Mitochondrial DNA sequences of the hypervariable regions HVI and HVII were analysed in 83 Caucasians living in central Italy to expand the database for forensic identification purposes, and 75 different haplotypes resulting from 62 polymorphic positions in HVI and 44 in HVII were observed. The most frequent haplotype (263G, 309.1C, 315.1C) was shared by 7 individuals, 2 haplotypes were shared by 2 individuals, and 72 were unique. The genetic diversity was found to be 0.99 and the random match probability 1.9%. A condition of sequence heteroplasmy was found in only one case at nt 16311, whereas a length heteroplasmy was found in the homopolymeric stretch of cytosines 303-315. Our results indicate that in direct sequencing beyond the poly-cytosine stretch, the overlap is due to length heteroplasmy, whereas the blurred signal occurs when the stretch is composed of more than 10 cytosines.

Capillary electrophoresis with laser-induced fluorescence was applied to the analysis of six STRs and the amelogenin sex test with the purpose of verifying accuracy and precision of the sizing method with the GS500 internal standard. Sequenced dye-labeled, PCR-amplified alleles from amelogenin, HumVWA31, HumTH01, HumF13A01, HumFIBRA, D21S11 and HumCSF1PO loci were run several times on the same capillary and on multiple capillaries and the offset of computer-measured fragment sizes from the expected molecular weights was calculated and analysed. All loci except D21S11 showed a poor degree of accuracy. Precision results from run-to-run and day-to-day injections displayed a maximum standard deviation (SD) > 0.15 nt for HumVWA31, HumF13A01, D21S11 and HumFIBRA, although the maximum range of calculated sizes in multiple runs was lower than 1 basepair. No variation in precision was observed according to the quality of the DNA template. Allele typing by comparison with allelic ladders for each locus is recommended.

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A sample of 265 subjects from central Italy was analyzed at the HumCD4 locus by polymerase-chain-reaction (PCR). Phenotypes were identified by comparison with a sequenced ladder, after high-resolution horizontal polyacrylamide gel electrophoresis (PAGE) followed by silver staining. A set of representative alleles was sequenced by Taq-cycle-sequencing with dye terminator labeling and capillary gel electrophoresis strategies. Eight common alleles—5,6,7,8,9,10,11,12—and a rare larger 14, never before described in Caucasians, were found. Allele and genotype frequencies were similar to those described in former studies on Caucasians, with a prevalency of alleles number 5, 6, 10. Sequence analysis showed that the polymorphism is due to a pentameric TTTTC basic motif, tandemly repeated, and that from allele number 10 onwards the fourth repeat presents a T to C translation (CTTTC). Instead, allele number 9 may exist in two forms, because 75% of alleles examined in this study presented the CTTTC motif at the fourth position, while the remaining 25% had the basic repeat structure.

No abstract