Cytology and systematics in JapaneseArisaema (Araceae)

Chromosome numbers are determined from 37 populations attributed to 22 taxa of JapaneseArisaema. Of them, chromosome numbers ofA. limbatum var.conspicuum (2n=26),A. minus (2n=26),A. nambae (2n=28) andA. seppikoense (2n=26) are determined for the first time. New chromosome numbers, 2n=26, are reported forA. aequinoctiale, A. limbatum, A. stenophyllum, A. undulatifolium andA. yoshinagae. Three modes of basic chromosome numbers,x=14,x=13 andx=12, occur in JapaneseArisaema. Precise karyotypic comparisons of 20 taxa reveal that taxa withx=14 andx=13 share 26 major chromosome arms and have an obvious chromosomal relationship. One of two submeta-centric chromosomes inx=13 corresponds to two telo-centric chromosomes inx=14. InA. ternatipartitum with 2n=6x=72, ten out of 12 basic chromosomes are the most similar in size and arm ratio with larger ten chromosomes ofA. ringens among JapaneseArisaema examined. A basic chromosome number ofx=14 is the commonest in the genusArisaema and the remaining basic chromosome numbers,x=13 andx=12, seem to be derived through dysploidal reduction by translocating large segments of major arm of telo-centric chromosome onto other minor arm of telo-centric followed by loss of the remainings including a centromere, and by loss of two telo-centrics fromx=14, respectively. Some systematic problems of JapaneseArisaema are discussed based on new cytological data.Arisaema hatizyoense, A. minus andA. nambae are accepted as independent species.     

Molecular Phylogeny of Eupatorieae (Asteraceae) Estimated from cpDNA RFLP and its Implication for the Polyploid Origin Hypothesis of the Tribe

Neomirandea (x=17 and 25), Ageratina (x=17) and Sclerolepis (x=15) with the higher chromosome base numbers, and the other includes Mikania (x=17) and the remaining genera with lower chromosome base numbers (x=10–11). However, the monophyly of the former clade is supported with a low bootstrap value. In the latter clade, Mikania (x=17) diverged first, then Stevia (x=11), and finally eight genera with x=10 diverged in succession. This result supports the hypothesis that the genera in the tribe Eupatorieae with x =10 evolved from an ancestor with a higher base number, and the tribe is of polyploid origin. Received 13 September 1999/ Accepted in revised form 20 January 2000     

Sugarcane genome architecture decrypted with chromosome‐specific oligo probes

Sugarcane (Saccharum spp.) is probably the crop with the most complex genome. Modern cultivars (2n = 100–120) are highly polyploids and aneuploids derived from interspecific hybridization between Saccharum officinarum (2n = 80) and Saccharum spontaneum (2n = 40–128). Chromosome‐specific oligonucleotide probes were used in combination with genomic in situ hybridization to analyze the genome architecture of modern cultivars and representatives of their parental species. The results validated a basic chromosome number of x = 10 for S. officinarum. In S. spontaneum, rearrangements occurred from a basic chromosome of x = 10, probably in the Northern part of India, in two steps leading to x = 9 and then x = 8. Each step involved three chromosomes that were rearranged into two. Further polyploidization led to the wide geographical extension of clones with x = 8. We showed that the S. spontaneum contribution to modern cultivars originated from cytotypes with x = 8 and varied in proportion between cultivars (13–20%). Modern cultivars had mainly 12 copies for each of the first four basic chromosomes, and a more variable number for those basic chromosomes whose structure differs between the two parental species. One?four of these copies corresponded to entire S. spontaneum chromosomes or interspecific recombinant chromosomes. In addition, a few inter‐chromosome translocations were revealed. The new information and cytogenetic tools described in this study substantially improve our understanding of the extreme level of complexity of modern sugarcane cultivar genomes.     

Primary trisomics obtained from autotriploid by diploid reciprocal crosses in cucumber

To promote cytogenetical studies on cucumber (Cucumis sativus L., 2n = 2x = 14), the reciprocal crosses were made between autotriploid and diploid for selecting the primary trisomics. Meanwhile, chromosome behavior during meiosis in autotriploid cucumber was investigated to look for cytological evidences for origin of primary trisomics. Many viable F₁ seeds were obtained from reciprocal crosses between autotriploid and diploid. The number of chromosomes of 56 surviving progenies varied from 14 to 28, with plants having 2n = 15 occurring at the highest frequency (51.8%). Primary trisomics were firstly obtained in this study. Four types of primary trisomics were isolated and they could be distinguished from each other, as well as diploid. Variable chromosome configurations, e.g. univalent, bivalents and trivalents were observed in many pollen mother cells of the autotriploid at metaphase I. Binomial chromosome distribution was observed at anaphase I and frequency of 8/13 was 6.25%. The meiosis of autotriploid, especially the class of gametes with eight chromosomes, gave the cytological evidence of producing 2x + 1 type gamete and could be induced into primary trisomic plants from progeny of autotriploid–diploid crosses. These studies have established a ground work for selecting a series of primary trisomics, and further using them for associating linkage groups with specific chromosomes in cucumber.     

Inter- and intra-genomal chromosome pairing in an interspecific hybrid and its bearing on basic chromosome number inPennisetum

Meiosis in the interspecific hybrid betweenPennisetum typhoides (2n=14; genomeAA) andP. purpureum (2n=28; genomesA′A′BB) has been studied with particular reference to allosyndetic and autosyndetic pairing of chromosomes. Although up to nine bivalents occurred in the hybrid, never more than five were observed to be (heteromorphic)AA′ bivalents (range 1–5). It has been concluded thatA andA′ genomes are onlypartially homologous. It has further been inferred that the two genomes are evolutionarily related and could have arisen from a common progenitor withx=5 chromosomes or from related species withx=5 chromosomes. Autosyndetic pairing of chromosomes within thetyphoides complement (A genome) and within theA′ genome ofpurpureum have been reported here for the first time. Intra-haploid pairing to a probable maximum of two bivalents within each of the three genomes of the hybrid, viz.,A, A′ andB, further suggestsx=5 as the phyletically basic number in the genusPennisetum. It has been inferred thatx=7 is a secondarily basic number, having been derived fromx=5. The occurrence of a species withn=5 inPennisetum, viz.,P. ramosum, substantiates this view. Further support in favour of this conclusion comes from the secondary association of bivalents in dipoidP. typhoides. Thus, the apparently diploid species,Pennisetum typhoides with2n=14 chromosomes is considered to be a “secondary diploid” having a secondarily balanced number ofx=7. On the basis of the results obtained by the author is conjunction with the available evidence from the literature, it is suggested thatx=5 may be the original basic number for the entire grass family and seven, the most preponderant number in it, and other higher numbers derived from it subsequently during the course of evolution.     

Karyomorphology of some Moraceae and Cecropiaceae (Urticales)

The karyomorphology of 16 species in 13 genera representing Moraceae and Cecropiaceae was investigated in an effort to contribute to a better understanding of chromosome features and evolution in the families. All genera investigated have similar karyomorphology, but differences are found in (1) chromosome features of Interphase nucleus (simple, simple-complex, or complex chromocenter type), (2) basic chromosome number (x=13 or 14), (3) size variation (mono-or bimodial), and (4) frequencies of chromosomes with median centromeres (m-chromosome) (25–85%) and those with subterminal (or terminal) centromeres (st-chromosome) (14–69%). Comparisons with Ulmaceae as an outgroup of the remainder of Urticales suggest that the simple chromocenter type,x=14 comprising bothm- andst-chromosomes, and the monomodial karyotype are plesiomorphies in Moraceae and Cecropiaceae. Most of Moraceae and Cecropiaceae retain generalized chromosome features of the order, but have involved a few evolutionary changes in karyomorphology. Based on some detailed karyomorphological data, inter- and infrafamilial relationships are also briefly discussed.     

Karyomorphology and evolution in some Hamamelidaceae and Platanaceae (Hamamelididae; Hamamelidales)

Karyomorphology in 14 species of 12 genera representing a variation of Hamamelidaceae and in one species of Platanaceae (Platanus only) is investigated in an effort to contribute to an understanding of chromosome evolution and inter- and intrafamilial relationships. All genera investigated show similar chromosome features at resting stage and prophase, excepting that at resting stageRhodoleia shows the simple, rather than the simple-complex, chromocenter type as in other genera. At metaphase all the genera investigated of Hamamelidaceae, like other ‘lower’ Hamamelididae, have chromosomes with median centromeres (m-chromosomes), those with submedian centromeres (sm-chromosomes) and those with subterminal (or terminal) centromeres (st-t-chromosomes) at different frequencies, although frequencies ofst-t-chromosomes are always less than 33%. InPlatanus,m-chromosomes are lacking and insteadst-t-chromosomes are predominant (86%), a feature seemingly very specialized. We confirmedx=7 in Platanaceae,x=12 in Hamamelidoideae and Rhodoleioideae, andx=8 in Exbucklandioideae and Altingioideae (Hamamelidaceae). An analysis of chromosome morphology supports the hypothesis thatx=12 in the former two subfamilies is of tetraploid origin fromx=6, rather than of triploid origin fromx=8. We further give brief comments on the suprageneric classification of Hamamelidaceae that was recently proposed by Endress.     

In vitro induction of a trisomic of Agave tequilana Weber var. Azul (Agavaceae) by para-fluorophenylalanine treatment

The effect of para-fluorophenylalanine (PFP) on the production of trisomic plants of Agave tequilana Weber var. Azul produced through somatic embryogenesis was investigated. Normal diploid plants with 2n = 2x = 60 were obtained in the control treatment and with 4 mg L⁻¹ PFP exposure, while use of 8 and 12 mg L⁻¹ PFP led to production of trisomics with 2n = 2x = 61. Normal diploid plants showed a bimodal karyotype with five pairs of large chromosomes and 25 pairs of small chromosomes. Trisomic plants also had a bimodal karyotype with a group of three chromosomes in position five of the chromosome set. More than 13 homologous chromosome pairs exhibited structural changes. Differences in chromosome arm ratio (long arm/short arm) were also found in eight chromosome pairs; all these aberrations in the chromosome complement of trisomic plants were probably caused by inversions, deletions, and/or duplications produced by high concentrations of PFP. The gross chromosome structural changes and the presence of a single extra chromosome could have been induced by the effect of PFP on the mitotic spindle by inducing nondisjunction of sister chromatids, resulting in hyperploids (2n + x) and hypoploids (2n − x). Flow cytometric analysis of nuclear DNA content was performed using nuclei isolated from young leaves of normal and trisomic plants. The 2C DNA content of 8.635 pg (1Cx = 4,223 Mbp of trisomic plants was different (p < 0.001) than that of normal plants (2C DNA = 8.389 pg (1Cx = 4,102 Mbp). The difference in genome size was correlated with the large structural changes in the trisomic plant genomes.     

Early evolution of the chromosomal structure of Triticum turgidum–Aegilops ovata amphiploids carrying and lacking the Ph1 gene

After two selfing generations of two different Triticum turgidum –Aegilops ovata amphiploids carrying the Ph1 gene, or lacking it (ph1c mutant), karyotypes of their offspring were scored by GISH (genomic in situ hybridization). On average, the chromosome number was lower than expected (56 chromosomes) on the basis of the parental constitutions (T. turgidum, AABB, 2n=4x=28; Ae. ovata, M^oM^oU^oU^o, 2n=4x=28). The lost chromosomes belonged to the wild Aegilops species. The two families differed greatly by their number of intergenomic translocations, also detected by GISH. The ph1c family showed nine translocations over 12 plants while only one translocation was observed in the Ph1 family. All exchanges involved either the M^o and U^o chromosomes or the M^o and wheat chromosomes, the size of the exchanged segment ranging from 3% to 36% of the total chromosome length. The results suggest an epistatic effect of the ph1c deletion over the genetic diploidizing system that operates in Ae. ovata since translocated chromosomes are most-likely derived from homoeologous recombination. The potential of these results for wheat breeding programmes is also considered. Received: 28 November 2000 / Accepted: 20 March 2001     

Revision of ploidy status of Dioscorea alata L. (Dioscoreaceae) by cytogenetic and microsatellite segregation analysis

Dioscorea alata is a polyploid species with several ploidy levels and its basic chromosome number has been considered by most authors to be x = 10. Standard chromosome counting and flow cytometry analysis were used to determine the chromosome number of 110 D. alata accessions of the CIRAD germplasm collection. The results revealed that 76% of accessions have 2n = 40 chromosomes, 7% have 2n = 60 chromosomes and 17% have 2n = 80 chromosomes. Progenies were produced from 2n = 40 types of D. alata and the segregation patterns of six microsatellite markers in four different progenies were analysed. The Bayesian method was used to test for diploid versus tetraploid (allo- and autotetraploid) modes of inheritance. The results provided the genetic evidence to establish the diploidy of plants with 2n = 40 chromosomes and to support the hypothesis that plants with 2n = 40, 60 and 80 chromosomes are diploids, triploids and tetraploids, respectively, and that the basic chromosome number of D. alata is x = 20. The findings obtained in the present study are significant for effective breeding programs, genetic diversity analysis and elucidation of the phylogeny and the species origin of D. alata.     

Contributions to the cytotaxonomy of the Commelinaceae*

Further material of Gibasis geniculata (Jacq.) Rohw. (syn. Tradescantia geniculata Jacq.) and other Gibasis species collected in the field has been studied. The existence of several species in our earlier experimental material is confirmed. These include G. geniculata itself (2n= 32 or 48 small chromosomes), G. oaxacana D. R. Hunt (2n= 16 small chromosomes) and G. schiedeana (Kunth) D. R. Hunt, which has two chromosome forms, 2n= 10 and 2n= 16, both with large chromosomes. These forms are diploid and tetraploid based on x= 5 and x= 4 respectively and show a Robertsonian relationship with each other. The cytology of tetraploid (2n= 20) G. karwinskyana is confirmed and that of a diploid form (2n=10) described. The recently described G. consobrina D. R. Hunt (2n= 20) is shown to be cytologically comparable with G. karwinskyana, but to differ in significant details. Next, the cytology of G. pulchella (Kunth) Rafin., the type species of Gibasis, is described and also that of the allied G. matudae D. R. Hunt. Both G. pulchella (2n= 10, 15) and G. matudae (2n= 10) show interchange heterozygosity, with complete rings of ten chromosomes at meiosis in some plants of G. pulchella. Preliminary comments are made on the cytology of G. aguensis (Standl. & Steyerm.) Rohw. (2n= 10), another ally of G. pulchella, and on the G. linearis group, where the basic number appears to be x= 6 but two karyotype patterns have been found. A discussion of chromosome architecture in Gibasis in relation to the taxonomy of the genus concludes the paper.     

Reticulate evolution of the hybrid produced artificially by crosses between Osmunda banksiifolia and Osmunda lancea

Although ferns have been developed by hybridization and chromosome doubling, no natural polyploidy has yet been recorded in Osmundaceae. So, we produced hybrids artificially by crosses between Osmunda banksiifolia (2n = 2x = 44) and Osmunda lancea (2n = 2x = 44), and investigated their sporogenesis. From the O. banksiifolia × O. lancea hybrid with 44 univalent chromosomes, allotetraploids with 44 bivalent chromosomes were produced by chromosome doubling, and allotriploids with 22 univalent chromosomes and 22 bivalent chromosomes were then produced by back crosses. The results show when and how chromosome doubling occurs in hybrids. The success of artificial hybridization between O. banksiifolia and O. lancea, did not, however, reflect any product of natural hybridization between the two species.     

Decreased DNA content of birch (Betula) chromosomes at high ploidy as determined by cytophotometry

Relative amounts of DNA were determined on telophase nuclei by Feulgen cytophotometry for euploid taxa of birch (Betula) with somatic chromosome numbers of 28, 42, 56, 70, and 84. A direct correlation was found between observed DNA absorbance and chromosome number except for plants of B. papyrifera with 84 somatic chromosomes. The DNA density value for nuclei of the 84-chromosome plants fitted a 12.25 ratio instead of the expected 13.0 ratio. The DNA density value for these plants was calculated to be approximately equivalent to plants which would possess 63 somatic chromosomes. The average DNA value per chromosome was 2.73 for the 84-chromosome plants in contrast to 3.50 per chromosome in each of the lower euploids. Nuclear diameters of the 84-chromosome plants were directly related to chromosome number and not to DNA density value. The genomic number of Betula was considered to be x=7, rather than x=14, since a DNA value equivalent to 63 chromosomes is a multiple of 7 and not 14. Diploid birch species (2n=2x=28), therefore, would actually be tetraploids (2n=4x=28). The reduction in DNA content may be an adaptation for the establishment of higher ploidy in birches.     

Cytogenetics of tropical bulbous ornamentals VIII: Pollen grain mitosis in Zephyranthes puertoricensis

The immediate result of recombination during male meiosis within the complex interchange heterozygote polyploids in Zephyranthes could be assessed from the chromosomal constitution of pollen grains in one species with 2n=25. The numbers found in the pollen grains were n=1–16, 20 and 24. Considering n=12 (22.3%) as the haploid number, nearly 57.2% were subhaploid, out of which 19.6% had n=11. Occurrence of such a high proportion of subhaploid grains may be explained on the basis of the polyploid constitution (4x+1) of the species (x=6). In this sense, grains with a basic set of 6 chromosomes may be able to function. Therefore, in the real sense only grains with numbers less than 6 are submonoploid which constitute only 10.7%. In the subhaploid grains there was a preference for large and medium sized chromosomes, while in grains with higher numbers there was noted a tendency for smaller chromosomes to be represented more than once, indicating their non-disjunction during meiosis. Totally new types of nucleolar chromosomes emanating after recombination were also seen.This investigation has shown the extent of potential cytological polymorphism possible in heterozygotes like Zephyranthes puertoricensis. How much of this polymorphism is meaningful in creating the aneuploid pattern found in the genus, depends on the competitive advantage different numbers have during fertilisation vis-a-vis possibly similar polymorphism created during the female meiosis.     

Physical mapping of ribosomal DNA on several species of the subgenus Rosa

The localization of NORs by fluorescent in situ hybridization on metaphase spreads of five diploid (Rosa gigantea Coll., Rosa moschata Herrm., Rosa multiflora Thunb., Rosa rugosa Thunb. and Rosa sempervirens L., 2n=2x=14), one triploid (Rosa chinensis’semperflorens’ Koehne., 2n=3x=21) and one tetraploid (Rosa gallica ’versicolor’ L., 2n=4x=28) ancestral species of modern roses was studied. Two terminal hybridization signals were observed in all diploid species corresponding to a single NOR per genome in these species. Triploid R. chinensis showed three hybridization sites on the short arm of three morphologically similar chromosomes. Six hybridization sites, located at terminal positions of the short arms of three chromosome pairs, were observed in the tetraploid species. These signals corresponded to three pairs of NORs and all of them were located in chromosome pairs of different size. These observations, together with the analysis of meiotic pairing in PMCs, support the view that our specimen of R. chinensis is an autotriploid and that R. gallica’versicolor’ has an alloploidy nature. Received: 27 November 2000 / Accepted: 12 March 2001     

Karyomorphology ofCrossostylis (Rhizophoraceae)

We present the first report on somatic chromosome numbers and morphology in eight of 13 recorded species ofCrossostylis, one of inland genera of Rhizophoraceae. The chromosome number ofCrossostylis is 2n=28 in all species examined; therefore, the genus hasx=14, a number which is the smallest and unknown elsewhere in the family. Based onCrossostylis raiateensis, we further present that 24 of 28 chromosomes at metaphase have centromeres at median position, and the remaining four at submedian or subterminal position. The chromosome morphology seems to imply thatCrossostylis might be a tetraploid with the original base numberx=7, but an extensive study in the other inland genera is needed to find such a small chromosome number.     

Karyological studies of some representatives of <Emphasis Type="Italic">Tanacetum</Emphasis> L. (Anthemideae-Asteraceae) from north-east Anatolia

The karyotypes of nine Tanacetum taxa distributed in north-east Anatolia, Turkey, were determined and evaluated by cluster analysis and principal-components analysis. Chromosome numbers were 2n = 2x = 18 (8 taxa) and 4x = 36 (1 taxon). Somatic chromosome numbers of two taxa and a new ploidy level in one taxon are reported for the first time. Karyotype analysis indicated that chromosomes of Tanacetum taxa have predominantly median centromeres. The taxa studied differed significantly in the size of the short arms and long arms, and the arm ratio of each pair of homologous chromosomes, indicating structural rearrangements of the chromosomes have been involved in diversification of the taxa. They were placed in 2A, 3A, and 2B of Stebbins’ karyotype classification, showing the presence of a primitive symmetrical karyotype in the genus. Several systematic and evolutionary aspects of the genus are discussed on the basis of karyological data.     

Assessment of intergenomic recombination through GISH analysis of F1, BC1 and BC2 progenies of Tulipa gesneriana and T. fosteriana

Using 23 F1 hybrids, 14 BC1 and 32 BC2 progenies, the genome composition of Darwin hybrid tulips was analysed through genomic in situ hybridisation (GISH) of somatic chromosomes. All plants were diploids (2n = 2x = 24) with the exception of one tetraploid BC1 (2n = 4x = 48) and one aneuploid BC2 (2n = 2x + 1 = 25) hybrid. Morphometric analysis in F1 hybrids revealed a difference in the total length of chromosomes representing genomes of T. gesneriana and T. fosteriana, where the percentage of each genome equaled 55.18 ± 0.8 and 44.92 ± 0.6% respectively. GISH distinguished chromosomes from both parent genomes although there was a lack of consistent chromosome labelling in some cases. In both T. gesneriana and T. fosteriana chromosomes some segments of heterochromatin in the telomeric and intercalary regions exhibited a higher intensity of fluorescence. In situ hybridisation with 5S rDNA and 45S rDNA probes to metaphase chromosomes of F1 hybrids showed that these regions are rich in rDNA. A notable feature was that, despite genome differences, there was a considerable amount of intergenomic recombination between the parental chromosomes of the two species as estimated in both BC1 and BC2 offspring. The number of recombinant chromosomes ranged from 3 to 8 in BC1 and from 1 to 7 in BC2 progenies. All recombinant chromosomes possessed mostly a single recombinant segment derived from either a single crossover event or in a few cases double crossover events. This explains the fact that, unlike the situation in most F1 hybrids of other plant species, certain genotypes of Darwin hybrid tulips behave like normal diploid plants producing haploid gametes and give rise to mostly diploid sporophytes.     

Karyomorphological studies onWoodwardia sensu lato of Japan

Karyomorphological comparisons were made of five species of JapaneseWoodwardia. There were no marked differences at interphase and prophase among the five species.Woodwardia japonica, W. prolifera, andW. unigemmata were diploid with 2n=68 and the formulas of their metaphase karyotypes uniformly 4m(median centromeric chromosomes)+12sm(submedian)+52(st+t)(subterminal and terminal).Woodwardia orientalis was tetraploid with 2n=136 and 8m+24sm+104(st+t), and the ratio of each chromosomal type to total complement was identical to that of three diploid species. These four species had several characteristics in common:x=34, the longest chromosome of sm, and a mean chromosome length over 3.0 μm. AlthoughWoodwardia orientalis showed some similarity toW. prolifera, it seems to be an allotetraploid which originated by chromosome doubling of a hybrid ofW. prolifera and a diploid species as yet karyomorphologically unknown.Woodwardia kempii was tetraploid with 2n=124 and 8m+24sm+92(st+t), and differed from the others in havingx=31, the longest chromosomes of t, and a mean chromosome length under 3.0 μm. This species has been classified as an independent genus,Chieniopteris, and our karyomorphological study supports this treatment.     

A Thinopyrum ponticum and Triticum aestivum hybrid. Electrophoretic patterns of their endogenous phosphorylation, Diphosphonucleoside kinases, DNases and RNases

Endogenous protein phosphorylation, DNase and RNase electrophoretic patterns, and the detection of NDP-kinases by TLC (Thin Layer Chromatography) were performed in Thinopyrum ponticum (2n=10x=70), Triticum aestivum (2n=6x=42), and their hybrid seedlings in order to accomplish intergeneric hybridization. Octoploid intergeneric hybrids (2n=8x=56) were synthesized in less than 50% of the hybrids. The F₁ hybrid plants resembled Th. ponticum with regard to morphological features and were sterile. Hybrid seedlings revealed very low endogenous phosphorylation and very low NDP-kinase activity in comparison to their parents. In addition hybrid seedlings expressed a new nuclease. Received: 29 June 2000 / Accepted: 28 July 2000