Phyletic and evolutionary relationships ofBrachyscome lineariloba (Compositae)

A comparison of karyotypes ofBrachyscome breviscapis (2n = 8),B. lineariloba cytodemes E (2n = 10), B (2n = 12) and C (2n = 16) suggests that these species have a homoelogous basic set of four chromosome pairs, two large pairs and two small, and that theB. lineariloba cytodemes E, B and C are related toB. breviscapis by successive additions of small chromosomes. A pronounced asynchrony of chromosome condensation between these large and small chromosomes has been observed. In the artificial hybrids betweenB. dichromosomatica (2n = 4) ×B. breviscapis, and theB. lineariloba cytodemes, theB. dichromosomatica chromosomes are similar in size and condensation behaviour to the small chromosomes ofB. breviscapis and ofB. lineariloba cytodemes E, B and C. Meiotic pairing in these hybrids also demonstrates the strong affinities between these chromosomes. It is suggested thatB. breviscapis may be of amphidiploid origin between a species with two large early condensing chromosome pairs and another,B. dichromosomatica-like species with two small late condensing pairs. It seems most likely that the additional small and late condensing chromosomes inB. lineariloba cytodemes E, B and C are derived from theB. dichromosomatica-like parent, and that each addition increases vigour, fecundity and drought tolerance, allowing these cytodemes to colonize more open and arid environments. Transmission of the univalents in the quasidiploidB. lineariloba cytodeme E was verified as being via the pollen, and not via the embryo sacs.The cytology ofBrachyscome lineariloba (Compositae, Asteroidae), 10.     

Organisation and origin of a B chromosome centromeric sequence from Brachycome dichromosomatica

Brachycome dichromosomatica is an Australian native daisy that has two pairs of A chromosomes and up to three B chromosomes in some populations. A putative B-specific tandem repeat DNA sequence (Bd49) was isolated previously. Here we describe further characterisation of this sequence and investigate its possible origin. Southern analysis showed that all individual B chromosomes examined have highly methylated tandem repeats of Bd49 but differences in banding pattern for distinct B isolates suggested that the sequence is in a state of flux. Using in situ hybridisation, the sequence was shown to be located at the centromeric region of the B chromosome. Southern analysis of genomic DNA with Bd49 demonstrated that multiple copies of the sequence exist in the genomes of B. eriogona, B. ciliaris, B. segmentosa and B. multifida (none of which have B chromosomes) whereas other species tested (including 0B plants of B. dichromosomatica and 0B B. curvicarpa and B. dentata) have few or no copies. Genomic clones and Bd49-like sequences derived by the polymerase chain reaction (PCR) were obtained from five species but determination of phylogenetic relationships within the genus and inference as to the possible origin of the B chromosome were problematic because of extensive intragenomic heterogeneity of the sequences.     

Cytogeographical study of four aneuploids ofCarex oxyandra Kudo in Japan

Chromosome numbers were determined for 342 clones ofCarex oxyandra collected from 35 localities in Hokkaido, Honshu, Shikoku and Kyushu, Japan. Four intraspecific aneuploids, 2n=18, 20, 24 and 26, were found. In meiotic division, only bivalent chromosomes were observed in all clones at metaphases I and II, suggesting that the aneuploids are established gamodemes. In the mitotic metaphase chromosomes, trimodal variation in chromosome length was observed. The 2n=26 clones found on Mt. Hiko had two particularly small chromosomes. The cytodemes with higher number of chromosomes are distributed in more southern areas of Japan.Carex oxyandra, therefore, accompanied with chromosome fragmentations, might spread the geographical distribution to the southern parts. The morphological characters of leaves, spikes, scales, perigynia and nuts were similar among the four cytodemes, except for the small leaves on plants from Yaku Island.     

Chromosomal location and reorganization of the 45S and 5S rDNA in theBrachyscome lineariloba complex (Asteraceae)

The chromosomal locations of the 45S (18S-5.8S-26S) and 5S ribosomal DNA in theBrachyscome lineariloba complex and two related species have been determined by the use of multicolor fluorescencein situ hybridization (McFISH). TheBrachyscome lineariloba complex includes five cytodemes with 2n=4, 8, 10, 12 and 16. Each of the 5S and 45S rDNA loci occurs at two sites on chromosomes in cytodemes with 2n=4. While in cytodemes with 2n=8, 10, 12 and 16, the number of 5S rDNA sites increases from four to eight paralleled to the genomic addition of diploid (4 chromosomes) or haploid (2 chromosomes) dosage. Of the 5S rDNA sites, only one pair is major, except for the cytodeme with 2n=10. The remaining 5S rDNA sites are minor and seem to have reduced the unit number of the 5S rDNA during the successive genomic additions. The 45S rDNA site is detected only at two nucleolar organizing regions in all cytodemes regardless of successive genomic addition. The loss or diminution of 45S rDNA sequences seem to have proceeded more rapidly than 5S rDNA sequences in theB. lineariloba complex.     

The molecular organisation of a B chromosome tandem repeat sequence fromBrachycome dichromosomatica

A high copy, tandemly repeated, sequence (Bd49) specific to the B chromosome and located near the centromere in Brachycome dichromosomatica was used to identify lambda genomic clones from DNA of a 3B plant. Only one clone of those analysed was composed entirely of a tandem array of the B-specific repeat unit. In other clones, the Bd49 repeats were linked to, or interspersed with, sequences that are repetitious and distributed elsewhere on the A and B chromosomes. One such repetitious flanking sequence has similarity to retrotransposon sequences and a second is similar to chloroplast DNA sequences. Of the four separate junctions analysed of Bd49-like sequence with flanking sequence, three were associated with the same A/T-rich region in Bd49 and the fourth was close to a 25 bp imperfect dyadic sequence. No novel B-specific sequences were detected within the genomic clones. Received: 31 December 1995; in revised form: 1 May 1996 / Accepted: 10 May 1996     

The compact Brachypodium genome conserves centromeric regions of a common ancestor with wheat and rice

The evolution of five chromosomes of Brachypodium distachyon from a 12-chromosome ancestor of all grasses by dysploidy raises an interesting question about the fate of redundant centromeres. Three independent but complementary approaches were pursued to study centromeric region homologies among the chromosomes of Brachypodium, wheat, and rice. The genes present in pericentromeres of the basic set of seven chromosomes of wheat and the Triticeae, and the 80 rice centromeric genes spanning the CENH3 binding domain of centromeres 3, 4, 5, 7, and 8 were used as “anchor” markers to identify centromere locations in the B. distachyon chromosomes. A total of 53 B. distachyon bacterial artificial chromosome (BAC) clones anchored by wheat pericentromeric expressed sequence tags (ESTs) were used as probes for BAC-fluorescence in situ hybridization (FISH) analysis of B. distachyon mitotic chromosomes. Integrated sequence alignment and BAC-FISH data were used to determine the approximate positions of active and inactive centromeres in the five B. distachyon chromosomes. The following syntenic relationships of the centromeres for Brachypodium (Bd), rice (R), and wheat (W) were evident: Bd1-R6, Bd2-R5-W1, Bd3-R10, Bd4-R11-W4, and Bd5-R4. Six rice centromeres syntenic to five wheat centromeres were inactive in Brachypodium chromosomes. The conservation of centromere gene synteny among several sets of homologous centromeres of three species indicates that active genes can persist in ancient centromeres with more than 40 million years of shared evolutionary history. Annotation of a BAC contig spanning an inactive centromere in chromosome Bd3 which is syntenic to rice Cen8 and W7 pericentromeres, along with BAC FISH data from inactive centromeres revealed that the centromere inactivation was accompanied by the loss of centromeric retrotransposons and turnover of centromere-specific satellites during Bd chromosome evolution.     

Detection of 5S rDNA and other repeated DNA on supernumerary B chromosomes ofTriticum species (Poaceae)

The supernumerary B chromosomes ofTriticum speltoides andT. tripsacoides were analyzed in mitotic metaphases of spike primordium cells by C-banding and in situ hybridization (ISH) analyzes. B chromosomes ofT. speltoides have larger telomeric and interstitial C-bands, whereas those ofT. tripsacoides are almost completely devoid of C-bands. A prominent ISH site of rye related DNA sequences (using probe pSc119) was detected on B chromosomes ofT. tripsacoides and only a minor ISH site was observed on theT. speltoides B chromosomes. However, two ISH sites of 5S rRNA loci were detected at a terminal and an interstitial location of theT. speltoides B chromosomes. These sites were absent on B chromosomes ofT. tripsacoides. The results are discussed with respect to the phylogenetic origin of these B chromosomes.     

Internal transcribed spacer sequence analyses indicate cytoevolutionary patterns within Brachycome Cass. (Asteraceae)

The sequences of the Internal Transcribed Spacer regions (ITS1 and ITS2) within the genes coding for cytoplasmic ribosomal (r) RNAs on the A chromosome complement of 34 members of the higher plant genus Brachycome (synonym Brachyscome) have been compared. The ITS1 sequence of species within the B. lineariloba complex contains a 56 bp tract that is absent from at least 12 Brachycome species but is present in other species within Brachycome as well as other Asteraceae. Phylogenetic data support the suggestion that the number of chromosomes reduced in several independent Brachycome lineages during speciation. Comparisons with the B chromosome ITS2 of B. dichromosomatica cytodeme A1 suggests an origin of the B chromosome at a time prior to the divergence of the four cytodemes of B. dichromosomatica.     

Chromosome banding in the genusPinus

Somatic chromosomes (2n=24) ofPinus luchuensis Mayr at metaphase were observed by fluorescent banding methods with chromomycin A₃ (CMA) and DAPI. CMA-bands appeared at the interstitial and/or proximal regions of nearly all chromosomes. DAPI-bands appeared at the interstitial and/or centromeric regions of nearly all chromosomes, and pairs of DAPI-dots appeared at the centromeric regions. Each homologous pair of chromosomes in the chromosome complement was identified by the CMA and DAPI fluorescent banding patterns. The interstitial CMA-bands were mostly localized at the secondary constrictions of the Feulgen-stained chromosomes. The fluorescent banding pattern ofP. luchuensis was very similar to that ofP. thunbergii, but was different from that ofP. densiflora.     

Inter- and intraplant C-banding polymorphism in one population ofAegilops comosa subsp.comosa var.comosa (Poaceae)

The Giemsa C-banding pattern of the chromosomes of the native self-pollinatedAegilops comosa subsp.comosa var.comosa was studied. Six of the seven chromosomes of the haploid genome were found to be polymorphic for C-banding patterns. Chromosome A had four variants, chromosome E three variants and each of the chromosomes B, D, and F two variants. Chromosomes E and G were polymorphic for arm length and arm ratio.This paper is part of the doctoral dissertation ofA. Georgiou.     

Taxonomic studies ofAsarum sensu lato

The karyotype and the C-banding pattern in two species ofHexastylis andAsarum epigynum were analysed in detail, and the results obtained were compared with those of the other species ofAsarum, Asiasarum andHeterotropa previously reported. The present results were partially different from the previous reports related to the karyotypes of these species. The karyotype observed in two species ofHexastylis (2n=26) was represented by ten pairs of metacentric chromosomes and three pairs of small subtelocentric chromosomes, which is very similar to that ofAsiasarum in eastern Asia. The C-banding patterns ofHexastylis andAsiasarum, however, were clearly different from each other. A striking difference was found in one of the three pairs of small subtelocentric chromosomes. A Formosan speciesAsarum epigynum had the somatic chromosome number 2n=12 and a highly asymmetrical karyotype composed of mainly subtelocentric chromosomes. These karyological features were remarkably different from those of the other groups inAsarum s.l.     

Repeated DNA sequences inTriticum (Poaceae): Chromosomal mapping and its bearing on the evolution of B and G genomes

The somatic chromosomes ofTriticum turgicum var.durum cv. Langdon andT. dicoccoides (AABB tetraploids),T. timopheevii, andT. araraticum (AAGG tetraploids) were assayed for distribution patterns of a highly repeated 120bp DNA sequence by in situ hybridization. The repeated sequence appears to be an ancient sequence shared withSecale andAegilops. The distribution patterns of the chromosomes were compared to the patterns of the A and B genome chromosomes ofT. aestivum cv. Chinese Spring (AABBDD hexaploid).T. turgidum andT. dicoccoides were observed to have identical in situ hybridization patterns. In both species, nine chromosomes with a total of 21 sites of hybridization were observed. The pattern, with few exceptions, was identical to that of Chinese Spring.T. araraticum andT. timopheevii were observed to have different patterns. InT. araraticum, six chromosomes with 21 total hybridization sites are present while inT. timopheevii nine chromosomes with 19 total sites exist. Major differences in hybridization patterns were observed between the B and G genomes. The divergence of the tetraploid wheats in this study appears to have resulted in changes in location, not in amount, of the ancient repeated sequence.     

Genomic in situ hybridization inArachis (Fabaceae) identifies the diploid wild progenitors of cultivated (A. hypogaea) and related wild (A. monticola) peanut species

Genomic in situ hybridization offers a powerful tool for investigating genome organisation and evolution of taxa known, or suspected, to be allopolyploids. The question of the diploid progenitors of cultivated peanut (Arachis hypogaea, 2n=4x=40) has been the subject of numerous studies at cytogenetical, cytochemical, biochemical and molecular levels, but no definitive conclusions have been reached. The biotinylated total genomic DNA from potential diploidArachis species were separately hybridized in situ to root tip chromosomes ofA. hypogaea and wild speciesA. monticola (2n=4x=40) without or mixed with an excess of unlabelled DNA from the species not used as a probe. Among the range of different species combinations used, the strong and uniform signals given by labelledA. ipaensis DNA when hybridized toA. hypogaea andA. monticola in combination with unlabelledA. villosa DNA indicates that overall molecular composition of twenty chromosomes ofA. hypogaea andA. monticola is very similar toA. ipaensis chromosomes. ProbingA. hypogaea andA. monticola chromosomes with labelled genomic DNA fromA. villosa mixed with unlabelled DNA fromA. ipaensis likewise labelled strongly and uniformly the other twenty chromosomes. BarringA. ipaensis, all the diploidArachis species presently investigated had characteristic centromeric bands in the twenty chromosomes within the complement indicating a clear division ofA. ipaensis from other species. InA. hypogaea andA. monticola only twenty chromosomes showed centromeric bands. These results (i) confirm the allopolyploid nature ofA. hypogaea andA. monticola, (ii) strongly support the view that wildA. monticola and cultivatedA. hypogaea are very closely related, and (iii) indicate thatA. villosa andA. ipaensis are the diploid wild progenitors of the tetraploid species studied. The present results also reveal that the nucleolus organizing region (NOR) originating fromA. villosa alone is expressed in the two tetraploid species.     

Karyotype analysis and heterochromatin differentiation with Giemsa C-banding and fluorescent counterstaining inCephalanthera (Orchidaceae)

Detailed studies of the chromosomes of the three Austrian species of the genusCephalanthera showed them all to have basically similar karyotypes. BothC. damasonium (2n = 36) andC. longifolia (2n = 32) have three large and several classes of smaller chromosome pairs. The karyotype ofC. rubra (2n = 44) is composed of four large and several groups of smaller pairs. The heterochromatin in these species amounts to about 10% of total karyotype length. All the chromosomes have Giemsa-positive centromeres, but only a few have intercalary or terminal bands. Using differential fluorescent staining with DAPI/actinomycin D, quinacrine/actinomycin D (both A-T specific), and chromomycin A₃/distamycin A (G-C specific) three different types of major heterochromatic bands can be characterized in respect of their satellite DNA composition: highly A-T rich, slightly A-T rich, and very G-C rich. The chromosomes ofC. longifolia contain more A-T rich C-bands than those ofC. damasonium, while the latter's have more G-C rich heterochromatin. In both species several C-bands appear as secondary constrictions or gaps in the Feulgen-stained chromosomes, but most likely, in each species there is only one pair of chromosomes where the secondary constrictions function as nucleolus organizing regions. No major intraspecific variation could be observed except on one small chromosome pair ofC. longifolia which had a heteromorphic C-band in most individuals. Possible pathways of karyotype evolution involving polyploidy and Robertsonian events are discussed.     

Structural and numerical chromosomal polymorphism in natural populations ofAlopecurus (Poaceae)

Considerable structural and numerical chromosomal variation has been found in natural populations ofAlopecurus. Interchange heterozygotes, identified by multivalent formation during meiosis, have been recovered in four out of six species studied and are reported for the first time in the diploidsA. bulbosus, andA. myosuroides, and the tetraploidA. pratensis. B chromosomes have been found in two species,A. pratensis andA. myosuroides and also in inter-specific hybrids betweenA. pratensis andA. arundinaceus. The characteristics, distribution and meiotic behaviour of both interchange heterozygotes and B chromosomes are described.     

The karyotype ofFestucopsis serpentini (Poaceae Triticeae) from Albania studied by banding techniques and in situ hybridization

The karyotypes of two populations ofFestucopsis serpentini (2n = 2x = 14) endemic to Albania were investigated in detail by Giemsa C- and N-banding, AgNO₃ staining, and in situ hybridization with an rDNA probe. The complements consisted of 14 large chromosomes, 10 metacentric and 4 SAT-chromosomes, a metacentric and a submetacentric pair. SAT-chromosomes from one population carried exclusively minute satellites, whereas SAT-chromosomes from another population also carried larger polymorphic satellites, suggesting a geographical differentiation. The existence of four chromosomes with nucleolus forming activity was established through AgNO₃ staining; however, the rDNA probe additionally hybridized to intercalary positions in the short arms of two metacentric chromosomes revealing two inactive rDNA sites. C-banding patterns comprised from zero and up to four very small to larger, generally telomeric bands per chromosome giving low levels of constitutive heterochromatin. Similarities in chromosome morphology and C-banding patterns identified the homologous relationships of all chromosomes in one population, but of three pairs only in the other. Reliable identification of homologous chromosomes between plants was only possible for the SAT-chromosomes. A comparison between the C-banded karyotypes ofF. serpentini andPeridictyon sanctum supports their position in two genera.     

Unusual chromosome pairing and B chromosomes inBriza spicata (Poaceae)

One plant from a population ofBriza spicata (Poaceae) was found to have highly irregular meiotic behaviour. It is characterized by having a reduced chiasma frequency, a large between cell variance in chiasma frequency and the formation of multivalents involving pairs of A chromosomes. The B chromosome present in this plant also forms multivalents with a pair of A chromosomes. It is suggested that the normal control of strict bivalent pairing has broken down and homoeologous chromosomes are associating as multivalents. Furthermore, the partial homology of the B chromosome with a pair of A chromosomes is revealed.     

B-Chromosomes inAllium flavum (Liliaceae) and some related species

B-Chromosomes ofAllium flavum, A. stamineum andA. carinatum were C-banded. InA. flavum different types of B's were found, one of them possessing a nucleolus organizer. In the B's ofA. flavum andA. stamineum the banding patterns resemble those found in the standard chromosomes. The B inA. carinatum is only terminally banded. InA. flavum chiasma localization in the B's appears to be dependent on C-band location, just as is the case in the A-chromosomes. An increase in B's was found to cause a small (but significant) increase in chiasma frequency in the PMC's. This may result from an alteration in the nucleotype by the B's.     

Sympatric occurrence of three cytotypes and four morphological types of B chromosomes of Astyanax scabripinnis (Pisces, Characiformes) in the River Ivaí Basin, state of Paraná, Brazil

Chromosomes of Astyanax scabripinnis from the Tatupeba stream, Ivaí basin (state of Paraná, Brazil), were analyzed. Astyanax scabripinnis population presents 3 different diploid numbers (2n = 46, 48 and 50) and B chromosomes in each cytotype. Eighty per cent of the females among individuals of cytotype I (2n = 50) has a metacentric B macrochromosome, whereas three different types of B chromosomes were identified in individuals of cytotype II (2n = 48). Cytotype III (2n = 46) showed two B chromosomes of different morphologic types (metacentric macrochromosomes and acrocentric) in all specimens and cells analyzed. Constitutive heterochromatin pattern for the three cytotypes showed weak markings in centromeric regions and conspicuous blocks in the telomere regions of ST and A chromosomes. Whereas C-banding showed that B chromosomes were totally or partially heterochromatic, a discussion on their behavior and origin was also undertaken.     

Genome size variation and C-band polymorphism inAlstroemeria aurea, A. ligtu andA. magnifica (Alstroemeriaceae)

Among a total of 43 accessions ofAlstroemeria aurea, A. ligtu andA. magnifica nuclear DNA amounts (2C-values) showed significant intraspecific variation, 1.09, 1.21 and 1.15 fold, respectively, when determined through flow cytometric measurements of fluorescence of propidium iodide (PI) stained nuclei. After staining with another fluorochrome, 4,6-diamidino-2-phenylindole (DAPI), an intraspecific variation of 1.10, 1.11 and 1.12 fold, respectively, was found. C-band polymorphisms were present among and within the accessions of all three species. In some cases only very small differences in C-banding pattern were observed. In other cases, however, differences were more prominent. Besides C-band polymorphism, there were also instances of chromosome length polymorphism, which concerned the total chromosome complement or single chromosomes. The variation in nuclear DNA amount inA. aurea andA. ligtu was more or less continuous, except for one accession ofA. ligtu subsp.simsii. Artificial selection and possibly introgression of chromosomes from other species may have moulded the karyotypes of some of the accessions ofA. aurea, a species that has been under cultivation for more than 160 years. The variation as observed inA. magnifica subsp.magnifica was discontinuous and could be due to a broad species concept.