markers or, alternatively, that spatial patterns of mitochondrial DNA variation may need more sensitive statistical methods to be recognized.

Estimating allele frequencies of hypervariable DNA systems

Several polymorphisms of human DNA have been shown to be hypervariable due to the recurrence of a variable number of tandem repeats (VNTRs) in the lengths of allelic restriction fragments. The recurrence of allelic variants in this novel class of polymorphisms seems to comply well with a model of continuous random variables. Based on this assumption, we have compiled some simple algorithms for classification of continuous data and estimation of classes of relative frequencies and have implemented these routines for the management of databases storing hypervariable single locus DNA genetic systems. The algorithms are compiled in BASIC language and can be incorporated in task-oriented computer programs. Three procedures are discussed, based in turn on: (a) using predetermined, arbitrary classes; (b) point estimations of frequencies for single fragments using error measurements associated with the kilobase value assignment; (c) estimates of phenotype frequencies according to error measurements. Error measurements are obtained from a statistic of values pertaining to several restriction fragments (genomic controls) repeatedly tested in different experiments. Problems related to these approaches are discussed.

A repository of 14 PCR-loci Italian gene frequencies in the world wide web

A collection of 6830 typing results produced by the Immunohematology Laboratory at the UCSC, pertaining to 11 STRs (FES/FPS, vWA31, HUMTH01, F13A1, MBP, D21S11, D7S460, D18S51, CD4, TPOX, CSF1PO) and 3 AmpFLPs (D1S80, APO-B, COL2A1), is publicly available as an electronic archive at a website.

Exclusion of an individual charged with rape by allele-specific DNA profiling on fetal tissues

A mentally handicapped young woman was raped by an unknown individual, who repeatedly abused of her. The crime was discovered only after she fell pregnant and aborted. By gestational age (16–18 weeks), the fetus was assumed to have been conceived when the woman was hospitalized. Hencefrom, a nurse was suspected to be her rapist, and was prosecuted. During the trial, our laboratory was asked to perform DNA profiles over some fetal tissues, the woman and the suspect. Following the analysis detailed here, evidence was given that the alleged rapist could not be the father.

The dark side of the UK National DNA Database.

Sir—Adrian Linacre (May 31, p 1841) is an enthusiastic proponent of the UK’s National DNA Database (NDNAD). We wish to offer a more critical perspective of the database. In 2002, a young girl was murdered in a Tuscan village by a foreigner. From a bloodstain left at the crime scene, the suspect’s DNA profile was produced and circulated on the Interpol net. In February, 2002, P N Hankin, a barman in England was identified from the NDNAD and charged with the murder. He denied the charge and protested that he had been at his workplace on the day of the murder, which was corroborated by several eye-witnesses. He was, however, arrested. Eventually, British officers reported a fresh profile from a sample volunteered by Hankin, with one previously unseen mismatch. He was subsequently cleared of all charges. This case highlights some of the limitations of DNA profiling. Inferring a person has committed a crime on the basis of a profile match alone is problematic. The rest of the evidence should not be overlooked in favour of the DNA match. The use of DNA databases does not prevent forensic scientists from misrepresenting the population associated with a crime (the forensic population). A forensic population is not an abstract category, but a small group of individuals identified by background evidence. In the past, concerns have been raised about the existence of genetic isolates and the forensic use of gene frequency multiplication rule. These hypotheses have been strenuously and effectively criticised. But informing a person’s guilt probablility by comparison with the national (or whatever other abstract) population is a flawed approach that has little to do with population genetics axioms and their exceptions. Linacre’s statement that “such high probabilities lead the investigator to place the person at the crime scene” reiterates an old cliche—technically known as the fallacy of transposed conditional—here embodied in the desultory idea that every man on earth can pass by a Tuscan village on a certain day and commit a crime. Databases contain undetected errors—the larger the database, the more prone it is to errors. As it turns out, hidden errors are part of forensic evidence, in the sense that we can modify our beliefs according to the chance of their occurrence. We believe national DNA databases offer an opportunity to study errors and their real-scale effects. Why overstate the importance of these databases when the real population of criminal suspects is so small and the risk of misclassification with DNA from a national database so tangible. The forensic community needs to adopt a more circumspect attitude towards the use of DNA evidence from national databases in criminal cases and recognise the problems inherent in such an approach. *Vince L Pascali, Giampietro Lago, Marina Dobosz

Allele frequencies distribution of two VNTR markers (YNH24; YNZ22) in PST I digests from random Italian individuals (population of Rome)

Since the first report of a highly polymorphic locus in Man (Wyman and White, 1980), numerous hypervariable DNA sequences have been described. These markers have been given the name VNTR (Nakamura et al, 1987), and represent nowaday a major tool in forensic biology and paternity testings.

Classification of alphal antitrypsin phenotypes by high-resolution two-dimensional electrophoresis (2-D PAGE)

The ability of two-dimensional gel electrophoresis (2-D PAGE) to visualize soluble proteins is well documented (Anderson and Anderson 1977). By this tecnique, exploiting charge/mass differences to separate polypeptides, many polymorphic serum proteins were exactly located in 2-D PAGE maps of human serum (Anderson and Anderson 1979). The main advantages of this procedure are: first, a better molecular characterization of mutants is obtainable, secondly a simultaneous detection of several proteins is possible in the same slab gel;finally, the monitoring of non-conventional phenotypes,as the null variants (which would otherwise be difficult to detect) could be obtained (Asakawa et al. 1985).