Abstract: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 Jaccard's 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 Nei's gene diversity and the Shannon's 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.

To determine the suitability of micropropagation techniques developed for conserving rare medicinal herb Ungernia victoris we estimated the genetic fidelity of plants produced through direct regeneration from the bulb scale segments and organogenesis from long-term callus culture. Average value of the Jaccard's distances between explant-derived regenerants and maternal plants calculated from RAPD data was 0.5 %, while that of estimated between callus-derived regenerants and maternal cell line was 4.2 %; average distances between the objects among the explant-derived and callus-derived regenerants were 0.7 % and 2.5 %, respectively. The data obtained suggest that conditions for in vitro culture applied in this work provide relatively high genetic stability of the species upon the direct regeneration in vitro and regeneration from the long-term cultured callus.

An analysis of five 10-year-old U. victoris cell lines of common origin revealed mixoploidy, significant level of cell polyploidization, quantitative and qualitative changes in individual RAPD-fragments. No regularity was observed in the lines clustering depending on the nutrient medium phytohormone composition or genealogy of the lines. The cytogenetic, molecular and genetic approaches are recommended to ensure reliable determination of somaclonal changes, because the variations in ploidy levels do not always result in the RAPD-polymorphism.

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A comprehensive study of a typical steppe perennial

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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 (D L ) and resolving power (R p ) 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 D L is more efficient than R p ; therefore, we selected primers based on the D L value. In total, 12 primers with the largest values of D L 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 D L 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.

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