Katsuya Nagano

Contrasting patterns of the 5S and 45S rDNA evolutions in the Byblis liniflora complex (Byblidaceae)

To clarify the evolutionary dynamics of ribosomal RNA genes (rDNAs) in the Byblis liniflora complex (Byblidaceae), we investigated the 5S and 45S rDNA genes through (1) chromosomal physical mapping by fluorescence in situ hybridization (FISH) and (2) phylogenetic analyses using the nontranscribed spacer of 5S rDNA (5S-NTS) and the internal transcribed spacer of 45S rDNA (ITS). In addition, we performed phylogenetic analyses based on rbcL and trnK intron. The complex was divided into 2 clades: B. aquatica–B. filifolia and B. guehoi–B. liniflora–B. rorida. Although members of the complex had conservative symmetric karyotypes, they were clearly differentiated on chromosomal rDNA distribution patterns. The sequence data indicated that ITS was almost homogeneous in all taxa in which two or four 45S rDNA arrays were frequently found at distal regions of chromosomes in the somatic karyotype. ITS homogenization could have been prompted by relatively distal 45S rDNA positions. In contrast, 2–12 5S rDNA arrays were mapped onto proximal/interstitial regions of chromosomes, and some paralogous 5S-NTS were found in the genomes harboring 4 or more arrays. 5S-NTS sequence type-specific FISH analysis showed sequence heterogeneity within and between some 5S rDNA arrays. Interlocus homogenization may have been hampered by their proximal location on chromosomes. Chromosomal location may have affected the contrasting evolutionary dynamics of rDNAs in the B. liniflora complex.

Chromosomal study of lettuce and its allied species (Lactuca spp., Asteraceae) by means of karyotype analysis and fluorescence in situ hybridization.

In this study, in addition to the karyotype analysis, the chromosomal distributions of 5 S and 18 S rDNAs, and the Arabidopsis-type (T3AG3) telomeric sequences were detected by means of fluorescence in situ hybridization (FISH) to promote the information of chromosomal organization and evolution in the cultivated lettuce and its wild relatives, L. sativa, L. serriola, L. saligna and L. virosa. The karyotype analysis revealed the dissimilarity between L. virosa and the remaining species. In all four Lactuca species studied, one 5 S rDNA and two 18 S rDNA loci were detected. The simultaneous FISH of 5 S and 18 S rDNAs revealed that both rDNA loci of L. sativa, L. serriola and L. saligna were identical, however, that of L. virosa was different from the other species. These analyses indicate the closer relationships between L. sativa/L. serriola and L. saligna rather than L. virosa. Arabidopsis-type telomeric sequences were detected at both ends of their chromatids of all chromosomes not in the other regions. This observation suggests the lack of telomere-mediated chromosomal rearrangements among the Lactuca chromosomes.

A chromosome study of two centromere differentiating Drosera species, D. arcturi and D. regia

Abstract Using sequential fluorescent staining method and fluorescence in situ hybridization (FISH) technique, karyomorphological and molecular cytogenetic investigations of two centromere differentiating Drosera species, D. arcturi and D. regia were carried out. Drosera arcturi had chromosome number of 2n = 58, while D. regia had chromosome number of 2n = 34. Many chromosome bands stained with CMA positive and DAPI positive (CMA+DAPI+) were the most common in both species. CMA positive and DAPI negative (CMA+DAPI–) sites were shown in two chromosomes of both species. Four sites stained with CMA+DAPI–appeared on both sides of the constrictions of two larger chromosomes in D. arcturi, while two CMA+DAPI–sites appeared at terminal positions of two chromosomes in D. regia. Two-color FISH of 5S and 45S rDNAs showed two regions with major 45S rDNA signals in the both species, and four sites with clear 5S rDNA signals in D. arcturi. Drosera arcturi did not show any primary constriction in all chromosomes, except for two larger chromosomes. In contrast, D. regia had localized-centromeric position or well-differentiated primary constrictions in most metaphase chromosomes.

Use of RAPD analysis to assess the threat of interspecific hybridization to the critically endangered Polemonium kiushianum in Japan

Polemonium kiushianum is a critically endangered species of which only eight populations exist in semi-natural grasslands of the Mt. Aso area of Kyushu, Japan. Habitat modification and the risk of hybridization with non-indigenous horticultural congeners, such as P. caeruleum subsp. caeruleum and P. caeruleum subsp. yezoense var. yezoense, pose increasing threats to P. kiushianum. To develop a DNA marker that distinguishes P. kiushianum from other Polemonium species, we performed random amplified polymorphic DNA (RAPD) analysis and selected an approximately 500-bp fragment generated by the OPB06 RAPD primer. In addition, we designed a primer pair, H11F/R, based on the nucleotide sequences of the fragments derived from P. caeruleum subsp. caeruleum and P. caeruleum subsp. yezoense var. yezoense. The results with the H11F/R primers indicated that most extant P. kiushianum plants in natural populations are not genetically contaminated by hybridization with non-indigenous horticultural species.