[Genomic Markers Associated with Cold-Hardiness in Brassica rapa L.]

Brassica rapa L. is a valuable and widespread species, but its cultivation in risk farming areas requires high-quality cold-hardy varieties to be developed. Mechanisms of the cold stress response in plants involve expression of numerous genes, including ribosomal ones, and are related to plant chromosome variability. FISH- and PCR-based methods were used to study intraspecific chromosome variability in the number and localization of 45S and 5S rDNA clusters and also to examine a set of molecular markers associated with cold-hardiness in winter B. rapa cultivars from high-risk farming areas. Several SSR (Na10-CO3 and BrgMS5339-1) and SCAR (BoCCA1-F/BoCCA1-1R1 and BoCCA1-F/BoCCA1-2R1) markers were identified as suitable for diagnosing cold-resistant and cold-susceptible genotypes in B. rapa. Compared with fodder cultivars, oilseed and leaf cultivars were shown to have more molecular markers associated with cold-hardiness and a higher level of polymorphism for the chromosomal distribution of 45S and 5S rDNAs, including chromosome heteromorphism. Thus, the least cold-resistant genotypes were found to display the lowest level of chromosome variability in the distribution of the 45S and 5S rDNA clusters and vice versa. The findings could be useful for the development of new cold-tolerant B. rapa varieties.

[Intraspecific divergence in wheats of the Emmer group using in situ hybridization with the Spelt1 family of tandem repeats].

Fluorescence in situ hybridization (FISH) was used to study the distribution of Spelt1 repetitive DNA sequences on chromosomes of 37 accessions representing eight polyploidy wheat species of the Emmer evolutionary lineage: Triticum dicoccoides Körn, T. dicoccum (Schrank) Schuebel, T. durum Desf., T. polonicum L., T. carthlicum Nevski, T. aethiopicum Jabbz., T. aestivum L., and T. spelta L. Substantial polymorphism in the number, distribution, and the sizes of the Spelt1 loci was revealed. On the chromosomes of the accessions examined, Spelt1 tandem repeats were found in seven different positions (per haploid chromosome set). These were "potential hybridization sites", including the subtelomeric regions of either short or long arms of chromosomes 2A and 6B, the short arm of chromosome 1B, and the long arms of chromosomes 2B and 3B. However, in individual genotypes, only from one to three Spelt1 loci were revealed. Furthermore, no hybridization with Spelt1 probe was detected on chromosomes from 12 accessions. Thus, the total number of Spelt1 sites in karyotypes varied from zero to three, with the average number of 1.16. This was substantially lower than in the species of the Timopheevi section and diploid Aegilops speltoides Tausch, a putative donor of the B genome. The decrease of the content of Spelt1 sequences in the genomes of the Emmer group wheats in comparison with the species of the Timopheevii group and diploid Ae. speltoides was assumed to result from the repetitive sequences reorganization during polyploidization and the repeat elimination during wheat evolution.

[Intraspecific divergence in wheats of the Timopheevi group as revealed by in situ hybridization with tandem repeats of the Spelt1 and Spelt52 families].

Fluorescent in situ hybridization (FISH) was used to study the distribution of the Spelt1 and Spelt52 repetitive DNA sequences on chromosomes of ten accessions representing three polyploid wheat species of the Timopheevi group: Triticum araraticum (7), T. timopheevii (2), and T. kiharae (1). Sequences of both families were found mostly in the subtelomeric chromosome regions of the G genome. The total number of Spelt1 sites varied from 8 to 14 in the karyotypes of the species under study; their number, location, and size differed among the seven T. araraticum accessions and were the same in the two T. timopheevii accessions and T. kiharae, an amphidiploid T. timopheevii-Aegilops tauschii hybrid. The Spelt52 tandem repeat was detected in the subtelomeric regions of chromosomes 1-4; its sites did not coincide with the Spelt1 sites. The chromosome distribution and signal intensity of the Spelt52 repeats varied in T. araraticum and were the same in T. timopheevii and T. kiharae. The chromosome distributions of the Spelt1 and Spelt52 repeats were compared for the polyploid wheats of the Timopheevi group and diploid Ae. speltoides, a putative donor of the G genome. The comparison revealed a decrease in hybridization level: both the number of sites per genome and the size of sites were lower. The decrease was assumed to result from repeat elimination during polyploidization and subsequent evolution of wheat and from the founder effect, since the origin of Timopheevi wheats might involve the genotype of Ae. speltoides, which is highly polymorphic for the distribution of Spelt1 and Spelt52 sequences and is similar in the chromosome location of the repeats to modern wheat.

[Chromosomal localization of 5S and 45S ribosomal DNA in species of Linum L. section Linum (syn=Protolinum and Adenolinum)]. / Khromosomnaia lokalizatsii 5S i 45S ribosomnoi DNK u vidov roda Linum L. sektsii Linum (syn.=Protolinum i Adenolinum).

Fluorescence in situ hybridization (FISH) was for the first time used to study the chromosomal location of the 45S (18-2.5S-26S) and 5S ribosomal genes in the genomes of five flax species of the section Linum (syn. Protolinum and Adenolinum). In L. usitatissimum L. (2n = 30), L. angustifolium Huds. (2n = 30), and L. bienne Mill. (2n = 30), a major hybridization site of 45S rDNA was observed in the pericentric region of a large metacentric chromosome. A polymorphic minor locus of 45S rDNA was found on one of the small chromosomes. Sites of 5S rDNA colocalized with those of 45S rDNA, but direct correlation between signal intensities from the 45S and 5S rDNA sites was observed only in some cases. Other 5S rDNA sites mapped to two chromosomes in these flax species. In L. grandiflorum Desf. (2n = 16) and L. austriacum L. (2n = 18), large regions of 45S and 5S rDNA were similarly located on a pair of homologous satellite-bearing chromosomes. An additional large polymorphic site of 45S and 5S rDNA was found in the proximal region of one arm of a small chromosome in the L. usitatissimum. L. angustifolium, and L. bienne karyotypes. The other arm of this chromosome contained a large 5S rDNA cluster. A similar location of the ribosomal genes in the pericentric region of the pair of satellite-bearing metacentrics confirmed the close relationships of the species examined. The difference in chromosomal location of the ribosomal genes between flax species with 2n = 30 and those with 2n = 16 or 18 testified to their assignment to different sections. The use of ribosomal genes as chromosome markers was assumed to be of importance for comparative genomic studies in cultivated flax, a valuable crop species of Russia, and in its wild relatives.

Molecular cytogenetic analysis of tetraploid and hexaploid Aegilops crassa.

The distribution of highly repetitive DNA sequences on chromosomes of tetraploid and hexaploid cytotypes of Aegilops crassa (Dcr1Xcr and Dcr1XcrDcr2 genomes) was studied using C-banding and in situ hybridization analyses with the pSc119, pAs1 and pTa794 DNA clones. In total, 14 tetraploid and five hexaploid accessions were examined. All chromosomes can be identified by their C-banding and ISH pattern with the pAs1 DNA clone. Only a few pSc119 hybridization sites were observed in the telomeric regions of several chromosomes. We found a high level of C-banding polymorphism and only minor variations in labeling patterns. The position of C-bands generally coincided with the location of the pAs1 sequence. Three 5S rDNA loci were detected in tetraploid Ae. crassa, whereas five pTa794 ISH sites were observed in 6x Ae. crassa. All the hexaploid accessions differed from the tetraploids by a reciprocal non-centromeric translocation involving chromosomes A and N. Three additional translocations were detected in the accessions analyzed. The Dcr1 genome of 4x Ae. crassa is highly modified compared with the D genome of the progenitor species Ae. tauschii. Because of the large amount of chromosomal rearrangements, the origin of the Xcr genome remains unknown. The second Dcr2 genome of 6x Ae. crassa is different from the Dcr1 genome but is similar to the D-genome chromosomes of Ae. tauschii, indicating that no additional large rearrangements occurred at the hexaploid level.