[Revealing of allelic polymorphism in the populations of parthenogenetic lizards Darevskia dahli (Lacertidae) using locus-specific PCR].

Locus-specific PCR was used to study the genetic polymorphism in three populations of parthenogenetic lizard species Darevskia dahli. The analysis was carried at the two (GATA)n-containing loci (Du215 and Du281) using the sample of 26 individuals. A total of eight Du215 and three Du281 allelic variants were detected. It was demonstrated that all the lizards examined were heterozygous at these loci. In 12 animals, unusual Du215 allelic variant was revealed, the origin of which was thought to be associated with different types of genomic rearrangements, or segmental duplication. The populations studied were substantially different relative to the levels of allelic polymorphism, which could be explained by different habitation conditions, leading to accumulation of mutations in noncoding genome regions.

[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.

[Finding of Bov-B LINE retroelement in parthenogenetic and bisexual lizard species of the genus Darevskia (Lacertidae)].

The Bov-B LINE retrotransposon was first discovered in Ruminantia and was long considered to be specific for this order. Later, this mobile element was described in snakes and some lizard species. Analysis of phylogenetic relationships of Bov-B LINE elements from different ruminants, snakes, and lizard species led to the suggestion on horizontal transfer of this retrotransposon from Squamata to Ruminantia. In the Squamata group, Bov-B LINE element was found in all snakes and some lizard species examined. The element was not detected in the genomes of some species of the genera Lacerta and Podarcis. In the present study, using PCR amplification and sequencing of PCR products, Bov-B LINE element was identified in the genomes of parthenogenetic and bisexual species of the genus Darevskia (Lacertidae), as well as in such species as Lacerta agilis and Zootoca vivipara, where this retrotransposon had not been not detected before.

[Analysis of genetic variation in unisexual and bisexual lizard species of the genus Leiolepis from Southeast Asia].

Using multilocus DNA fingerprinting with microsatellite probes (CAC)5, (GACA)4, (GGCA)4 and (GATA)4, intraspecific variation of the Southeast Asian lizards belonging to the genus Leiolepis (bisexual species Leiolepis reevesii and triploid parthenogenetic species Leiolepis guentherpetersi) was first examined. The L. guentherpetersi lizards were characterized by monophyletic DNA fingerprint profiles for the loci detected by the (GACA)4, (GGCA)4, and (CAC)5 probes, in terms of intrapopulation similarity index constituting S = 0.96. This was different from the individual-specific profiles of the lizards from bisexual, presumably parental species, L. reevesii (S = 0.6; P < 0.001). Genetic homogeneity of triploid L. guentherpetersi lizards at the loci examined serves as one of the arguments for the parthenogenetic nature of this species. Genetic variability of triploid parthenogenetic species L. guentherpetersi appeared to be comparable with that reported earlier for the Caucasian rock lizards of the genus Darevskia, namely, D. dahlia, D. armeniaca, and D. unisexualis (P > 0.05). The results of DNA fingerprinting analysis of the same L. guentherpetersi samples with the (GATA)4 hybridization probe were unexpected. Variability of parthenogenetic species L. guentherpetersi at the (GATA)n markers was remarkably higher than that at other DNA markers (S = 0.35; P = 3.08 x 10(-11)), being comparable to the variation of the (GATA)n DNA markers in bisexual species L. reevesii (P = 0.74). The reasons for high polymorphism of the (GATA)n-containing loci in L. guentherpetersi still remain unclear. This polymorhism is probably associated with high instability of the loci, which can be revealed by means of family analysis of parthenogenetic offspring.

[Molecular markers of nuclear DNA in the study of evolution and speciation process in an example of "Lacerta agilis complex" (Sauria: Lacertidae)].

In order to elucidate the molecular-genetic relations of some Lacerta s. str. lizard populations, subspecies and species in comparison with some other genera we used methods revealing four types of nuclear DNA markers. Among these were taxonprint, RAPD, Inter-MIR-PCR markers and also satellite DNA monomer sequences. The aim was to compare the phylogeny and systematics of this reptilian group based on morphological and molecular criteria. This problem has a general importance for understanding a speciation process. Our results show a good correlation between both approaches when genera and species levels were studied. Systematic status of five subspecies of L. agilis were supported but not in all cases, some subspecies have no meaningful genetic differences by three types of molecular markers, but all of them were differed by RAPD markers. The data confirm the subdivision of L. agilis populations into west and east clades proposed by other authors earlier on the basis of mitochondrial DNA and morphology. The population structure of one of the subspecies--L. agilis exigua, was studied on a number of populations distributed from Ural region up to Kabardino-Balkaria by IMP method. There were no significant differences among these 14 populations investigated. The data testify the rapid distribution of the species after the end of Pleistocene glaciation.

[Study of allelic polymorphism of (GATA)n-containing loci in parthenogenetic lizards Darevskia unisexualis (Lacertidae)]. / Izuchenie allel'nogo polimorfizma (GATA)n-soderzhashchikh lokusov partenogeneticheskikh iashcherits Darevskia unisexualis (sem. Lacertida).

The genesis of mini- and microsatellite loci, which is under extensive study in humans and some other bisexual species, have been virtually overlooked in species with clonal mode of reproduction. Earlier, using multilocus DNA fingerprinting, we have examined variability of some mini- and microsatellite DNA markers in parthenogenetic lizards from the genus Darevskia. In particular, mutant (GATA)n-restrictive DNA fragments were found in Darevskia unisexualis. In the present study, we examined intraspecific polymorphism of three cloned loci of D. unisexualis--Du323, Du215, and Du281--containing (GATA)7GAT(GATA)2, GAT(GATA)9, and (GATA)10TA(GATA) microsatellite clusters, respectively. Different levels of intrapopulation and interpopulation variability of these loci were found. Locus Du281 showed the highest polymorphism--six allelic variants (in the sample of 68 DNA specimens). Three alleles were found for locus Du215. The Du325 locus was electrophoretically invariant. The primers chosen for loci Du323, Du215, and Du281 were also used for PCR analysis of homologous loci in two presumptive parental bisexual species, D. valentini and D. nairensis. The PCR products of the corresponding loci of the parental species had approximately the same size (approximately 200 bp) as their counterparts in D. unisexualis, but the polymorphism levels of the paternal, maternal, and hybrid species were shown to be somewhat different. These data on the structure of the D. unisexualis loci provide a possibility to study genetic diversity in the parthenogenetic species D. unisexualis and other related unisexual and bisexual species of this genus, which can provide new information on the origin of parthenogenetic species and on the phylogenetic relationships in the genus Darevskia. These data can also be used for resolving problems of marking the lizard genome, which is still poorly studied.

[Superinstability (TCT/TCC)n of microsatellite DNA in parthenogenetic lizards Darevskia unisexualis (family Lacertidae)]. / Supernestabil'nost' (TCC/TCC)n mikrosatellitnykh DNK u partenogeneticheskikh iashcherits Darevskia unisexualis (sem. Laceridae).

Using multilocus DNA fingerprinting, we have examined variability of (TCT)n microsatellite and M13 minisatellite DNA repeats in populations, families, and tissues of Caucasian parthenogenetic rock lizards Darevskia unisexualis (Lacertidae). It has been shown for the first time that population and family DNA samples of D. unisexualis (75 samples in total) have individually specific DNA fingerprinting patterns of (TCT)n fragments. Analysis of inheritance of (TCT)n microsatellites in 46 first-generation progeny in 17 parthenogenetic D. unisexualis families revealed their extremely high instability. Mutant TCT fingerprint phenotypes were found in virtually each animal of the progeny. Moreover, varying fragments in the progeny and their original variants in the mothers were shown to simultaneously contain (TCT)n and (TCC)n polypyrimidine clusters. At the same time, no variability of (TCT)n fragments has been detected in the tissues and organs of mature parthenogenetic lizards and in the analogous tissues of the two-week-old progeny of this year. This suggests the absence of somatic mosaicism and methylation of the corresponding loci in the samples. Along with the hyperinstability of (TCT/TCC)n polypyrimidine clusters, we have shown that the population and family DNA fingerprinting patterns of M13 minisatellites were invariable and monomorphic in the same DNA samples of D. unisexualis. Our results indicate that mutations at loci containing polypyrimidine microsatellites significantly contribute to the total genomic variability of parthenogenetic lizards D. unisexualis.

[Variation of mini- and microsatellite DNA repeats in parthenogenetic lizard Darevskia armeniaca as revealed by DNA fingerprinting analysis]. / Fingerprintnyi analiz variabel'nosti mini- i mikrosatellitnykh povtorov DNK u partenogeneticheskikh iashcherits Darevskia armeniaca.

Population and family samples of two morphological forms (mutant and normal with respect to dorsal color) of pathogenetic lizard Darevskia armeniaca were examined by means of DNA fingerprinting using M13 mini- and (GATA)n and (TCC)n microsatellite DNA markers. The morphological forms examined were characterized by clonally inherited, species-specific patterns of the DNA markers, which were different from the species-specific DNA fingerprints of the other parthenogenetic species of the genus Darevskia (D. dahli. D. unisexualis, and D. rostombekovi). The mean index of similarity (S) obtained for a sample of 36 individuals from three isolated populations using three types of DNA markers was 0.966. This was similar to the variability level observed in D. dahli (0.962) (P > 0.05), but higher than that in D. unisexualis (0.950) (P < 0.05) and D. rostombekovi (0.875) (P < 0.01). Inheritance of M13 minisatellite and (TCC)n microsatellite DNA markers in the F1 offspring of parthenogenetic lizards was examined. It was shown that variability and clonal diversity of the fingerprint phenotypes observed in the populations and families of D. armeniaca could be at least partly explained by RFLP mutations in microsatellite repeats.

[Molecular evolution of satellite DNA CLsat in lizards of the Darevskia species (Sauria: Lacertidae): correlation with species diversity]. / Molekuliarnaia évoliutsiia satellitnoi DNK CLsat iashcherits roda Darevskia (Sauria: Lacertidae): korreliatsiia s vidovym raznoobraziem.

The structure and evolution of a satellite DNA family was examined in lizards from the genus Darevskia (family Lacertidae). Comparison of tandem units of repeated DNA (satDNA), CLsat, in all species from the genus Darevskia has shown that their variability is largely based on single-nucleotide substitutions, which constitute about 50 diagnostic positions underlying classification of the family into three subfamilies. Maximum differences between the subfamilies reached 25%. At this level of tandem unit divergence between the subfamilies, no cross-hybridization between them was observed (at 65 degrees C). The individual variability of one subfamily within the species was on average 5% while the variability between species consensuses within a subfamily was 10%. The presence of highly conserved regions in all monomers and some features of their organization show that satellites of all Darevskia species belong to one satDNA family. The organization of unit sequences of satellites CLsat and Agi also detected by us in another lizard genus, Lacerts s. str. was compared. Similarity that was found between these satellites suggests their relatedness and common origin. A possible pathway of evolution of these two satDNA families is proposed. The distribution and content of CLsat repeat subfamilies in all species of the genus was examined by Southern blotting hybridization. Seven species had mainly CLsatI (83 to 96%); three species, approximately equal amounts of CLsatI and CLsatIII (the admixture of CLsatII was 2-3%); and five species, a combination of all three subfamilies in highly varying proportions. Based on these results as well as on zoogeographic views on phylogeny and taxonomy of the Darevskia species, hypotheses on the evolution of molecular-genetic relationships within this genus are advanced.

[Satellite DNA restriction site variability as a molecular basis of taxonoprint method: evidence from the study of Caucasian rock lizards]. / Variabel'nost' saitov restriktsii satellitnoi DNK kak molekuliarnaia osnova metoda taksonoprinta (na primere skal'nykh iashcherits Kavkaza).

The restriction site distribution in satellite DNA of 17 Caucasian rock lizard species of the genus Lacerta (Darevskia gen. nov.), (Squamata, Lacertidae) was analyzed. The distribution patterns were shown to reflect the degree of satellite DNA evolutionary divergence, which could be revealed by taxonprint method, i.e., through the analysis of genomic DNA with a set of restriction endonucleases and subsequent computer-aided treatment. Thus, the taxonprint method offers an opportunity to examine the satellite DNA divergence in closely related species and infer their phylogeny of the species studied without reserting to costly and labor-consuming procedures. This is the advantage of using this technique at the early stages of genomic DNA phylogenetic analysis for rapid and effective estimation of relationships between closely related species as well as in the cases when DNA cloning and sequencing are too expensive or not feasible.

[Variation of mini- and microsatellite DNA markers in populations of parthenogenetic rock lizard Darevskia rostombekovi]. / Izmenchivost' mini- i mikrosatellitnykh markerov DNK v populiatsiiakh partenogeneticheskoi skal'noi iashcheritsy Darevskia rostombekovi.

Variation and clonal diversity in populations of the parthenogenetic rock lizard Darevskia rostombekovi was examined by means of multilocus DNA fingerprinting using mini- and microsatellite DNA markers M13, (GATA)4, and (TCC)50). The animals examined were shown to exhibit a clonally inherited, species-specific pattern of DNA markers (fingerprint profile) that is different from the species-specific patterns of parthenogenetic species D. dahli, D. armeniaca, and D. unisexualis. The mean intraspecific similarity index S was 0.950 (0.003) for a sample of 19 animals from three isolated populations of North Armenia. This significantly differed from the estimate of this parameter for a sample of 21 animals including two individuals from mountainous, relict population from the vicinity of the Sevan Lake, which was equal to 0.875 (0.001). A comparison of DNA fingerprints showed differences between 21 individuals attaining 79 DNA fragments of 1801 mini- and microsatellite markers included in the analysis. The results obtained show that intraspecific variation in D. rostombekovi is higher than that in the previously studied parthenogenetic species D. dahli (S = 0.962) and D. unisexualis (S = 0.950) (P < 0.001). Taking into account that D. rostombekovi is considered monoclonal on the basis of allozyme data, the problem of clonal variability is discussed with regard to the evidence on nuclear DNA markers. It is suggested that the hybrid karyotype of D. rostombekovi, which is more unstable than that of D. dahli and D. unisexualis, generates a series of chromosomal rearrangements (mutations). This may lead to the appearance of a geographically isolated chromosomal race (clone) in the population inhabiting the southeastern coast of the Sevan Lake.

[Reticulate evolution of parthenogenetic species of the Lacertidae rock lizards: inheritance of CLsat tandem repeats and anonymous RAPD markers]. / Setchataia évoliutsiia partenovidov skal'nykh iashcherits sem. Lacertidae: nasledovanie tandemnogo povtora CLsat i anonimnykh markerov metoda RAPD (RAPID).

The genetic relatedness of several bisexual and of four unisexual "Lacerta saxicola complex" lizards was studied, using monomer sequences of the complex-specific CLsat tandem repeats and anonymous RAPD markers. Genomes of parthenospecies were shown to include different satellite monomers. The structure of each such monomer is specific for a certain pair of bisexual species. This fact might be interpreted in favor of co-dominant inheritance of these markers in bisexual species hybridogenesis. This idea is supported by the results obtained with RAPD markers; i.e., unisexual species genomes include only the loci characteristic of certain bisexual species. At the same time, in neither case parthenospecies possess specific, autoapomorphic loci that were not present in this or that bisexual species.

[Increased variability of (TCC)n microsatelline loci in populations of the parthenogenetic lizard Lacerta unisexualis Darevsky]. / Povyshennaia variabel'nost' (TCC)n mikrosatellitnykh lokusov v populiatsiiakh partenogeneticheskogo vida iashcherits Lacerta unisexualis Darevsky.

In four isolated populations of parthenogenetic Caucasian rock lizard Lacerta unisexualis, variability of (TCC)n loci was examined using multilocus DNA fingerprinting. Unexpectedly high variability of (TCC)n microsatellites was found in all four populations. The mean similarity index was 0.825, which is higher than similarity estimates obtained for other mini- and microsatellite loci in L. unisexualis and parthenogenetic species L. dahli and L. armeniaca studied earlier. The high variation level of (TCC)n loci was shown to be at least partially associated with the presence of a diverged (TCC)n sequence fraction in the L. unisexualis genome. Mutations at some other genetically unstable (TCC)n loci may cause their structural diversity in populations of L. unisexualis.

[Variability of GATA-microsatellite DNA in populations of the parthenogenetic species of lizard Lacerta unisexualis Darevsky]. / Variabel'nost' GATA-mikrosatellitnykh DNK v populiatsiiakh partenogeneticheskogo vida iashcherits Lacerta unisexualis Darevsky.

Multilocus DNA fingerprinting was used to analyze the genome variation of mini- and microsatellite DNA regions in parthenogenetic Caucasian rock lizard Lacerta unisexualis. The DNA fingerprints obtained with probe M13 were nearly identical in all populations examined (the average similarity index S = 0.992). The fingerprints obtained with probe (GATA)4 varied (S = 0.862). Polymorphic fragments were assumed to correspond to allelic variants of genetically unstable GATA loci. Comparison of the fingerprints of animals from four geographically isolated populations revealed several population-specific GATA microsatellite markers. Based on their distribution among the populations, the corresponding alleles were assumed to originate from a common ancestral allele.

[Taxonoprint DNA analysis of certain lizard species from the Lacertidae family]. / Taksonoprintnyi analiz DNK nekotorykh vidov iashcherits semeistva Lacertidae.

It was earlier shown that electrophoretic patterns of the highly repetitive segments of DNA's of some animal species studied seem to have species specificity ("DNA taxonoprint method") [2]. In this paper the results of investigation of more than 20 lizard species of the Lacerta genus in comparison with representatives of some other genera and family are presented. It was shown that in taxonoprints there are one band, at least, but usually more bands common for all Lacerta species. Specimens of genus Podarcis have more similarity with genus Lacerta than genus Eremias. The representatives of Teiidae familie have no resemblance with family Lacertidae at all. Among Lacerta species there are 10, which form the separate group being very similar by most of the taxonoprints. This fact goes in line with hypothesis that 5 parthenogenetic species of this group evolve due to interspecies hybridization of another 5 bisexual species of this group. The other species studied have specific peculiarities in various taxonoprints permitting to draw some conclusions on their relations.