G. Pappalardo

Genetic predisposition to familial neuroblastoma: Identification of two novel genomic regions at 2p and 12p

Objectives: The rarity of familial neuroblastoma (NB) has allowed only a few linkage studies, most of which did not show any evidence of linkage to regions involved in somatic alterations or to genes implicated in other neurocristopathies seldom associated with NB. We screened a highly informative family with recurrent NB by genome-wide linkage analysis aimed at identifying chromosomal regions for NB predisposing genes. Methods: A genome-wide screen was performed using 382 microsatellite markers. Multipoint model-based linkage analysis was carried out under a dominant mode of inheritance for the disease using the ‘affected only’ approach. Results: Our analysis identified two haplotypes co-segregating with the disease on chromosomes 2p and 12p, and yielded maximum lod-score values of 3.01 (p < 0.0001) for markers on both intervals. Conclusions: Evidence of linkage was reported at 16p in North American families, whereas our studies excluded this interval and indicated other loci for disease predisposition, thus confirming the remarkable genetic heterogeneity of NB. These results suggest an oligogenic inheritance in NB involving more loci in genetic determination of the disease.

Different informativeness of the three hypervariable mitochondrial DNA regions in the population of Bologna (Italy)

Mitochondrial DNA (mtDNA) sequence variations at hypervariable regions HVI, HVII and HVIII were analysed in 100 unrelated Italians from Bologna. The values of the statistical parameters are in agreement with the range of European populations. We suggest that the less informative HVIII region may be useful to distinguish HVI-HVII identical sequences in forensic analysis especially when nuclear DNA cannot be investigated.

Automated fluorescence analysis of CAG repeats at the human androgen receptor gene (HUMARA): evaluation of polymorphism in an Italian sample and report of a new allele.

The HUMARA CAG repeats polymorphism was studied in an Italian population sample. Polymerase chain reaction amplification and automated fluorescent analysis were used. A total of 19 and 15 repeats was observed in female and male subjects, respectively, and one new allele was found. The authors conclude that this X-linked short tandem repeat, typed without ambiguity and with a heterozygosity of 0.902, is useful in parentage testing of female subjects.

Population data of mitochondrial DNA region HVIII in 150 individuals from Bologna (Italy)

Abstract Mitochondrial DNA (mtDNA) sequence variations at the HVIII region in 150 unrelated Italian individuals from Bologna were analysed. A comparison of our sequence data with the Anderson sequence showed that nucleotide substitutions predominate over insertions and deletions. Among nucleotide substitutions the most frequent was a T–C transition.

The DRPLA CAG Repeats in an Italian Population Sample: Evaluation of the Polymorphism for Forensic Applications

The DRPLA CAG repeats polymorphism has been studied in an Italian population sample. PCR amplification, manual PAGE and silver staining were employed. A total of 16 different alleles, spanning the range from 5 to 21 CAG triplettes, was observed. The heterozygosity was 0.81 and no significant deviation from Hardy-Weinberg equilibrium was found 81 meioses from parentage testing were also analyzed and a Mendelian pattern of inheritance was observed in all cases. In addition, we could successfully type DRPLA locus in some forensic specimens, 1 ng of DNA allowing clear definition of alleles. The authors conclude that the DRPLA CAG repeats analysis may be useful for forensic applications.

APO B Polymorphism in the District of Bologna (Italy)

VNTRs are high polymorphic segments, called micro and minisatellites and due to the degree of allelic variability and the Mendelian inheritance they are useful markers in forensic application. Their polymorphism is attribuited to allelic variation in the number of tandemly repeated DNA sequences. One such hypervariable region is APOB 3′ HVR, located 75 bp downstream from the second polyadenilation signal at the 3′ end of human APOB, wich maps to human chromosome 2 (Knott et al 1986). The polymorphism is due to the repeat of two structurally related sequences “x” and “y” of 15 bp in lengh (Boerwinkle et al. 1989, Ludwig et al 1989).

Simultaneous DNA analysis of HLA-DPB and -DQB loci from single hairs: A forensic case report

Polymorphic DNA sequences can be amplified over a rnillionfo1d with polymerase chain reaction (PCR), therefore samples can be typed even when the DNA is degraded or very small amount is available. The HLA Class II genes HLA-DR, -DQ, and -DP present a high degree of polymorphism and are suitable as genetic markers for individual forensic identification (Korman et al 1985). Higuchi et al (1988) typed for HLA-DQA single hairs by using PCR. We report a case concerning five hairs found on the hands of a murdered man. HLA-DPB polymorphism was analyzed from all the single hairs, for two simultaneously with HLA-DQB polymorphism. The typing was compared to that obtained from plucked hairs of the victim and the suspect.

A method to help the detection of false homozygous samples at the D17S5 locus

Abstract Here we describe a method to check single-banded samples for the correct evaluation of this polymorphic region. Forward primers annealing within the D17S5 repeat sequences were designed and utilised in PCR with Horns reverse primer. The method allows false homozygous states to be revealed and may be suitable to avoid mistyping in population studies and in forensic casework.

Myotonic dystrophy CTG repeats in an Italian population sample: evaluation of the polymorphism for forensic applications.

The myotonic dystrophy (DM) CTG repeat polymorphism has been studied in an Italian population sample. Polymerase chain reaction (PCR) amplification, manual polyacrylamide gel electrophoresis (PAGE), and silver staining were employed. Alleles were typed by comparison with a sequenced allelic ladder. A total of 25 different alleles, spanning the range from 5 to 31 CTG triplets, was observed. The heterozygosity was 79%, and no significant deviation from Hardy-Weinberg equilibrium was found. Eighty-one meioses from parentage testing were also analyzed, and a Mendelian pattern of inheritance was observed in all cases. In addition, we could successfully type the DM locus in 20 laboratory-prepared bloodstains, with 1 ng of DNA allowing clear definition of alleles. We conclude that the CTG repeats at the DM locus may be useful for forensic applications.

Detection of a New Large Allele at the D17S5 (YNZ22) Locus

The variable number of tandem repeat regions (VNTRs) are a great source of polymorphism in the human genome. VNTR alleles have been identified initially by Southern blotting analysis . To date, as the sequences surrounding the polymorphic region have been defined, rapid typing of VNTRs polymorphism has been performed using PCR. PCR-based assay is rapid and can offer DNA typing also when the amount of DNA is too small to be typed with Southern blotting.