Milosz Smolik

Genetic variability of Polish and Russian accessions of cultivated blue honeysuckle (Lonicera caerulea).

Inter-simple sequence repeat (ISSR) amplification was used to analyze polymorphisms of micro-satellite sequences in the honeysuckle genome and to evaluate genetic diversity among fourteen Polish and Russian blue honeysuckle accessions (Lonicera caerulea var. edulis, L. caerulea no. 7661, L. caerulea no. 7987, Jolanta, Atut, Wojtek, Czarna, Zielona, Dlinnoplodna, Czelabinka, Signoglazka, N1, N2 and A). The plant material was selected from the Department of Pomology, the Dendrological Garden in Rogowo (Poland), and breeder collections. A total of 40 primers, containing different simple sequence repeat motifs, were tested for amplification. Out of the 40 primers, only 11 gave interpretable banding patterns in all blue honeysuckle accessions. A total of 129 ISSR loci were amplified, of which 83 (64%) were polymorphic and 24 (19%) accession-specific. ISSR-PCR with genomic DNA from blue honeysuckle yielded DNA fragments ranging from 260 to 3250 bp in size. UPGMA cluster analysis with bootstrapping (1000 replications) and used to construct a dendrogram and to estimate the genetic distances between Lonicera accessions. The ISSR-based phylogeny was consistent with Lonicera caerulea origin based on morphological and phenological evidence. The phylogenetic relationships based on the accession studies and the breeding usefulness are discussed.

Evaluation of genetic variability in choosen apple (Malus × domestica Borkh.) cultivars by ISSR-PCR analysis

The aim of the study was to determine the genetic variability in eight apple cultivars: Delikates, Cortland, James Grieve, Lired, Jonathan, Golden Delicious, Jonagold, and Idared from the collection of Fruit Growing Research Station in Rajkowo of the West Pomeranian University of Technology, Szczecin. The cultivar Delikates was obtained from the crossing of two cultivars: Cortland, and James Grieve, whereas cultivar Lired is a James Grieve’s sport. The second one cultivar—Jonagold was obtained from the crossing of Jonathan and Golden Delicious. The cultivar Idared is a hybrid obtained from the crossing of Jonathan and Wagener. Out of 40 primers, 17 were chosen for the final study. Those amplified a total of 183 loci (872 amplicons) out of which 34 (18.5%) were monomorphic, 128 (69.5%) were polymorphic and 22 (12%) cultivar-specific. Specific ISSR products were detected for each apple cultivar. A dendrogram was constructed using the UPGMA method which revealed two distinct clusters: I—Delikates, Cortland, James Grieve and Lired, II—Jonathan, Golden Delicious, Jonagold and Idared. Genetic similarity between Delikates, Cortland and James Grieve was 68.6, 70.8%, respectively and between cultivar Jonagold, Jonathan and Golden Delicous was 79.8, 85.2%, respectively.

R-ISSR - for fingerprinting, mapping and identification of new genomic loci in rye (secale cereale l.).

The results of the research confirming the possibility of applying various combinations of RAPD and ISSR primers in one multiplex PCR and the generation of a new type of R-ISSR products for the rye genome were presented in this work. The following was applied in the research: five rye genotypes including two inbred lines (153/79-1 and Ot1-3), hybrid F1 and two bulks (tolerant and susceptible) formed from recombinant inbred lines—RILs (F9) varying in the response to abiotic stress caused by nutrient deficiencies at the seedling stage. While evaluating the possibility of applying R-ISSR to the assessment of the rye variability, five of its genotypes were amplified separately with the RAPD and ISSR primers in each PCR reaction. These primers were combined in R-ISSR amplifications. The products of RAPD, ISSR and R-ISSR amplification were separated in 1.5% agarose gel. 32 R-ISSR combinations were examined, combining 20 and 8 selected RAPD and ISSR primers, respectively. 658 loci were amplified, including 230 RAPD, 180 ISSR and 271 R-ISSR, including 157 new loci. Over 91 loci were found, with an identical electrophoretic mobility for three methods. It was shown that R-ISSR products with electrophoretic mobility on agarose gels, identical to the co-migrating RAPD or ISSR, are not products of RAPD or ISSR, but they possess sequences of heteroamplicons—R-ISSR. The occurrence of sequences of primers used to R-ISSR was demonstrated while sequencing seven selected products of the above type. The ISSR primers with a low Tm were proven to generate repeatable fingerprints in the thermal profile of the reaction specific for RAPD and combined with the RAPD primer—repeatable R-ISSR profiles. A similar range of variability as described in RAPD or ISSR was observed in the R-ISSR profiles. The correlation coefficient between genetic similarity matrices for five rye genotypes, calculated with the Mantel test, amounted to rAB.C = 0.870.