[Genetic Differentiation among Natural Populations of the Lizard Complex Darevskia raddei as Inferred from Genome Microsatellite Marking].

The article presents the genetic parameters of the populations of lizards of the Darevskia raddei complex (D. raddei nairensis and D. raddei raddei) and the populations of D. valentini calculated on the basis of the analysis of variability of 50 allelic variants of the three nuclear genome microsatellite-containing loci of 83 individuals. It was demonstrated that the F(st) genetic distances between the populations of D. raddei nairensis and D. raddei raddei were not statistically significantly different from the F(st) genetic distances between the populations of different species, D. raddei and D. valentini. At the same time, these distances were statistically significantly higher than the F(st) distances between the populations belonging to one species within the genus Darevskia. These data suggest deep divergence between the populations of D. raddei raddei and D. raddei nairensis of the D. raddei complex and there arises the question on considering them as separate species.

[MOLECULAR GENETIC CHARACTERISTICS OF ALLELE VARIANTS OF MICROSATELLITE LOCI DU281, DU215, AND DU323 IN THE PARTHENOGENETIC LIZARDS DAREVSKIA ROSTOMBEKOVI (FAMILY LACERTIDAE) ].

One of the main questions-in the study of the unisexual (parthenogenetic) species of vertebrates is the determination of their genetic diversity. Nuclear and mitochondrial genetic markers can be used for this purpose. One of the most effective genetic markers is the microsatellite DNA, which mutates at a high rate. Development and characteristics of such markers are necessary in studies of parthenogenetic species. In this work, the analysis of the allele polymorphism of three microsatellite loci was performed for the first time via locus-specific PCR in the populations of parthenogenetic species Darevskia rostombekovi (n = 42) and bisexual parental species D. raddei (n = 6) and D. portschinskii (n = 6). All examined individuals of the parthenogenetic D. rostombekovi were heterozygous. Two to five alleles, depending on the locus, were found in the studied populations of the parthenogenetic species. It was shown that the differences were due to the varying structure of the microsatellite cluster and to single nucleotide substitutions at fixed distances in the DNA regions adjacent to the cluster. The allele structure variations form haplotype markers specific for each allele and inherited from the parental bisexual species. It was determined which alleles of the parthenogenetic species were inherited from the maternal species and which from the paternal species. Characteristics of distribution, frequency of occurrence, and combination of alleles of microsatellite loci, which determine the distinctive features of each D. rostombekovi population were obtained. The data can be used in the future to determine the clonal diversity and possible ways of its formation in the populations of the parthenogenetic species D. rostombekovi.

Identification of structural DNA variations in human cell cultures after long-term passage.

Random amplified polymorphic DNA (RAPD) analysis was adapted for genomic identification of cell cultures and evaluation of DNA stability in cells of different origin at different culture passages. DNA stability was observed in cultures after no more than 5 passages. Adipose-derived stromal cells demonstrated increased DNA instability. RAPD fragments from different cell lines after different number of passages were cloned and sequenced. The chromosomal localization of these fragments was identified and single-nucleotide variations in RAPD fragments isolated from cell lines after 8-12 passages were revealed. Some of them had permanent localization, while most variations demonstrated random distribution and can be considered as de novo mutations.

[Intra- and interspecific polymorphism of (AAT)n in microsatellite locus du47D in parthenogenetic species of the genus Darevskia].

The molecular structure of the allelic variants of (AAT)n of the Du47D microsatellite locus was determined in parthenogenetic lizards Darevskia dahli, D. armeniaca, and D. rostombekovi. Comparative analysis of these alleles showed that they were characterized by perfect structure of microsatellite cluster, and were different in the number of (AAT) monomeric units, as well as in the combinations of species-specific substitutions and deletions in the microsatellite flanking regions. Molecular structure of microsatellite cluster, species-specific single nucleotide polymorphism (SNP), and different representation of alleles Du47 in the samples of parthenogenetic species examined point to the origin of the alleles from different bisexual species, which is consistent with the hybrid nature of unisexual species of the genus Darevskia. In addition, these data reflect different combination patterns of interspecific hybridization events with the participation of the same bisexual species upon the formation of hybrid genomes of parthenogenetic species. Possible application of the allelic variants of microsatellite loci of parthenogenetic lizards as the genetic markers for the analysis of the genomes of parthenogenetic species in the light of evolution, ecology, and parthenogenetic type of reproduction in vertebrates is discussed.

[Molecular structure of allelic variants of microsatellite loci Du281 and Du47 in unisexual and bisexual lizard species of the genus Darevskia].

Experimental data on the molecular structure and variability of microsatellite loci in unisexual and bisexual lizard species of the genus Darevskia were analyzed. The allelic variants of Du281 and Du47 were found to differ in the number of monomers, the structure of microsatellite clusters, and point mutations in these clusters and flanking DNA. Interspecific comparison of alleles of these loci revealed both variable regions in the microsatellite clusters and allele-specific evolutionarily conserved nucleotide groups. In general, the results of comparative structural analysis of allelic variants testify to a high genetic similarity of the unisexual and bisexual lizard species studied and reveals the characteristic features of their interspecies variability.

[Molecular and genetic characterization of allelic variants of microsatellite loci in parthenogenetic Caucasian rock lizards Darevskia armeniaca (Lacertidae)].

Using a pair of primers selected for the loci deltau 215, deltau 281, and deltau 323 of Darevskia unisexualis monolocus PCR analysis of orthologous loci was carried out in populations of the related parthenogenetic species D. armeniaca and in populations of bisexual parental species D. valentini and D. mixta. It was shown that the studied loci were polymorphic and represented in populations of D. armeniaca by several allelic variants. We cloned and sequenced PCR amplification products of the allelic variants of deltau 215, deltau 281, and deltau 323 loci. It was found that allelic differences of microsatellite loci were caused by variation in a number of tandem repeats in the microsatellite clusters and point mutations in the flanking regions. Interspecies comparison of the orthologous locus deltau 215 between parthenogenetic species D. armeniaca and parental species showed that two allelic variants of deltau 215 in D. armeniaca were inherited from the parental bisexual species D. mixta and D. valentini. The third allelic variant was not found in parental species and appeared because of mutation processes in genome of parthenospecies. For the first time, the information about the molecular nature of allelic polymorphism of these microsatellite loci of parthenogenetic species D. armeniaca was received in this study.

[Nucleotide sequences of the microsatellite locus Du215 (arm) allelic variants in the parthenospecies Darevskia armeniaca (Lacertidae)].

Using monolocus PCR analysis with the pairs of primers designed for the Du215 locus of Darevskia unisexualis, allelic polymorphism at the orthologous locus in the populations of the related parthenospecies D. armeniaca was investigated. It was demonstrated that Du215 (arm) locus was polymorphic and in the populations of parthenospecies D. armeniaca (n = 127) represented by at least three allelic variants, differing from each other by the size and composition of microsatellite cluster, and by single nucleotide substitutions in flanking DNA. Unlike the Du215 locus, Du215 (arm) was shown contain not only GATA, but also (GACA) repeats, which were absent in D. unisexualis. Thus, in this study, the data on the molecular nature of allelic polymorphism at one of the microsatellite loci of the parthenospecies D. armeniaca were reported.