Interspecific Relationships among Charrs Based on Phylogenetic Analysis of Nuclear Growth Hormone Intron Sequences

DNA samples were obtained from six charr species: Salvelinus alpinus, S. malma, S. confluentus, S. leucomaenis, S. namaycush and S. fontinalis and two outgroups: Salmo salar and S. trutta. The C and D introns from the duplicate growth hormone loci (GH1 and GH2) were amplified using the polymerase chain reaction and sequenced from each of the species. Phylogenetic analyses with and without gaps were performed using the beta version of PAUP. The GH introns are evolving relatively slowly, so the data were combined with sequences from the ITS1 and ITS2 for a phylogenetic analysis of the charr species. The GH intron data and the combined data supported two groups of sister species among charrs: S. alpinus and S. malma, and S. confluentus and S. leucomaenis.     

Cytological characterization of the tandem repetitive sequences and their methylation status in the Antirrhinum majus genome

Tandem repetitive sequences are DNA motifs common in the genomes of eukaryotic species and are often embedded in heterochromatic regions. In most eukaryotes, ribosomal genes, as well as centromeres and telomeres or subtelomeres, are associated with abundant tandem arrays of repetitive sequences and typically represent the final barriers to completion of whole-genome sequencing. The nature of these repeats makes it difficult to estimate their actual sizes. In this study, combining the two cytological techniques DNA fiber-FISH and pachytene chromosome FISH allowed us to characterize the tandem repeats distributed genome wide in Antirrhinum majus and identify four types of tandem repeats, 45S rDNA, 5S rDNA, CentA1, and CentA2, representing the major tandem repetitive components, which were estimated to have a total length of 18.50 Mb and account for 3.59% of the A. majus genome. FISH examination revealed that all the tandem repeats correspond to heterochromatic knobs along the pachytene chromosomes. Moreover, the methylation status of the tandem repeats was investigated in both somatic cells and pollen mother cells from anther tissues using an antibody against 5-methylcytosine combined with sequential FISH analyses. Our results showed that these repeats were hypomethylated in anther tissues, especially in the pollen mother cells at pachytene stage.     

The utilization of a Pacific salmon Oncorhynchus nerka subsidy by three populations of charr Salvelinus spp.

The L_F‐at‐age trajectories differentiated two populations of Dolly Varden charr Salvelinus malma and a population of Arctic charr Salvelinus alpinus from the eastern end of Iliamna Lake, Alaska. Salvelinus malma from the Pedro Bay ponds were the smallest for a given age, followed by Salvelinus alpinus from the lake, and S. malma from the Iliamna River were much larger. The utilization of a large sockeye salmon Oncorhynchus nerka subsidy by the three Salvelinus spp. populations was then investigated by comparing diet data and mixing model (MixSIR) outputs based on carbon and nitrogen stable isotopes. Stomach contents indicated that both S. malma populations fed on O. nerka products, especially eggs and larval Diptera that had scavenged O. nerka carcasses, whereas S. alpinus fed on a variety of prey items such as three‐spined sticklebacks Gasterosteus aculeatus and snails. Stable‐isotope analysis corroborated the diet data; the two S. malma populations incorporated more O. nerka‐derived nutrients into their tissues than did S. alpinus from the lake, although all populations showed substantial utilization of O. nerka‐derived resources. Salvelinus alpinus also seemed to be much more omnivorous, as shown by stable‐isotope mixing models, than the S. malma populations. The dramatic differences in growth rate between the two S. malma populations, despite similar trophic patterns, indicate that other important genetic or environmental factors affect their life history, including proximate temperature controls and ultimate predation pressures.     

Comparative genetics at the gene and chromosome levels between rice (<Emphasis Type="Italic">Oryza sativa</Emphasis>) and wildrice (<Emphasis Type="Italic">Zizania palustris</Emphasis>)

Using comparative genetics, genes, repetitive DNA sequences and chromosomes were studied in the Oryzeae in order to more fully exploit the rice genome sequence data. Of particular focus was Zizania palustris L., n = 15, commonly known as American wildrice. Previous work has shown that rice chromosomes 1, 4 and 9 are duplicated in wildrice. The Adh1 and Adh2 genes were sequenced and, based on phylogenetic analyses, found to be duplicated in wildrice. The majority of the sequence diversity in the Adh sequences was in intron 3, in which were found several MITE insertions. Cytological and molecular approaches were used to analyze the evolution of rDNA and centromeric repetitive sequences in the Oryzeae. In wildrice, copies of the 5S rDNA monomer were found at two loci on two different chromosomes near the centromeres, as in rice. One nucleolar organizer region (NOR) locus was found adjacent to the telomere, as in rice. RCS1, a middle repetitive sequence in rice, was present in all of the centromeres of wildrice. RCS2/CentO, the highly repetitive component of Oryza sativa L. centromeres, was conserved in eight of the Oryza species examined, but was not found in wildrice. Three other middle repetitive centromeric sequences (RCH1, RCH2/CentO and RCH3) were also examined and found to have variable evolutionary patterns between species of Oryza and Zizania.Communicated by B. Friebe     

Genetic Differentiation of Three Sympatric Charr Species from the Genus SalvelinusInferred from PCR–RFLP Analysis of Mitochondrial DNA

We examined differentiation of three sympatric charr species Dolly Varden charr Salvelinus malmaWalbaum, white-spotted charrS.leucomaenisPallas, and Levanidov charrS. levanidovi Chereshnev, Scopetz, Gudkov. Using restriction fragment length polymorphism analysis (RFLP), three mitochondrial DNA fragments (ND1/ND2, ND5/ND6, and Cytb/D-loop) amplified in PCR were compared. The divergence of mtDNA nucleotide sequences betweenS. levanidovi and S. leucomaenis was about 9.7%, between S. levanidovi and S. malma, 8.7%, and between S. malma andS. laecomaenis, 7.5%. These results indicate genetic isolation of the species examined and support the earlier suggestion on closeness of Levanidov charr to the common ancestor of the genus Salvelinus.     

<Emphasis Type="Italic">Astyanax</Emphasis> <Emphasis Type="Italic">bockmanni</Emphasis> Vari and Castro, 2007: an ambiguous karyotype in the <Emphasis Type="Italic">Astyanax</Emphasis> genus

Despite the widespread distribution of Astyanax bockmanni in streams from Upper Paraná River system in central, southeastern, and southern Brazil, just recently, it has been identified as a distinct Astyanax species. Cytogenetic studies were performed in two populations of this species, revealing conservative features. A. bockmanni shows 2n = 50 chromosomes, a karyotypic formula composed of 10 M + 12SM + 12ST + 16A and multiple Ag-NORs. Eight positive signals in subtelocentric/acrocentric chromosomes were identified by fluorescent in situ hybridization (FISH) with 18S rDNA probes. After FISH with 5S rDNA probes, four sites were detected, comprising the interstitial region of a metacentric pair and the terminal region on long arms of another metracentric pair. Little amounts of constitutive heterochromatin were observed, mainly distributed at distal region in two chromosomal pairs. Additionally, heterochromatin was also located close to the centromeres in some chromosomes. No positive signals were detected in the chromosomes of A. bockmanni by FISH with the As-51 satellite DNA probe. The studied species combines a set of characteristics previously identified in two different Astyanax groups. The chromosomal evolution in the genus Astyanax is discussed.     

Comparative catches and food habits of dolly varden and arctic charrs,Salvelinus malma and S. alpinus,at Karluk,Alaska, in 1939–1941

Synopsis For many years the seriousness of the predations of dolly varden (Salvelinus malma) upon the eggs, alevins and juveniles of Pacific salmons (Oncorhynchus) has been a controversial subject. Over $ 300,000 was spent in Western Alaska from 1920 to 1941 for bounties on dolly varden in the belief that they were serious salmon predators. The writter undertook this study because of an intense desire to evaluate the need and justification for such a bounty system. The incidental examination of stomach contents of charrs which were captured for tagging or for tag recovery purposes from Karluk Lake and Karluk River on Kodiak Island, Alaska, and from Shelikof Strait and Uyak Bay during the summer months of 1939, 1940 and 1941 presented an excellent opportunity to pursue such a project. Stomachs were collected and contents analyzed from 1,992 arctic charr (Salvelinus alpinus) and 2,565 dolly varden charr (Salvelinus malma), taken by beach seine, gill net, fyke net, hook and line and by weir trap from Karluk Lake; from 956 dolly varden taken by seine and weir trap from the Karluk River; and from 462 dolly varden taken by beach seine and commercial salmon traps in Uyak Bay. Among the more than 5,000 charr stomachs examined, only 42 were found to contain salmon smolts, parr or alevin. Examination of 500 stomachs taken from dolly varden charr captured in the lower Karluk River at the height of the red salmon (Oncorhynchus nerka) smolt migration in May of 1939 revealed very little evidence of predation on the smolts in fresh water. There was some evidence that this relationship may have changed after the migrating charr had become adjusted to salt water, although the examination of 460 stomachs of dolly varden captured in salt water revealed nothing particularly incriminating in its role as a salmon predator. The fact that the downstream migration of the dolly varden reached its peak before that of the red salmon smolts was considered significant. Examination of the 4,500 charr stomachs at Karluk Lake in the months from April through September led to the discovery that two distinct species of charrs were present, one, the dolly varden, primarily stream-inhabiting and anadromous, and the other, the arctic charr, primarily lake-inhabiting and nonanadromous. Although red salmon eggs constituted 32% of the total volume of food ingested by both species of charr at Karluk Lake, there was considerable evidence that they were consumed in salvage or scavenger feeding, and there was practically no evidence that they were consumed as a result of predatory feeding. Only five, or one-tenth of one percent, of the thousands of charr stomachs examined at Karluk Lake contained red salmon parr or alevin.A summary of the contrasts in the feeding and migratory habits and physical characteristics of Salvelinus malma and Salvelinus alpinus led to the conclusion that alpinus was a more serious potential salmon predator than malma at Karluk Lake. In view of the fact that alpinus was seldom reached by the predator-control program whereas malma was being systematically destroyed only because it was more easily captured due to its migratory tendencies, it was recommended that the predator control program be terminated in Karluk waters.Published post mortem in shortened and slightly edited version. Reprint requests to the Editor.     

基于荧光原位杂交的藜属植物核型分析

为精确地识别藜属植物染色体组的核型特征,该文研究了4种来自青海高原的野生藜属植物(灰绿藜、藜、菊叶香藜及杂配藜)和1种从美国引进的栽培藜麦品种PI614932-HX(3)基于染色体荧光原位杂交(rDNA FISH)的核型。利用5S rDNA和45S rDNA对5种藜属植物有丝分裂中期的染色体进行FISH研究。藜属植物的核型分析结果表明:(1)藜属植物中存在二倍体(2n=2x=18)和四倍体(2n=4x=36)两种倍性,藜麦和灰绿藜为四倍体,其余3种为二倍体。(2)藜麦、灰绿藜、藜、菊叶香藜及杂配藜的核型公式分别为2n=4x=36=34m(2AST)+2sm,2n=4x=36=32m(4AST)+4sm,2n=2x=18=16m(4AST)+2sm,2n=2x=18=18m及2n=2x=18=16m+2sm。(3)染色体由大部分的中部着丝粒染色体(m)和少部分近中部着丝粒染色体(sm)组成。(4)核型类型除了菊叶香藜为1B以外,其余均属于2B类型。(5)在藜麦、灰绿藜及藜中具有分布位置不同、数量不等的双随体。5S rDNA、45S rDNA FISH结果表明:(1)藜麦和灰绿藜的染色体上存在2对5S rDNA位点和1对45S rDNA位点,藜、杂配藜的染色体上存在1对5S rDNA位点和1对45S rDNA位点,菊叶香藜的染色体上只存在1对5S rDNA位点。(2)5S rDNA和45S rDNA位点均位于染色体的短臂上。该研究首次获得了藜属植物基于5S rDNA和45S rDNA荧光原位杂交核型,为藜属植物亲缘关系研究和细胞生物学研究提供了分子细胞遗传学依据。     

Centromeric repetitive DNA sequences in the genus Brassica

Representatives of two major repetitive DNA sequence families from the diploid Brassica species B. campestris and B. oleracea were isolated, sequenced and localized to chromosomes by in situ hybridization. Both sequences were located near the centromeres of many chromosome pairs in both diploid species, but major sites of the two probes were all on different chromosome pairs. Such chromosome specificity is unusual for plant paracentromeric repetitive DNA. Reduction of stringency of hybridization gave centromeric hybridization sites on more chromosomes, indicating that there are divergent sequences present on other chromosomes. In tetraploid species derived from the diploids, the number of hybridization sites was different from the sum of the diploid ancestors, and some chromosomes had both sequences, indicating relatively rapid homogenization and copy number evolution since the origin of the tetraploid species.     

中国17种蔷薇属植物45S和5S的rDNA分布研究

为了探寻蔷薇属植物亲缘关系及系统发育研究的分子细胞遗传学证据,该研究采用双色FISH(荧光原位杂交)技术,对原产中国7个组的17种蔷薇属植物的45S和5S rDNA进行了定位分析。结果表明:(1)多数蔷薇属植物1组染色体对应1个45S rDNA位点和1个或2个5S rDNA位点,偶尔出现1~2个rDNA位点的丢失,但复伞房蔷薇(Rosa brunonii)的1组染色体对应了2个45S rDNA位点。(2)二倍体的蔷薇属植物至少有1对5S rDNA位点与45S rDNA位点共定位,而四倍体材料的5S rDNA位点与45S rDNA位点没有共定位,但所有四倍体材料均至少有1种rDNA信号纯合,表明它们应为二倍体直接加倍产生的同源四倍体。(3)绝大多数材料45S rDNA位于染色体短臂、5S rDNA位于染色体长臂,但缫丝花(R. roxburghii f. roxburghii)有1个5S rDNA信号位于染色体的短臂上,表明它与蔷薇属其他种的亲缘关系较远。(4)阿克苏地区和伊犁地区的疏花蔷薇的核型不同,且45S和5S rDNA的数量和位置不同,分子细胞遗传学证据也支持阿克苏地区的疏花蔷薇应为疏花蔷薇的新变种。(5)该研究中共有8个二倍体和6个四倍体蔷薇属植物的双色FISH为首次报道。研究认为,无论二倍体还是四倍体蔷薇属植物中出现的异形同源染色体、rDNA信号位置在同源染色体上的差异以及rDNA信号的增加和丢失,可能都与染色体结构变异和染色体重组有关,在分子细胞遗传学水平上证明染色体结构变异和染色体重组在蔷薇属植物演化过程中具有重要的作用。     

Comparative FISH analysis of C-positive regions of chromosomes of wood mice (Rodentia,Muridae, <Emphasis Type="Italic">Sylvaemus</Emphasis>)

The homology of DNA of C-positive centromeric regions of chromosomes in wood mice of the genus Sylvaemus (S. uralensis, S. fulvipectus, S. sylvaticus, S. flavicollis, and S. ponticus) was estimated for the first time. DNA probes were generated by microdissection from the centromeric regions of individual autosomes of each species, and their fluorescence in situ hybridization (FISH) with metaphase chromosomes of representatives of all studied wood mouse species was carried out. Unlike in the chromosomal forms and races of S. uralensis, changes in the DNA composition of the chromosomal centromeric regions in the wood mouse species of the genus Sylvaemus (including closely related S. flavicollis and S. ponticus) are both quantitative and qualitative. The patterns of FISH signals after in situ hybridization of the microdissection DNA probes with chromosomes of the species involved in the study demonstrate significant differences between C-positive regions of wood mouse chromosomes in the copy number and the level of homology of repetitive sequences as well as in the localization of homologous repetitive sequences. It was shown that C-positive regions of wood mouse chromosomes can contain both homologous and distinct sets of repetitive sequences. Regions enriched with homologous repeats were detected either directly in C-positive regions of individual chromosomes or only on the short arms of acrocentrics, or at the boundary of C-positive and C-negative regions.     

Development of Molecular Cytogenetics and Physical Mapping of Ribosomal RNA Genes in Lupinus

Identification of individual chromosomes in Lupinus is not possible due to gradient in size and similar morphology. To overcome this problem, molecular cytogenetics was developed for Lupinus. As an initial step in karyotype analysis, fluorescent in situ hybridization (FISH) was performed to determine genomic distribution of rRNA genes in L. hispanicus, L. luteus and L. × hispanicoluteus. It was found that all three diploid species posses two chromosome pairs carrying 18S-5.8S-25S rDNA and one chromosome pair carrying 5S rDNA. The use of probes for rDNA permitted unambiguous identification of three different pairs of chromosomes and revealed conservation of the number of rDNA loci among the three species. The study represents the first step in physical mapping of Lupinus genome through FISH by providing distinct chromosome landmarks. This revised version was published online in July 2006 with corrections to the Cover Date.     

Chromosome Analysis and Molecular Characterization of Highly Repeated DNAs in the Aphid Acyrthosiphon Pisum (Aphididae, Hemiptera)

Despite the interest in aphid biology, information on chromatin organization of their holocentric chromosomes is still limited to few species. In order to fill this gap, we have performed an extensive survey on pea aphid mitotic chromosomes using both classical and molecular cytogenetic techniques. Our results after silver, CMA₃ and DAPI-staining, C-banding and fluorescent in situ hybridization (FISH) using 28S rDNA and 5S rDNA as probes evidenced a tendency of repetitive DNAs to be concentrated on the X chromosomes. FISH experiments with the telomeric probe (TTAGG) _n revealed bright hybridization signals on each telomere of all Acyrthosiphon pisum chromosomes. No interstitial signals were seen. This revised version was published online in August 2006 with corrections to the Cover Date.     

A tandemly repetitive centromeric DNA sequence of the fish Hoplias malabaricus (Characiformes: Erythrinidae) is derived from 5S rDNA

A substantial fraction of the eukaryotic genome consists of repetitive DNA sequences that include satellites, minisatellites, microsatellites, and transposable elements. Although extensively studied for the past three decades, the molecular forces that generate, propagate and maintain repetitive DNAs in the genomes are still discussed. To further understand the dynamics and the mechanisms of evolution of repetitive DNAs in vertebrate genome, we searched for repetitive sequences in the genome of the fish species Hoplias malabaricus. A satellite sequence, named 5SHindIII-DNA, which has a conspicuous similarity with 5S rRNA genes and spacers was identified. FISH experiments showed that the 5S rRNA bona fide gene repeats were clustered in the interstitial position of two chromosome pairs of H. malabaricus, while the satellite 5SHindIII-DNA sequences were clustered in the centromeric position in nine chromosome pairs of the species. The presence of the 5SHindIII-DNA sequences in the centromeres of several chromosomes indicates that this satellite family probably escaped from the selective pressure that maintains the structure and organization of the 5S rDNA repeats and become disperse into the genome. Although it is not feasible to explain how this sequence has been maintained in the centromeric regions, it is possible to hypothesize that it may be involved in some structural or functional role of the centromere organization.     

A highly repetitive DNA component common to all Cervidae: its organization and chromosomal distribution during evolution

In recent work we have isolated and characterized a highly repetitive DNA (MMV satellite IA) from Muntiacus muntjak vaginalis, the species with the most reduced karyotype in the Cervidae family. We have now analysed the genomes of nine related species for the presence of MMV satellite IA components, and have determined their organization and chromosomal distribution. Repetitive satellite IA type DNA is present in all species of the Cervidae, and also in the bovine, but not in a species of the Tragulidae suggesting that these sequences were generated after the phylogenetic separation of Bovidae and Tragulidae. Studies on the organization of the satellite IA DNA in the various species revealed three main repeat lengths: 1400, 1000 and 807 bp. The relative proportion of satellite IA sequences present in any one of the three registers is strikingly different within the various species and can be correlated with the phylogeny of the Cervidae. The chromosomal locations of the satellite IA sequences were determined in seven species by in situ hybridization. It turned out that the chromosomal rearrangements leading to the reduction in the number of chromosomes during karyotype evolution have led to the elimination of satellite I DNA at most locations. In all tandem fusions, the satellite IA sequences located at the centromeres of the ancestral acrocentric chromosomes are lost. In contrast, during the centric fusion that generates the M. m. vaginalis X chromosome satellite IA sequences are amplified. Sequence motifs, which are known to be involved in recombinational events are present in the satellite IA and might have contributed to the unique karyotype variation in the Cervidae.     

Cytogenetic characterization of <Emphasis Type="Italic">Lippia alba</Emphasis> and <Emphasis Type="Italic">Lantana camara</Emphasis> (Verbenaceae) from Brazil

The aim of this work was to determine the cytogenetic characteristics of Brazilian Lippia alba (Mill) N. E. Brown and Lantana camara Plum. that could be useful for future characterization of these genera. Our analyses revealed that Li. alba has 2n=30 chromosomes consisting of ten metacentric and five submetacentric pairs, while La. camara has 44 metacentric chromosomes. The large blocks of heterochromatin seen in both species suggest an apomorphic condition. Six 45S rDNA sites were detected in both species by fluorescence in situ hybridization (FISH). Two and four 5S rDNA sites were observed in Li. alba and La. camara, respectively. Meiotic analysis revealed a normal chromosomal behaviour. The number of chromosomes and the presence of 45S rDNA and 5S rDNA sites do not exclude a possible polyploid origin. The cytogenetic differences between La. camara and Li. alba may be useful markers for differentiating these species.     

Multiplication of 28S rDNA and NOR activity in chromosome evolution among ants of the Myrmecia pilosula species complex

Chromosomal localization of rDNA in samples of five taxa of the Myrmecia pilosula species complex (Hymeoptera: Formicidae: Myrmeciinae) with 2n=3 (M. croslandi), 8 (M. imaii), 10 (M. banksi), 18 (M. haskinsorum), and 27 (M. pilosula) was carried out by fluorescence in situ hybridization (FISH) using cloned M. croslandi rDNA (pMc.r2) including the coding region for 28S rRNA. Results show that (1) the 28S rDNA in the genome of these ants is repetitive and is localized in pericentromeric C-bands, (2) the number of chromosomes carrying rDNA is two in M. croslandi, M. imaii and M. banksi, six in M. haskinsorum and ten in M. pilosula, and (3) only one or two clusters of rRNA genes generate nucleoli in each species. We suggest that the rDNA in the ancestral stock of the M. pilosula complex was localized originally in a pericentromeric C-band, and multiplied by chance with time during saltatory increases in C-banding following episodes of centric fission. Most rDNA multiplied on various chromosomes seems to be inactivated and eliminated from the genome, together with C-bands, by -inversion or centric fusion, with the remnant rDNAs dispersed in the genome by centric fission and -inversion.     

Improvement of high‐resolution fluorescence in situ hybridisation mapping on chromosomes of Brassica oleracea var. capitata

The low resolution of chromosome‐based Fluorescence in situ hybridisation (FISH) mapping is primarily due to the structure of the plant cell wall and cytoplasm and the compactness of regular chromosomes, which represent a significant obstacle to FISH. In order to improve spatial resolution and signal detection sensitivity, we provide a reproducible method to generate high‐quality extended chromosomes that are ~13 times as long as their pachytene counterparts. We demonstrate that proteinase K used in this procedure is crucial for stretching pachytene chromosomes of Brassica oleracea in the context of a modified Carnoy's II fixative (6:1:3, ethanol:chloroform:acetic acid). The quality of super‐stretched chromosomes was assessed in several FISH experiments. FISH signals from both repetitive 5S rDNA and single‐copy ARC1 on super‐stretched chromosomes are brighter than those on other different types of chromosome due to enhanced accessibility to targets on stretched pachytene chromosomes. In conclusion, the resulting extended chromosomes are suitable for FISH mapping for repetitive DNA sequences and the localisation of a single‐copy locus, and FISH performed on super‐stretched chromosomes can achieve significantly higher sensitivity and spatial resolution than other chromosome‐based FISH mapping techniques.     

Ribosomal RNA genes in soybean and common bean: chromosomal organization, expression, and evolution

Ribosomal RNA (5S and 45S) genes were investigated by FISH in two related legumes: soybean [Glycine max (L.) Merr.] and common bean (Phaseolis vulgaris L.). These species are both members of the same tribe (Phaseoleae), but common bean is diploid while soybean is a tetraploid which has undergone diploidization. In contrast to ploidy expectations, soybean had only one 5S and one 45S rDNA locus whereas common bean had more than two 5S rDNA loci and two 45S rDNA loci. Double hybridization experiments with differentially labelled probes indicated that the soybean 45S and 5S rDNA loci are located on different chromosomes and in their distal regions. Likewise, the common bean 45S and 5S rDNA loci were on unique chromosomes, though two of the 5S rDNA loci were on the same chromosome. FISH analysis of interphase nuclei revealed the spatial arrangement of rDNA loci and suggested expression patterns. In both species, we observed one or more 5S rDNA hybridization sites and two 45S rDNA hybridization sites associated with the nucleolar periphery. The 45S rDNA hybridization patterns frequently exhibited gene puffs as de-condensed chromatin strings within the nucleoli. The other condensed rDNA sites (both 5S and 45S) were spatially distant from the nucleolus in nucleoplasmic regions containing heterochromatin. The distribution of rDNA between the nucleoplasm and the nucleoli is consistent with differential gene expression between homologous alleles and among homoeologous loci.     

Comparative karyotype analysis in Haplopappus Cass. and Grindelia Willd. (Asteraceae) by double FISH with rRNA specific genes

The genera Grindelia Willd. and Haplopappus Cass. belong to the family Asteraceae - Astereae and are distributed in America and South America, respectively. Previous cytotaxonomic studies showed for South American species of Grindelia 2n=12 and for Haplopappus 2n=10 and 2n=12. Both Grindelia species (G. anethifolia, G. prunelloides), newly analyzed with molecular-cytological methods, exhibited symmetric karyotypes (AsI %=55.46 and 55.95) with metacentric chromosome sets (5m + 1m-sat) and 2n=12 chromosomes. The NOR was detected after fluorescence in situ hybridization (FISH) with 18/25S rDNA in the satellite chromosome 2. In contrast H. Happlopappus glutinosus, H. grindeloides and H. stolpii showed exclusively a higher asymmetric index (66.83%, 67.01% and 68.87%, respectively) with submetacentric chromosome sets (4sm + 1sm–sat). The sat-chromosomes 3 of H. glutinosus and H. grindelioides were both significantly different in their length from chromosomes 2 and 4. Furthermore in Grindelia the FISH with 5S rDNA could estimate signals in the short arms of chromosomes 3 or 4, that were not significantly differentiated in their length. Contrary to these findings in Grindelia, the position of 5S rDNA in Haplopappus was detected in the long arms of chromosome 1 (H. grindelioides and H. stolpii) and chromosome 2 (with two different loci) and chromosome 4 of H. glutinosus. The lengths of all measured chromosome arms with 5S rDNA were significantly different to those of the neighbours in the karyotypes. The two-color FISH of 5S and 18/25S rDNA had provided clear karyotypic markers for three (Haplopappus glutinosus) and two (H. grindelioides and H. stolpii) chromosomes. The number and position of rDNA signals were relatively highly conserved in the investigated five species without the double marked chromosome 2 of H. glutinosus, which shows an evolutionary dynamic of this 5S rRNA specific gene cluster. This investigation supports the assumption that the evolution of New World members of Grindelia and Haplopappus has not been accompanied by large karyotypic changes, but small chromosomal rearrangements have undoubtedly occurred (e.g. 5S rDNA localizations).