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A comprehensive study of a typical steppe perennial Iris pumila L. were carried out in the central zone of the European part of this species’ range, namely in Ukraine. Intraspecific differentiation, population size and isolation degree and its consequences, the threat of human impact were analyzed, as well as ecological amplitude and genetic variation in ISSR markers and selected chloroplast regions were determined. The species was found to have a low intraspecific differentiation that indicates the uniformity of the gene pool in the studied part of the range. Moreover, the results of isolation assessment, population and ecological study of I. pumila confirm the potential risk of extinction. A considerable part of the species populations exist as separated patches of rare ecosystems, isolated from the nearest neighbours due to intense plowing of the steppe zone. The generative reproduction is rare. In contrast, ISSR analysis revealed comparably high genetic diversity in all the sampled populations. Furthermore, specific plastid haplotypes were demonstrated in some of them. The inconsistency between the results of population ecological study and the data of molecular genetic analysis indicates that the loss of genetic diversity in the species caused by habitat fragmentation and isolation under increasing anthropogenic pressure is likely to be slower than it would appear judging from the assessment of population parameters, which clearly show negative trends. This result also emphasizes the necessity for integrated approach to assessment of the extinction risk for particular species and careful analysis of all determinants of extinction.

The aim of the study was to develop the system of polymerase chain reaction (PCR)-based markers for the assessment of genetic diversity and population genetic studies of Gentiana lutea L. as well as to determine the utility of two indices of marker informativeness. The informativeness was determined for 40 PCR primers of different types (random amplified polymorphic DNA, inter simple sequence repeat, inter-retrotransposon amplified polymorphism, resistance gene analog polymorphism and conserved DNA-derived polymorphism markers) by evaluating discriminating power (DL) and resolving power (Rp) in a sample of 30 plants from two populations. Analysis of correlation between the index value and the number of differentiated pairs of genotypes in the given sample revealed that DL is more efficient than Rp; therefore, we selected primers based on the DL value. In total, 12 primers with the largest values of DL were chosen. Analysis of genetic relationship among 86 plants from six populations showed that the number of bands produced by the three of selected primers was sufficient to give average bootstrap support across six key nodes in the dendrogram higher than 85%, while using six of the primers resulted in average bootstrap value exceeding 99%. Thus, a minimal set of three to six selected primers are sufficient for a quick assessment of genetic diversity of G. lutea populations, depending on the sample size and degree of differentiation between populations, while the rest of the primers with DL values above 0.8 may be used for ecogenetic surveys. Preliminary results obtained with selected primers indicate the moderate level of genetic variation within the species and significant differentiation among individual populations.

We investigated informativeness and effectiveness of different marker types (ISSR, IRAP, REMAP, RGAP and LP-PCR that employ primers based on the conservative sequences of abiotic stress response genes) to study genetic diversity of Iris pumila L. By the number of amplicons per primer, number of polymorphic amplicons per primer and resolving power index (Rp), ISSR-markers were the most efficient followed by LP-PCR-markers. In order of decreasing value of indicators of genetic diversity “the percentage of polymorphic bands”, and “the average Jaccardś genetic distance between plants”, marker systems may be arranged as follows: ISSR > RAPD > LP-PC > RGAP ≈ IRAP. For ISSR-markers, the percentage of polymorphic bands was 1.3–1.7 times higher than for the others, and the average genetic distance was 1.2–1.3 times higher. Different marker systems were ranked by the value of Neiś gene diversity and the Shannonś index as follows: ISSR > RAPD ≈ LP-PCR > RGAP ≈ IRAP, with the highest and the lowest values differing 1.4 times. Genetic population structure was investigated with program Structure 2.3. The data of all marker systems suggest that all genomes under study belonged to one population. The PCoA and cluster analyses based on genetic distances showed distinctions in clustering generated from different markers data and summarized data, as well as the lack of strong clusters. Mantel test revealed significant positive correlation between the matrices of genetic distances generated by the data of almost all marker systems. The strongest correlation was found between RGAP- and IRAP-markers (r = 0.452, p = 0.01) and between RGAP and ISSR (r = 0.430, p = 0.01). ISSR, RAPD and LP-PCR proved to be more effective for the study of I. pumila genetic diversity, nevertheless, joint use of different marker systems will provide a more comprehensive assessment of variation in different genomic regions.