Pattern Of Chiasma Distribution Research Articles

THE CHROMOSOME ORGANIZATION OF GENES AND SOME TYPES OF EXTRAGENIC DNA IN ALLIUM

We have studied the physical organization of genes and non-coding DNA in two Allium species, A. cepa and A. fistulosum. Expressed sequence tags (ESTs) clones and polymerase chain reaction (PCR) products of gene fragments obtained with primers designed for the gene sequences available in the public GenBank were physically mapped onto chromosomes using a Tyr-FISH technique. The allinase and lacrymatory factor synthase (LFS) genes that encoded enzymes operating in one metabolic way were cloned sequences and physically mapped onto A. cepa and A. fistulosum chromosomes. The alliinase gene probe (1100 bp) hybridized to the distal region of the long arm of chromosome 4 of A. cepa. The LFS gene probe (550 bp) hybridized to the proximal region of the long arm of chromosome 5 in both species. Inter-simple sequence repeats (ISSRs) that are located between SSR loci were used for chromosomal location of microsatellites in A. fistulosum using common FISH. The chromosomal organization of the Tyl-copia group retrotransposons were investigated in A. fistulosum. Dispersed hybridization of the probe along the chromosome arms apart from telomeric ends was detected. Chromosomal distribution of DNA methylation pattern in A. fistulosum L. was studied using a specific antibodies against 5-methylcytosine (anti-5mC). Highly methylated distal regions in all chromosomes were found. Differences in the methylation level between corresponding regions of homologue chromosomes were shown. We describe recent progress in exploiting the ultrasensitive Tyr-FISH technique for development of visual gene maps for chromosomes of A. cepa and A. fistulosum. The results on the chromosomal location of individual genetic loci aided in assembling physical and genetic maps. We related the physical organization of expressed genes to the contrasting patterns of chiasma distribution and to the organization of repetitive DNA family and highly methylated DNA in these two Allium species. INTRODUCTION A half century ago we learned that higher eukaryotic organisms possess much more DNA in their genome than they likely need for genetic information (MacLean, 1973). In onion, the difference between gene fraction (3-5% of the genome) and extragenic DNA fraction (93-97%) is extremely high (Flavell et al., 1974; Stack and Comings, 1979). Onion has one of the largest genomes among monocots (16 415 Mbp per 1C nucleus). The extraordinary huge genome of onion delays the construction of genomic resources for this economically important crop. Onion is used in every home, daily. Onion is the second most valued vegetable crop (FAO, 2010). In Russia in 2010, onion was cultivated on 88,000 ha with total onion production mounted 1,536,300 tons (FAO, 2010). The best way to get a comprehensive insight into chromosome organization of genes and extragenic DNA is the genome sequencing of species. With the development of second generation sequencing techniques that are capable of sequencing thousands of a Corresponding author. Proc. 6 IS on Edible Alliaceae

Inversions of chromosome arms 4AL and 2BS in wheat invert the patterns of chiasma distribution

In many species, including wheat, crossing over is distal, and the proximal regions of chromosome arms contribute little to genetic maps. This was thought to be a consequence of terminal initiation of synapsis favoring distal crossing over. However, in an inverted rye chromosome arm, the pattern of metaphase I chiasmata was also inverted, suggesting that crossover frequencies were specific to chromosome segments. Here, wheat chromosome arms 2BS and 4AL, with essentially entire arms inverted in reverse tandem duplications (rtd), were studied in the MI of meiosis. Inversion-duplication placed the recombining segments in the middle of the arms. While the overall pairing frequencies of the inverted-duplicated arms were considerably reduced relative to normal arms, chiasmata, if present, were always located in the same regions as in structurally normal arms, and relative chiasma frequencies remained the same. The frequencies of fragment or fragment + bridge configurations in AI and AII indicated that of the two tandemly arranged copies of segments in rtds, the more distal inverted segments were more likely to cross over than the segments in their original orientations. These observations show that also in wheat, relative crossover frequencies along chromosome arms are predetermined and independent of the segment location. The segments normally not licensed to cross over do not do so even when placed in seemingly most favorable positions for it.

Unexpected behavior of an inverted rye chromosome arm in wheat

Distal location of chiasmata in chromosome arms is thought to be a consequence of the distal initiation of synapsis. Observations of meiotic behavior of a rye chromosome with an inverted arm show that patterns of chiasma distribution and frequency are also inverted; therefore, the patterns of synapsis and chiasma distribution are independent, and recombination frequency along a chromosome is position-independent and segment-specific. Since cases of random distribution of chiasmata and recombination are known in rye, a genetic mechanism must be present that licenses specific chromosome regions for recombination. Large differences in the metaphase I pairing of the inversion in various combinations of two armed and telocentric chromosomes confirm the major role of the telomere bouquet in early homologue recognition. However, occasional synapsis and chiasmate pairing of the distal regions of normal arms with the proximal regions of the inversion suggest that an alternative mechanism for juxtaposing of homologues must also be present. Synapsis in inversion heterozygotes was mostly complete but in the antiparallel orientation, hence defying homology, but non-homologues never synapsed. Instances of synapsis strictly limited to the chiasma-capable segments of the arm suggest that, in rye, both recombination-dependent and recombination-independent mechanisms for homologue recognition must be present.

Variation in chiasma frequency among eight accessions of Arabidopsis thaliana.

Natural variation in meiotic recombination frequency in Arabidopsis thaliana has been assessed by analyzing chiasma frequency variation among a range of geographically and ecologically diverse accessions. Fifty pollen mother cells at metaphase I of meiosis were analyzed from each of eight accessions and fluorescence in situ hybridization was applied to enable identification of all 10 chromosome arms. There was no significant variation in mean chiasma frequency between plants within accessions, but there was significant variation between accessions. Further analysis confirmed this finding and identified two particular accessions, Cvi and Ler, as having chiasma frequencies significantly lower than those of the other accessions. The analysis also revealed that the pattern of chiasma distribution between arms and among chromosomes is not consistent over accessions. Further detailed analyses were conducted on each individual chromosome (1-5) in turn, revealing that chromosome 4, one of the acrocentric chromosomes, is the least variable while the other acrocentric chromosome (2) is the most variable. These findings indicate the existence of recombination regulatory elements in Arabidopsis and we conclude that it may be possible in the future to identify these elements and determine their mode of action. The practical implications of such developments are considerable.

Colchicine-induced heritable modification of chiasma frequency and distribution in perennial ryegrass (Lolium perenne)

One-week-old seedlings of inbred lines of perennial ryegrass (Lolium perenne L.) were given a 3-h period of treatment with 0.2% colchicine, and this treatment was found to have heritable effects on chromosome behaviour at meiosis. The mean pollen mother cell chiasma frequency was reduced, and there was a corresponding increase in the number of univalents per cell and also changes in cell and bivalent variances. Patterns of chiasma distribution within bivalents were also modified. The effects were seen in the generation in which the treatment was given and they were also observed in the following sexual generation.

Chiasma-based genetic map of the mouse X chromosome.

The X chromosome pair was identified in diakinesis/metaphase I stage mouse oocytes using a repeat sequence DNA probe and fluorescence in situ hybridisation. Chiasma positions along the X bivalent were measured in 57 oocytes from 4 females. Overall, our observations showed that while there were no obvious "hotspots" for chiasma formation along the X chromosome, there was a tendency to favour the distal end. Minimum inter-chiasma distances were substantial indicating the occurrence of strong genetic interference. Estimates of both genetic distances and recombination fractions for any interval along the chromosome can be calculated from the chiasma data. The average chiasma frequency for the X bivalent was 1.37 giving an estimated total genetic map length of 68.5 cM. In general, the pattern of chiasma distribution along the X chromosome resembled that anticipated from recombination distances in published consensus linkage maps. There were, however, some intriguing differences between the two approaches. The reason for these discrepancies are unknown but may be related to lack of precision in cytogenetic mapping of loci, inter-strain and/or interspecies differences in the genetic controls over the distribution of crossover events. One advantage of the chiasma analysis approach is its suitability for investigating these problems.

Long-range effects of colchicine sensitivity on meiosis in Lolium multiflorum L. (Italian ryegrass)

A 3-h treatment period with 0.2 per cent colchicine on 1-week-old seedlings of inbred lines of Italian ryegrass (Lolium multiflorum L.) has long-range effects on chromosome behaviour at meiosis. There is a reduction of mean pollen mother cell chiasma frequency, an increase in univalents and in cell and bivalent variances, and also changes in the pattern of chiasma distribution within bivalents. The effects are seen in the generation in which the treatment is given, and in the following sexual generation.

Pairing competition between identical and homologous chromosomes in autotetraploid rye heterozygous for interstitial C-bands

Pairing competition between identical and homologous non-identical chromosomes is analysed in autotetraploid metaphase I cells of rye where one pair of identical partners bears an interstitial C-band in the long arm of chromosome 6R whereas the other pair of identical partners lacks such a C-band. This makes it possible to study pairing preferences in two distinct regions of the same chromosome arm. A significant excess of associations involving homologous partners is always observed in the proximal segment (from the centromere to the C-band), whereas a good fit with the expected random ratio, or else identical pairing preferences, is found in the distal segment (from the C-band to the telomere). Differences in the processes of pairing and chiasma formation in 6RL, and/or a readjustment in the pattern of chiasma distribution due to heterozygosity for the interstitial C-band as a result of homologous nonidentical pairing, may be responsible for the different behaviour detected in the two regions of the marked arm.

Synaptonemal complex spreading in Allium cepa and Allium fistulosum. II. Pachytene observations: the SC karyotype and the correspondence of late recombination nodules and chiasmata

Pachytene synaptonemal complexes and recombination nodules were analysed, by surface spreading, in the closely related species Allium fistulosum and Allium cepa (both 2n = 16), which show highly contrasting patterns of chiasma distribution. Pachytene observations show that all eight pairs of homologues are fully paired in both species, despite the pronounced localisation of chiasmata in A. fistulosum. Synaptonemal complex karyotype analysis reveals similar marker complexes in both species. These are presumed homoeologues, which, possibly due to the uneven distribution of the higher DNA amount found in A. cepa, rank in slightly different positions in the two karyotypes. Darkly staining ellipsoidal late recombination nodules were observed associated with PTA stained pachytene synaptonemal complexes. The positional distribution of late recombination nodules along synaptonemal complexes corresponds almost exactly to the distribution of chiasmata along metaphase I bivalents in the two species. These observations strongly support the proposal that late recombination nodules are involved in reciprocal meiotic recombination. The frequencies of late recombination nodules at pachytene showed deficits (30% in A. fistulosum, 70% in A. cepa) compared to metaphase I chiasma frequencies. It is suggested that the greater deficit of late recombination nodules in A. fistulosum could be related to a longer duration of meiosis in this species resulting from its greater genomic DNA content.Key words: synaptonemal complex, recombination nodules, Allium.

Synaptonemal complex spreading in Allium cepa and A. fistulosum

The general features and fine structure of homologous chromosome alignment and pairing have been investigated in two species of Allium (A. fistulosum and A. cepa), which have similar karyotypes but very different patterns of chiasma distribution. Although there is no support for the occurrence of a general pre-meiotic alignment of homologous chromosomes, both species show some alignment of homologues as an immediate prelude to synaptonemal complex (SC) formation. In both species pairing usually commences at sub-terminal sites and is succeeded by numerous separate intercalary initiations of pairing in interstitial and distal regions and then in proximal regions. The last parts to pair, in both species, are pericentromeric and telomeric regions. There is, therefore, no evident relationship between the sequence of pairing and chiasma distribution in these species. Regularly alternating convergences and divergences of aligned axial cores (ACs), termed multiple association sites, are frequently observed. It is proposed that these represent potential pairing initiation sites and from observations on their spatial distribution it is argued that they may be evenly distributed through most of the genome. Small spherical or ellipsoid nodules are found at association sites and between closely aligned ACs which persist in the SC segments present during zygotene, but most of them disappear abruptly at the end of zygotene. These are termed zygotene nodules (ZN) and it is proposed that they are involved in matching corresponding sites on homologous chromosomes as well as possibly having a recombinational role. Their composition, structure, mode of action and relationship to pachytene recombination nodules are at present unknown.

Effects of different supernumerary segments on chiasma distribution of the polymorphic species Chorthippus jucundus (Orthoptera: Acrididae)

The analysis of 18 wild populations of the polymorphic species Chorthippus jucundus has revealed the presence of five distal extra segments. They differ in location, size and staining properties. Two of them (L1H and L1h) are located in the largest element and do not C-band, while the remainder (S8H, S8h and S8h1) appear in the shortest chromosome and show positive C-banding. Those located in the same chromosome differ basically in their size. However, any of these blocks induce a similar change in the pattern of chiasma distribution within the affected bivalent. Additionally four patterns of C-banding as regards the presence of distal bands in some autosomes have been detected in the different populations.

Generation by a polymorphic supernumerary segment of recombination in a normally achiasmate proximal region in Acrotylus insubricus (Scopoli) (Orthoptera, Acrididae)

Four of 20 males collected from a Spanish natural population of Acrotylus insubricus (Scopoli) present a supernumerary segment that is distally located in one of the chromosomes of the M9 bivalent. In meiosis this segment is positively heteropyenotic during the first prophase and reverses its pycnosis at first metaphase. The extra material shows "grey" staining after C-banding and fluoresces brightly with acridine orange. The nucleolar expression in this species is not affected by the presence of the supernumerary segment. However, it provokes a considerable alteration in the pattern of chiasma distribution in the M9 bivalent if compared with standard individuals. Certain aspects related to the possible structural influence of the segment in the displacement of the chiasmata are discussed.Key words: Acrotylus, supernumerary segment, chiasma distribution.

Heterochromatin readjusting chiasma distribution in two species of the genus Arcyptera: The effect among individuals and populations

Chiasma variation has been studied in two selected chromosomes from two species of the genus Arcyptera according to the heterochromatin distribution per bivalent. The differences in chiasma distribution found between each karyomorph revealed an underlying tendency for chiasmata to occupy characteristic positions in the bivalents depending on the heterochromatin distribution. It seems that the heterozygosity of the bivalent more than the presence of heterochromatic segments is readjusting a standard pattern of chiasma distribution. The findings of this survey are discussed in relation to the significance and role of genetical recombination in natural populations depending on the frequency of karyomorphs per populations and the pattern of chiasma distribution per individual.

Further studies on chiasma distribution and interference in the human male.

Some unusual patterns of chiasma distribution were noted in a preliminary investigation of meiosis in an infertile male with an apparently normal mitotic karyotype and a normal mean autosomal cell chiasma frequency. A detailed investigation of chiasma distribution on all 22 autosomes revealed that several chromosomes showed a significant change in chiasma distribution and/or mean inter-chiasma distance in comparison with previously published controls. These findings are discussed in relation to the general patterns of chiasma localization in the human male and the role of interference.

Cytogenetics of Lolium perenne

An analysis of the patterns of chiasma distribution within bivalents, in a number of inbred lines of Lolium perenne, reveals a striking potential for variation in this character, as well as a polygenic genetic basis for its control. In parent plants the chiasmata have a regular pattern of localisation which restricts their formation to the ends of the chromosome arms: this strict localisation is progressively lost with inbreeding and novel patterns of distribution are found where chiasmata occur in more interstitial and proximal regions. The variation is continuous and originates from genotypic differences between the different parent plants, rather than from segregation among families deriving from individual parents.

The Effect of a Deficiency on Chiasma Distribution and Frequency in a Male ofStauroderus Scalaris(Orthoptera)

SUMMARYAn individual of Stauroderus scalaris with a heterozygous reduction of the L3 procentric heterochromatin has been found in a Spanish population. Chiasma frequency and distribution have been studied in the three long pairs of the complement and compared with those of a normal individual. Although there are no significant differences in the L3 bivalent chiasma frequency, it has been observed that chiasmata move away from the centromere in the heterozygous individual giving a different pattern of chiasma distribution with respect to a normal individual. Different aspects related to chiasma frequency and distribution in other bivalents are considered.

Variation of Chiasma Formation in Populations of <i>Aegilops spoltoides</i>

Chiasma frequency at PMCs was studied in two successive generations of Aegilops speltoides, the one consisting of six populations, the other of four families. Each family was the progeny of a single plant taken from the corresponding population of the first generation.The analysis of data showed significant differences to exist between populations and between families, as well as between plants within populations and families. Among the overall material three subgroups were recognised.The first one, including two populations and the corresponding families, displayed the lowest inter- and intraplant variation of chiasma frequency in both generations, but a more asymmetrical pattern of chiasma distribution was found in families. The next group, represented by one population-family system, dimorphic by constitution, showed similar variability in population and family, with regards to chiasma frequency. The distribution of chiasmata has proven partly similar, including the same proportion of the predominant bivalent type.The third category was the most variable, containing a B-carrying class, in which mean chiasma frequency differed significantly between the two generations, being lower in B-family. The suggestion is that B chromosomes, when accumulated, exert a reducing effect on chiasma formation.

Is chiasma determination sequential?

MOST cytological studies of chiasma distribution in higher plants and animals are limited to qualitative statements on the pattern of chiasma distribution along bivalents. A few quantitative studies exist, however, based on detailed measurements of chiasma positions along the extended diplotene bivalents of grasshopper and locust spermatocytes1–3. These studies differ in certain respects but agree in concluding that chiasma formation or at least chiasma determination (assuming that the determination of sites for exchange is separable from exchange itself4) occurs sequentially along bivalents. To this extent these studies largely endorse the view originally proposed by Mather5,6 which was inferred from the property of chiasma interference and from observations on the frequencies of crossing over in different chromosome regions of Drosophila melanogaster.

Male and female meiosis in grasshoppers. I. Stethophyma grossum.

Male meiosis in the grasshopper Stethophyma grossum is well known as an example of proximal chiasma localisation. An investigation of female meiosis in oocytes of this species shows that both the frequency and distribution of chiasmata are quite different from the male situation. Mean chiasma frequency per cell (14.98) in considerably higher in females than in males (11.28) which agrees with the trend established in other comparative studies of male and female meiosis. More strikingly, males and females also show not only different but quite opposite patterns of chiasma distribution. In spermatocytes of males, chiasmata are strictly localised proximally in most bivalents, but in oocytes of females very few chiasmata form in proximal regions and nearly all chiasmata form either in distal or interstitial regions. The genetical significance of these findings is considered.

The effect of supernumerary chromosomes on meiosis in Puschkinia libanotica (Liliaceae)

The presence of the partly heterochromatic supernumerary chromosomes in pollen mother cells of Puschkinia libanotica raises the chiasma frequency in the A chromosome bivalents. The pattern of chiasma distribution along each of the five A bivalents was related to the DNA labelling pattern of mitotic chromosomes. Regions that showed heavy labelling at the end of the DNA synthetic phase had fewer chiasmata than lightly labelled regions. As this relation is the opposite to that found by Rees and Evans in another species we regard any correlation between labelling pattern and chiasma distribution as fortuitous.