Genetic studies in 5 Greek population samples using 12 highly polymorphic DNA loci

Abstract Two minisatellite (D1S80, D17S5) and 10 microsatellite (D2S1328, TPO, D3S1358, D9S926, D11S2010, THO1, VWF, FES, D16S310, and D18S848) polymorphic loci were analyzed in 5 Greek population groups (eastern Macedonia, central Macedonia, Thessaly, Epirus, and Greeks from Asia Minor) using the polymerase chain reaction. The genotypes at these loci conformed to Hardy-Weinberg equilibrium, and pairwise comparisons between them were in agreement with the expectation of independence between loci. This along with the low values of the coefficient of gene differentiation (G^sub ST^) and the high heterozygosity levels of all loci allows the use of allele frequency data from the 12 hypervariable DNA markers for medicolegal casework in the Greek population groups studied. The small genetic distances indicate a genetic affinity among the 5 population samples. However, a few markers seem to allow some discrimination among the groups. No significant differences with other European populations were found for the loci studied.

KEY WORDS: DNA POLYMORPHISM, HYPERVARIABLE REGION LOCI, MINISATELLITES, MICROSATELLITES, GREECE

To date, a large number of hypervariable region (HVR) loci have been described and characterized in the human genome. This polymorphism arises from variation in the number of tandem repeats of a DNA sequence that is generated by mutations that lead to new length alleles. Mutation processes include replication slippage, unequal sister chromatid exchange, and gene conversion (Schlotterer and Tautz 1992; Jeffreys et al.1994). Because of their multiallelic variation and, consequently, high level of informativeness, these HVR loci can be used for human gene mapping (Nakamura et al. 1987), genetic analysis of inherited diseases (Peake et al. 1990; Hearne et al. 1992), study of human evolutionary history (Bowcock et al. 1994), and personal identification in the medical and forensic sciences (Jeffreys et al. 1991; Gill et al. 1995).

The HVR loci include minisatellites and microsatellites. Minisatellite loci are composed of 6-100-bp repeats, and they are preferentially located in the proterminal regions of human autosomes (Royle et al. 1988). They present stable inheritance patterns and are characterized by a large number of alleles per locus and by high heterozygosities (Nakamura et al. 1987; Wong et al. 1987). Thus they are extremely useful in evolutionary studies of genetically close populations. Microsatellite loci consists of several repeats of 1-5 bp, and they are widely dispersed throughout the genome at an estimated frequency of 1 microsatellite per Cr-10 kbp (Oldroyd et al. 1995). Because of their small size (100-400 bp), they can be easily amplified from minimal amounts of even old and/or highly degraded material.

Several genetic studies using multiallelic systems have been conducted for worldwide human populations representing the major geographic groups (Balazs et al. 1992; Devlin and Risch 1992; Deka et al. 1995; Perez-Lezaun et al. 1997). Furthermore, during the last few years, a number of specific local populations have been investigated (Furedi et al. 1995; Hochmeister et al. 1995; Martin et al. 1995; Sjerps et al. 1995; Rose et al. 1996).

In Greece allele frequency data are available for only a few HVR loci (mostly minisatellites) and refer to either the whole population (Hatzaki et al. 1995; Lambropoulos et al. 1995) or a certain geographic area of Greece (De Benedictis et al. 1993; Falcone et al. 1995).

In our ongoing survey 12 highly polymorphic DNA loci were tested in Greek population samples residing in 4 geographic areas of Greece (eastern Macedonia, central Macedonia, Thessaly, Epirus) and 1 population of Greeks from Asia Minor living in eastern and central Macedonia since 1923 (Figure 1). These population groups were chosen to provide material that is relevant to the cultural, historical, and genetic questions that present themselves when Europe is considered within the wider global context.

Eastern Macedonia is a region north of the Aegean where Neolithic settlements appeared. In central Macedonia there are many excavation sites, such as Dion, Aiges, Pella, Edessa, and Sindos, and the oldest Neolithic settlement was found in Nicomedia, between the Aliakmonas and Axios rivers, which were important for the transmission of farming to Europe from the Axios valley. In Thessaly the fertile plains are of central importance for the first farmers of Europe, where the Neolithic (5000-4000 B.C.) settlement of Sesklo was found. In the mountain region of Epirus there are possible preNeolithic elements (Tsaktsitas 1995). Finally, the Greeks from Asia Minor were chosen because presumably the first farmers of Europe emanated from the Near East, more specifically, the Fertile Crescent, in the southeastern and south-central part of Asia Minor. From there they moved through Thrace and Macedonia (northern Greece) to Europe (Cavalli-Sforza et al. 1994).

For the analysis of the 5 groups we chose 2 minisatellites and 10 microsatellites. For the microsatellites only tetranucleotide repeat loci were used, because analysis of dinucleotide loci has revealed polymerase slippage during amplification, resulting in artifactual stutter or shadow bands (Hauge and Litt 1993) and making unambiguous designation of alleles difficult. In contrast, tetranucleotide loci are less prone to these truncated polymerase chain reaction (PCR) products.