Molecular organization of 5S rDNA intergenic spacer in Gentiana pneumonanthe L. and G. punctata L.

Aim. The study was aimed at cloning and analysis of molecular organization of 5S rDNA intergenic spacer (IGS) in two Gentiana species of Ukrainian flora, G. pneumonanthe L. and G. punctata L. Methods. 5S rDNA IGS sequence was amplified using polymerase chain reaction (PCR) with a pair of primers specific for the gene coding region. The produced PCR products were fractionated by gel-electrophoresis, isolated, ligated into plasmid pUC18, cloned into E. coli, and then sequenced. Nucleotide sequences were aligned using the Muscle algorithm and analyzed in the Unipro UGENE software. Results. The intergenic spacer region of the 5S rRNA genes was cloned and sequenced for two Gentiana species of Ukrainian flora, G. pneumonanthe and G. punctata. Based on the analysis of the alignment of the IGS sequences of five Gentiana species from three sections, some features of molecular organization of IGS of 5S rRNA genes in the studied species were established. In particular, motifs typical for other angiosperm families were identified, such as conservative oligo-dT motif at the IGS 3'-end that served as a transcription termination site and AT-rich region preceding the coding region of 5S rRNA gene. However, in the region of transcription initiation, conservative GC-element in position -13 is changed to AC. Conclusions. The interspecific variation of molecular organization of 5S rDNA IGS was identified among Gentiana species that can be used to clarify the phylogenetic relationships between members of this genus.Keywords: Gentiana species, 5S rDNA intergenic spacer, molecular organization, phylogeny.

Organization of the 5s rDNA intergenic spacer of Quercus rubra L. and its relationship to the ukrainian Quercus species

Aim. Quercus rubra L. is a representative of the North American section Lobatae, which has naturalized in Ukraine and becomes an invasive species in some places. Accordingly, elucidation of its relationship with the Ukrainian species of section Quercus is relevant to assess the possibility of interspecific hybridization. The 5S rDNA intergenic spacer (IGS) represents a convenient molecular marker to address this question. Methods. DNA isolation, PCR amplification, cloning and sequencing. Results. The IGS of Q. rubra was cloned and sequenced. It was shown that after the divergence of the Lobatae and Quercus sections, the IGS sequences evolved mainly by accumulation of nucleotide substitutions. A high level of IGS similarity was found for species within the Labatae and Quercus sections, while the genetic distances between the representatives of these two sections appeared significantly higher. Conclusions. The high level of nucleotide substitutions indicates a considerable genetic distance between the Lobatae and Quercus sections and calls into question the possibility of spontaneous hybridization between Q. rubra and Ukrainian native species of the section Quercus. At the same time, the close genetic relationship between other North American species, Q. lobata and Ukrainian "white oaks" indicate that introduction of representatives of this group may create conditions for their spontaneous crossing with the native species, which could be undesirable for the conservation of Ukraine's natural forest-steppe communities. Sequencing of the 5S rDNA IGS can be used for molecular genotyping of the Quercus species and for identification of interspecific hybrids. Keywords: 5S rDNA, molecular evolution, Quercus, Lobatae.