Analysis Of Meiotic Chromosomes Research Articles

Visualizing Meiotic Chromosome Pairing and Segregation in Interspecific Hybrids of Rice by Genomic in situ Hybridization

Meiotic disturbances in F1 hybrids and their progenies are still major problems in wide hybridization. To investigate the genome affinity reflected in chromosome pairing and segregation, we studied chromosome behaviors during meiosis in two interspecific F1 hybrids [O. minuta×O. australiensis (Om×Oa, BCE genome) and Oa×O. ridleyi (Or, EHJ genome)] by using both traditional staining methods and genomic in situ hybridization (GISH). GISH analysis has been successfully performed on mitotic chromosomes to distinguish different Oryza genomes, but relatively fewer systematic analyses of meiotic chromosomes of interspecific hybrids have been reported. In the hybrids, highly irregular chromosome behaviors through meiosis resulted in producing microspores with unbalanced genome. At diakinesis of these two hybrids, most chromosomes present as univalent, with low frequency as bivalents and occasionally as trivalents. In a pollen mother cell, 2 to 8 bivalents and 0 to 4 trivalents were observed in the hybrid Oa×Or, and 1 to 8 bivalents and 0 to 5 trivalents were observed in the hybrid Om×Oa. GISH results indicated that 51.52% bivalents in Oa×Or and 79.65% bivalents in Om×Oa involved allosyndetic association, which indicates that recombination and introgression should be possible if viable backcrosses can be recovered even from triploid hybrids. In this study, we revealed that the meiotic disturbance due to low affinities between parental genomes is the major reason for the sterility of these two triploid interspecific hybrids. The two hybrids showing vigor in reproductive growth are potential genetic resources in future breeding programs. A better understanding of genomic affinities between these distant Oryza species can facilitate planning an effective breeding program by using wide hybridization, and efficient and routine GISH analysis is helpful to monitor alien introgression in the process.

INFECTED FRESHWATER SNAILS, BIOMPHALARIA ALEXANDRINA, THE INTERMEDIATE HOST OF SCHISTOSOMA MANSONI

The objective was to study the chromosomes of Biomphalaria alexandrina snails (class Gastropoda) in control and infected state which could be helpful in understanding how host-parasite relationships in feasible and effective control measures. The chromosomal changes were studied using the air-drying method. The results showed that B. alexandrina had a diploid chromosome number, 2n = 36. Also, the meiotic stages were detected as early- leptotene and late leptotene, zygotene, diplotene, metaphase I. The result also revealed the presence of a primitive sperm, with a conical head and a very long, uni-flagellate tail. A comparative meiotic chromosome analysis between the control and infected ones showed some significant differences, as pachytene and diplotene were more condensed.

Morphological studies and meiotic chromosome analysis of Epimedium elatum (Morr & Decne) - Rare endemic medicinal plant of Northwestern Himalayas in India.

<p><em>Epimedium elatum</em> (Berberidaceace) is a rare endemic medicinal herb of Northwestern Himalayas in India. Recent ethnopharmacological reports have demonstrated its traditional medicinal use against various bone related diseases in the Kashmir Himalayas. It owes its pharmaceutical importance due to high concentration of flavonoid glycosides like Epimedin A, B, C and Icariin which are known mainly for aphrodisiac, antiosteoporosis, anticancer, antioxidant, antiaging, antifatigue and antiviral activities. It is a neglected medicinal plant in Northwestern Himalayan region and may fall in the list of endangered species due to continuous anthropogenic pressures in its native habitats. In this study, we investigated distributional and altitudinal range of this prized species from twenty diverse eco-geographical zones of Kashmir Himalayas for the first time. We also report here its diversity in morphological attributes both in wild and captive cultivation. The species has a very small population size in most of the surveyed habitats with no natural protection. Under cultivation it showed increased plant height (63.09±4.9cm), more number of leaves (95.53±11cm) and flowers (160.76±20cm), indicating importance of high altitude medicinal garden for its immediate <em>ex situ</em> conservation. Further, the acetocarmine staining and squashing of young anthers confirmed it as a diploid species (2n=12) like other <em>Epimedium</em> species. Chromosome number and meiotic abnormalities are also reported for the first time in the species. Finally, constant anthropogenic pressures in Northwestern Himalayas demand immediate <em>in situ</em> and <em>ex situ</em> conservation programmes for <em>E.elatum</em>.<strong></strong></p>

Heteromorphism of “Homomorphic” Sex Chromosomes in Two Anole Species (Squamata, Dactyloidae) Revealed by Synaptonemal Complex Analysis

Iguanians (Pleurodonta) are one of the reptile lineages that, like birds and mammals, have sex chromosomes of ancient origin. In most iguanians these are microchromosomes, making a distinction between the X and Y as well as between homeologous sex chromosomes in other species difficult. Meiotic chromosome analysis may be used to elucidate their differentiation, because meiotic prophase chromosomes are longer and less condensed than metaphase chromosomes, and the homologues are paired with each other, revealing minor heteromorphisms. Using electron and fluorescent microscopy of surface spread synaptonemal complexes (SCs) and immunolocalization of the proteins of the SC (SYCP3), the centromere, and recombination nodules (MLH1), we examined sex chromosome synapsis and recombination in 2 species of anoles (Dactyloidae), Anolis carolinensis and Deiroptyx coelestinus, in which the sex chromosomes represent the ancestral condition of iguanians. We detected clear differences in size between the anole X and Y microchromosomes and found an interspecies difference in the localization of the pseudoautosomal region. Our results show that the apparent homomorphy of certain reptile sex chromosome systems can hide a cryptic differentiation, which potentially may influence the evolution of sexual dimorphism and speciation.

Spermatozoa Production by Triploid Males in the New Zealand Freshwater Snail Potamopyrgus antipodarum.

Asexual lineages derived from dioecious taxa are typically assumed to be all female. Even so, asexual females from a variety of animal taxa occasionally produce males. The existence of these males sets the stage for potential gene flow across asexual lineages as well as between sexual and asexual lineages. A recent study showed that asexual triploid female Potamopyrgus antipodarum, a New Zealand freshwater snail often used as a model to study sexual reproduction, occasionally produce triploid male offspring. Here, we show that these triploid male P. antipodarum (1) have testes that produce morphologically normal sperm, (2) make larger sperm cells that contain more nuclear DNA than the sperm produced by diploid sexual males, and (3) produce sperm that range in DNA content from haploid to diploid, and are often aneuploid. Analysis of meiotic chromosomes of triploid males showed that aberrant pairing during prophase I probably accounts for the high variation in DNA content among sperm. These results indicate that triploid male P. antipodarum produce sperm, but the extent to which these sperm are able to fertilize female ova remains unclear. Our results also suggest that the general assumption of sterility in triploid males should be more closely examined in other species in which such males are occasionally produced. © 2013 The Linnean Society of London, Biological Journal of the Linnean Society, 2013, 110, 227–234.

Meiotic chromosome analysis of the giant water bug, Lethocerus indicus.

The giant water bug, Lethocerus indicus (Lepeletier and Serville) (Heteroptera: Belostomatidae), a native species of Southeast Asia, is one of the largest insects belonging to suborder Heteroptera. In this study, the meiotic chromosome of L. indicus was studied in insect samples collected from Thailand, Myanmar, Loas, and Cambodia. Testicular cells stained with lacto-acetic orcein, Giemsa, DAPI, and silver nitrate were analyzed. The results revealed that the chromosome complement of L. indicus was 2n = 22A + neo-XY + 2m, which differed from that of previous reports. Each individual male contained testicular cells with three univalent patterns. The frequency of cells containing neo-XY chromosome univalent (∼5%) was a bit higher than that of cells with autosomal univalents (∼3%). Some cells (∼0.5%) had both sex chromosome univalents and a pair of autosomal univalents. None of the m-chromosome univalents were observed during prophase I. In addition, this report presents clear evidence about the existence of m-chromosomes in Belostomatidae.

A checkpoint escape

In most dividing cells, the spindle assembly checkpoint (SAC), which monitors kinetochore–microtubule interactions, signals to prevent anaphase onset until all chromosomes are correctly attached to the metaphase spindle. Here, Shao et al. reveal that Xenopus laevis oocytes do not respond to a SAC. The authors first developed a karyotyping technique that allows the analysis of meiotic chromosomes after nuclear envelop breakdown. When treating oocytes during meiosis I with microtubule-depolymerizing drugs that destroyed the spindle, they observed that homologous chromosomes could still divide into single pairs of chromatids, and on time. This suggested that oocytes could progress to meiosis II and arrest at metaphase II, as normal, without a spindle. Similarly, defective oocytes exhibiting monopolar spindles proceeded through monopolar anaphase without delay, indicating that, in X. laevis, the metaphase-to-anaphase transition is not under SAC regulation.

Meiotic chromosome pairing in Actinidia chinensis var. deliciosa

Polyploids are defined as either autopolyploids or allopolyploids, depending on their mode of origin and/or chromosome pairing behaviour. Autopolyploids have chromosome sets that are the result of the duplication or combination of related genomes (e.g., AAAA), while allopolyploids result from the combination of sets of chromosomes from two or more different taxa (e.g., AABB, AABBCC). Allopolyploids are expected to show preferential pairing of homologous chromosomes from within each parental sub-genome, leading to disomic inheritance. In contrast, autopolyploids are expected to show random pairing of chromosomes (non-preferential pairing), potentially leading to polysomic inheritance. The two main cultivated taxa of Actinidia (kiwifruit) are A. chinensis (2x and 4x) and A. chinensis var. deliciosa (6x). There is debate whether A. chinensis var. deliciosa is an autopolyploid derived solely from A. chinensis or whether it is an allopolyploid derived from A. chinensis and one or two other Actinidia taxa. To investigate whether preferential or non-preferential chromosome pairing occurs in A. chinensis var. deliciosa, the inheritance of microsatellite alleles was analysed in the tetraploid progeny of a cross between A. chinensis var. deliciosa and the distantly related Actinidia eriantha Benth. (2x). The frequencies of inherited microsatellite allelic combinations in the hybrids suggested that non-preferential chromosome pairing had occurred in the A. chinensis var. deliciosa parent. Meiotic chromosome analysis showed predominantly bivalent formation in A. chinensis var. deliciosa, but a low frequency of quadrivalent chromosome formations was observed (1 observed in 20 pollen mother cells).

A comparative analysis of karyotypes of three species of Macleaya—producers of a complex of isoquinoline alkaloids

Using a set of methods (C-banding, DAPI-staining, fluorescence hybridization in situ (FISH) with probes of 26S and 5S rDNA, and analysis of meiosis), the first comparative cytogenetic study of three species of Macleaya, producers of complex isoquinoline alkaloids, cordate Macleaya cordata (Willd.) R. Br. (2n = 20), small-fruited Macleaya microcarpa (Maxim.) Fedde (2n = 20) and Macleaya kewensis Turrill (2n = 20), was first carried out. On the basis of morphometric analysis, formulas of karyotypes were made for each species. Species ideograms for M. cordata, M. microcarpa, and M. kewensis were constructed taking into account the polymorphic variants of the C-banding patterns and indicating the location of 26S and 5S rDNA sites. A comparative study revealed that the karyotypes of M. microcarpa and M. kewensis have more in common with each other than with M. cordata. Analysis of meiotic chromosomes suggests of genetic stability of Macleaya genomes. The results of chromosome analysis were used to confirm the close relationship of Macleaya and to clarify their phylogenetic relationships.

Cytogenetic evidence of mixed disomic and polysomic inheritance in an allotetraploid (AABB) Musa genotype

Edible bananas originated mainly from two wild species, Musa acuminata Colla (AA) and Musa balbisiana Colla (BB), and triploid cultivars with an AAA, AAB or ABB genome are the most widely used. In the present study, chromosome pairing affinities are investigated in a sterile AB Indian variety and in its fertile colchicine-induced allotetraploid (AABB) derivative to determine the inheritance pattern of the tetraploid genotype. The potential implications of interspecific recombination and chromosomal composition of diploid gametes for Musa improvement are presented. The pairing of different chromosome sets at diploid and tetraploid levels was investigated through a combination of conventional cytogenetic and genomic in-situ hybridization (GISH) analyses of meiotic chromosomes, leading to a likelihood model of the pairing behaviour. GISH analysis of mitotic chromosomes was also conducted to reveal the chromosome constitution of hybrids derived from crosses involving the allotetraploid genotype. Analysis of chromosome associations at both ploidy levels suggested that the newly formed allotetraploid behaves as a 'segmental allotetraploid' with three chromosome sets in a tetrasomic pattern, three sets in a likely disomic pattern and the five remaining sets in an intermediate pattern. Balanced and unbalanced diploid gametes were detected in progenies, with the chromosome constitution appearing to be more homogenous in pollen than in ovules. Colchicine-induced allotetraploids in Musa provide access to the genetic background of natural AB varieties. The segmental inheritance pattern exhibited by the AABB allotetraploid genotype implies chromosome exchanges between M. acuminata and M. balbisiana species and opens new horizons for reciprocal transfer of valuable alleles.

Broadening Genetic Diversity in Canola Using the C‐Genome Species Brassica oleracea L.

ABSTRACTGenetic diversity in spring type canola (Brassica napus L., AACC genome, 2n = 38) cultivars is narrow. Limited effort has been made to utilize genetic diversity from the diploid species Brassica oleracea L. (CC genome, 2n = 18) due to lack of canola quality traits in seeds of this species. The objectives of this study were to assess the feasibility of introgressing canola quality traits from B. napus into B. oleracea for the purpose of developing canola quality B. oleracea as well as development of B. napus with greater genetic diversity from B. oleracea while retaining canola quality traits. Two inbred (B. napus × B. oleracea) × B. oleracea populations were generated using B. napus ‘Hi‐Q’ and A01‐104NA and B. oleracea var. alboglabra (L. H. Bailey) Musil (Chinese kale). These populations were assessed for seed quality, effectiveness of selection based on morphological traits, genetic diversity using simple sequence repeat (SSR) markers, and ploidy levels using flow cytometry and cytological analysis of meiotic chromosomes. Zero‐erucic and low glucosinolate types were recovered from a relatively small segregating population. Morphological grouping could not reliably be used to select B. oleracea plants with 2n = 18; all BC1S6 families had nuclear DNA content similar to the B. napus parents. Marker analysis revealed a high level of B. oleracea alleles among backcross lines. These findings suggest that introgression of genetic diversity from the diploid B. oleracea C‐genome into stable, canola quality B. napus type lines is feasible and may have great potential in developing genetically diverse spring type varieties.

A Report on Genetic Male Sterility in Uraria picta (Jacq.) DC.

A male sterile-female fertile mutant of Uraria picta (Jacq.) DC. (Family-Leguminosae, Papillionoidae), possessing dwarf habit, normal chromosome behaviour at meiosis and 100.0% sterile pollen grains (1.00% EMS, 6 h; 0.16% over the M2 mutant population), was identified and it segregated into 4 normal and 3 mutant plants at M3 (70 seeds sown, 11 plants germinated, but 7 survived till maturity). Out of 3 mutants, 1 plant showed abnormal meiosis, 100.0% sterile pollen grains with size variations and in some cases pollen grain agglutination, and the other 2 plants instead showed normal meiosis and 100.0% sterile pollen grains; these were designated as MS1 and MS2 respectively. Meiotic chromosome analysis and studies of pollen grains (pollen viability tests: Lugol's Iodine, Aniline blue, x-gal, Amido black, TTC, Neutral red and Methylene blue; DAPI staining for pollen nuclei; SEM analysis) in relation to untreated control revealed possible differential gene behaviour in MS1 (evident in microsporogenesis as well as microgametogenesis) and MS2 (microsporogenesis only). Male sterility is a non-structural nuclear type (monogenic recessive–ms1 ms1) and is being reported for first time in the species.

Analysis of meiotic chromosomes in ejaculates of infertile patients with an increased number of immature germ cells in semen samples.

Seventy-one semen samples of 60 infertile patients with a suggested high number of immature germ cells were assigned for chromosomal analysis. The sperm probes of nine patients did not contain immature germ cell stages apart from spermatids. In 22 probands we could only find early prophase stages or not readable metaphase structures. Only 25 patients could be analysed cytogenetically. The semen sample of each patient contained degenerated germ cell structures associated with desynaptic and fragmented/pulverized metaphase cells. It follows that in each analysed probe we could detect anomalies which can be attributed to errors in the gametic development process and which are possibly connected with the existing teratozoospermia and oligozoospermia. The easy practicality for cytogenetic analysis represents the most important advantage of the described method. The analysis of immature germ cell stages in semen samples makes it possible to bypass or to reduce the incidence of tests biopsies for meiotic chromosome investigation.

Cytogenetic analyses of intergeneric hybrids between barley and nine species of Elymus

Wild species in the Triticeae tribe are very valuable resources for agronomic improvement in cereal crop species. Intergeneric hybrids were produced between several barley cultivars and perennial species in the genera Elymus, Thinopyrum, and Pseudoroegneria. Caryopsis formation and subsequent plantlet regeneration from embryo culture were variable depending on the hybrid combinations. Chromosome numbers and hybrid identity were confirmed by GISH analysis on the somatic cells of the hybrids. While the hybrids showed very robust vegetative growth and exceeded the parental spikes in size, their floral morphologies resembled that of the wild species. Meiotic chromosome analysis revealed that the bivalent formation frequency per cell ranged from 0.06 in Hordeum vulgare 'Betzes' x Elymus curvatus to 3.0 in Elymus humidus x H. vulgare 'Manley'. By GISH analysis on the meiocytes of the hybrid E. humidus x 'Manley', the frequency of autosyndetic bivalents exceeded the allosyndetic bivalent formation, which gave an insight into the genome constitution of E. humidus as an autoallohexploid species. Regardless of the low allosyndetic chromosome pairing between barley and E. humidus, this combination may be useful for further input, since E. humidus is known to carry many valuable genes for biotic and abiotic stress tolerance.

Dissection of a Y-autosome translocation in Cryptomys hottentotus (Rodentia, Bathyergidae) and implications for the evolution of a meiotic sex chromosome chain

We describe the outcome of a comprehensive cytogenetic survey of the common mole-rat, Cryptomys hottentotus, based on G and C banding, fluorescence in situ hybridisation and the analysis of meiotic chromosomes using immunostaining of proteins involved in the formation of synaptonemal complex (SCP1 and SCP3). We identified the presence of a Y-autosome translocation that is responsible for a fixed diploid number difference between males (2n = 53) and females (2n = 54), a character that likely defines the C. hottentotus lineage. Immunostaining, combined with C banding of spermatocytes, revealed a linearised sex trivalent with X(1) at one end and X(2) at the other, with evidence of reduced recombination between Y and X(2) that seems to be heterochromatin dependant in the C. hottentotus lineage. We suggest that this could depict the likely initial step in the differentiation of a true neo-X, and that this may mimic an early stage in the mammalian meiotic chain formation, an evolutionary process that has been taken to an extreme in a monotreme mammal, the platypus.

Rad52-mediated DNA annealing after Rad51-mediated DNA strand exchange promotes second ssDNA capture

Rad51, Rad52, and RPA play central roles in homologous DNA recombination. Rad51 mediates DNA strand exchange, a key reaction in DNA recombination. Rad52 has two distinct activities: to recruit Rad51 onto single-strand (ss)DNA that is complexed with the ssDNA-binding protein, RPA, and to anneal complementary ssDNA complexed with RPA. Here, we report that Rad52 promotes annealing of the ssDNA strand that is displaced by DNA strand exchange by Rad51 and RPA, to a second ssDNA strand. An RPA that is recombination-deficient (RPA(rfa1-t11)) failed to support annealing, explaining its in vivo phenotype. Escherichia coli RecO and SSB proteins, which are functional homologues of Rad52 and RPA, also facilitated the same reaction, demonstrating its conserved nature. We also demonstrate that the two activities of Rad52, recruiting Rad51 and annealing DNA, are coordinated in DNA strand exchange and second ssDNA capture.

TheArabidopsis SKP1homologASK1controls meiotic chromosome remodeling and release of chromatin from the nuclear membrane and nucleolus

During early stages of meiotic prophase I the nucleus undergoes considerable reorganization, including the clustering of telomeres, the release of contacts between chromosomes and the nuclear membrane, the reorganization of the nucleolus, and chromatin remodeling. Using a light squashing technique for the analysis of meiotic chromosomes along with fluorescent in situ hybridization, transmission electron microscopy and immunolocalization studies with antibodies to modified histones, we demonstrate that ASK1 is essential for early nuclear reorganization events. A relatively large number of meiotic alterations have been identified in ask1-1 plants. We show that many of these defects, including alterations in homolog pairing, nucleolus migration and the missegregation of chromosomes, may arise from alterations in chromatin structure and the inability of chromosomes to resolve and release properly from the nuclear membrane and nucleolus during leptotene. These results raise the interesting possibility that ASK1 controls chromatin structure by targeting of either an early regulator of meiotic progression or possibly matrix attachment proteins for destruction.

ASK1, a SKP1 homolog, is required for nuclear reorganization, presynaptic homolog juxtaposition and the proper distribution of cohesin during meiosis in Arabidopsis

Nuclear reorganization and juxtaposition of homologous chromosomes at late leptotene/early zygotene are essential steps before chromosome synapsis at pachytene. We report the results of detailed studies, which demonstrate that nuclear reorganization and homolog juxtapositioning processes are defective in a null mutant, ask1-1. Our results from 4, 6-diamino-2-phenylindole (DAPI)-stained spreads showed that the "synizetic knot", which is typically found in wild type (WT) meiosis during late leptotene and zygotene, was missing in the ask1-1 mutant. Furthermore, ask1-1 meiocytes exhibited only limited homolog juxtaposition at centromere regions at early zygotene. Immunodetection of the cohesin protein SYN1 identified ask1 defects in cohesin distribution from zygotene to anaphase I. Analysis of meiotic chromosomes in ask1-1 and syn1 single mutants, as well as an ask1-1 syn1 double mutant indicate that ASK1 is required for normal SYN1 distribution during meiotic prophase I and suggest that ask1 associated defects may be primarily related to SYN1 mislocalization.

Karyotype evolution in Tilapia: mitotic and meiotic chromosome analysis of Oreochromis karongae and O. niloticus x O. karongae hybrids.

The karyotype of Oreochromis species is considered to be highly conserved, with a diploid chromosome complement of 2n = 44. Here we show, by analysis of mitotic and meiotic chromosomes, that the karyotype of O. karongae, one of the Lake Malawi 'chambo' species, is 2n = 38. This difference in chromosome number does not prevent the production of inter-specific hybrids between O. niloticus (2n = 44) and O. karongae (2n = 38). Analysis of the meiotic chromosomes of the O. niloticus x O. karongae hybrids indicates that three separate chromosome fusion events have occurred in O. karongae. Comparison of the O. karongae and O. niloticus karyotypes suggests that these consist of one Robertsonian fusion and two fusions of a more complex nature.

Molecular-cytogenetic analysis reveals sequence differences between the sex chromosomes of Oreochromis niloticus: evidence for an early stage of sex-chromosome differentiation

Sex determination in the Nile tilapia, Oreochromis niloticus, is primarily genetic, with XX females and XY males. A candidate sex-determining region in the terminal region of the largest chromosome pair has been identified by analysis of meiotic chromosomes. This region shows an inhibition of pairing and synapsis in the XY genotype, but not in XX or YY genotypes, suggesting that recombination is inhibited. Here we show that chromosome microdissection and subsequent amplification by degenerate oligonucleotide-primed PCR (DOP-PCR) can be used to produce in situ hybridization probes to this largest pair of O. niloticus chromosomes. Furthermore, analysis of the comparative hybridization of X and Y chromosome-derived probes to different genotypes provides the first demonstration that sequence differences exist between the sex chromosomes of O. niloticus. This provides further support for the theory that this chromosome pair is related to sex determination and further suggests that the sex chromosomes are at a very early stage of divergence.