甜菜无融合生殖单体附加系M14大孢子发生的超微结构

应用透射电镜技术探求甜菜单体附加系M14 (Beta vulgaris L.,VV+1C, 2n=18+1)有性生殖大孢子与二倍体孢子生殖大孢子发生时的超微结构特征,了解有性生殖和无融合生殖起始细胞的超微结构差别。甜菜单体附加系M14为兼性无融合生殖体,大孢子发生有韭型、蝶须型和蓼型三种类型。韭型和蓼型大孢子发生时均起源于大孢子母细胞,二分体之前从形态结构上难以区分;减数分裂前期I,细胞核中出现核液泡,形成联会复合体,细胞质改组,细胞壁上沉积胼胝质。韭型大孢子发生时,只进行减数第一次分裂,不发生减数第二次分裂,形成二分体,珠孔端二分体细胞退化,合点端二分体细胞发育为二倍体功能大孢子。蓼型二分体的珠孔端细胞在减数第二次分裂前或分裂过程中退化,合点端细胞经减数第二次分裂形成两个细胞,构成三分体,最终合点端大孢子发育为单倍体功能大孢子。蝶须型大孢子发生是M14中唯一的二倍体孢子无融合生殖方式,其大孢子发生时大孢子母细胞不发生减数分裂,不出现核液泡,未形成联会复合体,无细胞质改组,细胞壁上缺乏胼胝质的沉积和缺乏胞间连丝,这些可作为二倍体孢子无融合生殖的鉴定指标。     

Problems in breeding and cytology of sugar cane

In Part V the phenomenon endo-duplication of chromosomes within embryo-sac mother cells—cause of the increase in chromosome number within species hybrids of Saccharum—was discussed.Special cases of change in chromosome number by hybridization of sugar cane, which formerly were inconceivable, appeared to be based on endo-duplication.In Saccharum species hybrids with an increased chromosome number equalling twice the haploid number of the female parent plus once the haploid number of the male parent, are of such a frequent occurrence that they may be called normal.Also within other genera interspecific and intergeneric hybrids are known of which the somatic chromosome numbers show the same kind of increase. They are, however, less numerous than in Saccharum. Of agricultural plants we know them for instance in Solanum (potato), in intergeneric hybrids between Lolium and Festuca and in hybrids between Triticum monococcum and T. turgidum. In horticultural plants they occur in Cyclamen. About the mode of origin of such hybrids opinions are not uniform. In this article the possibility is considered whether the chromosome increase in such hybrids may be interpreted in the same way as in Saccharum.     

Abnormalities of endosperm development inLilium hybrids

Summary Of 191 interspecific crosses attempted between 27 different species representing different subgenera ofLilium only 14 gave rise to seeds and only 6 of these were viable. Sterility was associated with various types of abnormality in endosperm development. Many of these irregularities involved chromosome breakage and reunion but others entailed abnormal DNA replication and chromosome coiling. In one cross the individual endosperm nuclei contained four stranded structures, the behaviour of which at division was similar to that of bivalents at meiosis. The embryo-sac ofLilium is of theFritillaria type, containing both haploid and triploid polar nuclei. As a consequence the balance between the number of chromosome sets in the embryo, endosperm and maternal tissue is 2:5:2 and not 2:3:2 as is more commonly found in diploid species. It is suggested that sterility results from genetic imbalance of the endosperm itself rather than interaction of the endosperm with either embryo or maternal tissue.     

New C-banding pattern for chromosome identification in cucumber (Cucumis sativus L.)

A chromosome study of cucumber, C. sativus L., was performed using orcein and C-banding techniques. The diploid and tetraploid plants investigated here showed the somatic chromosome numbers 2n=14 and 28, respectively. The haploid chromosome complement was composed of five metacentric and two submetacentric chromosomes. All C. sativus chromosomes had clearly visible C-bands, and each chromosome could be identified unequivocally after C-banding staining, with 13 C-bands appearing in the haploid complement. The haploid complement had a 44.9% ratio of total C-band length to total chromosome length. Chromosomes 1, 2, 4, 5 and 7 had stable C-bands. Three large, dark C-bands appeared at the proximal regions of chromosomes 1 and 2. Chromosome I had quite a large C-band and with a 68.4% ratio of C-band length to short arm length. Chromosome 2 also had quite a large C-band in the pericentromeric region with a 57.6% ratio of C-band length to the full length of this chromosome and possessed an elongated primary constriction in early metaphase. In prometaphase, chromosome 2 showed that the long arm was completely separated from the short arm. The number of secondary constrictions could not be clearly observed because these chromosomes are small and they could not be counted in every metaphase cell. However, six chromosomes seemed to have secondary constrictions in the diploid plants. Two silver-stained bands were observed at primary constrictions of two of the large chromosomes.     

Formation of unreduced gametes is impeded by homologous chromosome pairing in tetraploid <Emphasis Type="Italic">Triticum turgidum</Emphasis> × <Emphasis Type="Italic">Aegilops tauschii</Emphasis> hybrids

It is believed that unreduced gametes with somatic chromosome numbers play a predominant role in natural polyploidization. Allohexaploid bread wheat originated from spontaneous hybridization of Triticum turgidum L. with Aegilops tauschii Coss. Unreduced gametes originating via meiotic restitution, including first-division restitution (FDR) and single-division meiosis (SDM), are well documented in triploid F₁ hybrids of T. turgidum with diploid Ae. tauschii (genomic constitution ABD, usually with 21 univalents in meiotic metaphase I). In this study, two T. turgidum lines known to carry genes for meiotic restitution were crossed to tetraploid Ae. tauschii. The resulting F₁ hybrids (genomes ABDD), had seven pairs of homologous chromosomes and regularly formed 14 univalents and seven bivalents at metaphase I. Neither FDR nor SDM were observed. The distribution of chromosome numbers among progeny obtained by self pollination and a backcross to T. turgidum showed the absence of unreduced gametes. These results suggest that high homologous pairing interfered with meiotic restitution and the formation of unreduced gametes. This may be related to asynchronous movement during meiosis between paired and unpaired chromosomes or to uneven distribution of chromosomes in anaphases, resulting in nonviable gametes.     

Cytological and genetical studies on hybrids between sorghum almum parodi (2n=40) and some diploid (2n=20) species of sorghum

Hybridization between S. almum (2n=40) and diploid (2n=20) species of Sorghum resulted in plants with 2n=40 and 2n=30 chromosomes. As segregation for many characters occurred in the offspring of those with 2n=40, hybrids of this type provide a means of transferring genes from the diploid to the tetraploid species of Eu-sorghum. Segregations for some genes in the progeny of these hybrids revealed heterozygosity in S. almum which may indicate that one of the ancestors of S. almum was a variety of S. vulgare very similar to the commercial grain sorghum.The triploids were only slightly fertile and the chromosome numbers of plants resulting from backcrossing to S. almum ranged from 30 to 46. Some of the plants with the higher chromosome numbers were self-fertile and segregation for genes which were present in the original diploid and tetraploid parents were observed in their off-spring. Backcrossing the triploid to the diploid parent produced fertile plants with 2n=20 and it is possible that the triploids could be used to transfer genes from the tetraploid to the diploid species of Eu-sorghum.The chromosome pairing in the triploids was similar to that expected in an autotriploid, but some non-homologous pairing was detected which may be the result of duplication of some chromosomes or chromosome segments within the genome (n=10) of S. vulgare.     

Discovery of 2n gametes in tetraploid oat Avena vaviloviana

Summary Sexual polyploidization via the action of 2n gametes (gametes with the sporophytic chromosome number) has been identified as the most important evolutionary mode of polyploidization among plant genera. This study was conducted to determine whether 2n gametes are present in the tetraploid level of the genus Avena (2n=4×=28) Twenty tetraploid Avena lines, representing four species and one interspecific hybrid, were screened for pollen grain size in order to differentiate between n and 2n pollen. Avena vaviloviana (Malz.) Mordv. line PI 412767 was observed to contain large pollen grains at a 1.0% frequency. Cytogenetic analyses of pollen mother cells of PI 412767 revealed cells with double the normal chromosome number (i.e., 56 chromosomes at metaphase I and anaphase I). The mode of chromosome doubling was found to be failure of cell wall formation during the last mitotic division that preceded meiosis. The resulting binucleate cells underwent normal meiotic divisions and formed pollen grains with 28 chromosomes. Based on the formation and function of 2n gametes, three models involving diploid and tetraploid oat lines are proposed to describe possible evolutionary pathways for hexaploid oats. If stable synthetic hexaploid oat lines could be developed by utilizing 2n gametes from diploid and tetraploid oat species through bilateral sexual polyploidization, the resulting hexaploids could be used in breeding programs for transferring genes from diploids and tetraploids to cultivated hexaploids.     

Cytological studies of 2n male gamete formation in sugarcane, Saccharum L.

The germplasm for modern sugarcane cultivars (Saccharum spp. hybrids)has been derived principally from S. officinarum (2n = 80), and S. spontaneum (2n = 40 to 128). Diploid gamete formation has been significant in developing cultivated sugarcane, but the cytological basis for the processes involved is not clearly understood. This research investigated microsporogenesis in nine clones of Saccharum spp. Hybrids and in S. officinarum and S. spontaneum. Diploid gamete formation occurred in all 11 lines, but was least frequent in S. spontaneum and S. officinarum which produced 0.5% and 0.8%2n gametes, respectively. In the hybrid lines, 2n gametes were formed infrequencies ranging from 0.9% to 4.4%. Cytological evidence was obtained for dyad and triad formation during microsporogenesis. Detailed analysis of chromosome behaviour at meiosis indicated that 2n male gamete formation is probably attributable to the absence of cytokinesis rather than a combination of asynchrony and non-disjunction. The clones were ranked on the basis of the frequencies with which they formed 4 × 1n microspores and the data were analysed using χ ² tests for homogeneity. These established that theSaccharum spp. hybrids could be designated as either ‘high’ or ‘low’ frequency haploid gamete producers. Conversely, the latter group, which formed diploid gametes most frequently (2.2%–4.4%), can be described as high frequency diploid gamete producers. The identification of clones most frequently forming diploid gametes may facilitate the more rapid recovery of desirable sugarcane genotypes because such clones could be selected for preferential use in clonal improvement. This revised version was published online in July 2006 with corrections to the Cover Date.     

Bilateral sexual polyploidization in potatoes

Summary Bilateral sexual tetraploidization was achieved by intercrossing diploid Phureja × haploid Tuberosum hybrids. Both parents contribute gametes with the somatic number of chromosomes (diplogynoids and diplandroids). More than 100 tetraploid progeny appeared in nine 2x–2x families. The tetraploids are generally more vigorous and significantly outyield their diploid full-sibs. The superiority of the tetraploids is interpreted on the basis of the mode of diplandroid (2n pollen) formation by first meiotic division restitution (FDR). Heterozygosis, epistasis and genetic diversity, if present in the parental diploid genotype, are largely maintained in the 2n gametes formed by FDR. and syngamy brings about more opportunities for heterotic responses in conjunction with polyploidization. Differences in tuber yield among tetraploid clones within families were found to be statistically significant (P <0.05). Thus, unlike somatic chromosome doubling, which can merely increase chromosome numbers, or somatic hybridization, which could in addition bring about heterosis, sexual polyploidization is also associated with genetic variability. The degree of inbreeding and the genotypic structure in the population of newly arisen tetraploids is discussed. The relationship among asexual reproduction, preservation of favorable nonadditive genetic effects by FDR. restoration of sexuality, and polyploidization is explored. The possible evolutionary significance of these factors is emphasized.     

Unreduced 3x gamete formation of allotriploid hybrid derived from the cross of Primula denticulata (4x)?×?P. rosea (2x) as a causal factor for producing pentaploid hybrids in the backcross with pollen of tetraploid P. denticulata

A triploid hybrid, which was obtained from interspecific crosses between tetraploid Primula denticulata (2n = 4x = 44) and P. rosea (2n = 2x = 22), successfully produced 11 plants by backcrossing with pollen of tetraploid P. denticulata. Analysis of ploidy level using flow cytometry and chromosome counting in the 11 BC₁ plants revealed that all progeny had much larger DNA contents and chromosome number than both parents. In this triploid-tetraploid (3x–4x) crossing, progeny was predominantly true or near pentaploid presumably produced by the fertilization between true or near triploid female gamete produced from triploid hybrid and diploid pollen of tetraploid P. denticulata. These results suggest that unreduced (3x) or near triploid female gametes were partially produced by single step meiosis, either first-division restitution or second-division restitution process. The zygotes formed by the fertilization between true or near triploid egg produced by single step meiosis in triploid hybrid and diploid pollen produced by normal meiosis of tetraploid P. denticulata might be the only survivors in embryogenesis.     

Analyses of phenotype and ARGOS and ASY1 expression in a ploidy Chinese cabbage series derived from one haploid

The aim of this research was to improve our understanding of how ploidy level influences phenotype and gene expression in Chinese cabbage (Brassica rapa L. ssp. pekinensis). Haploid plants (2n = 10) was induced by 0.2% colchicine to produce diploid (2n = 20) and tetraploid plants (2n = 40). The aneuploid (2n = 24) was also obtained by hybridization between diploid plants as the female and tetraploid plants. The ploidy levels of all plants were identified through chromosome counts and flow cytometry. Leaves and petals became larger as the ploidy level increased from haploid to diploid, and from aneuploid to tetraploid. Similarly, expression of ARGOS was regulated by genome size, increasing in parallel with the level of ploidy. Among the four ploidy types, expression was stronger in the floral buds than in the leaves. Expression by ASY1 also differed according to ploidy level, being highest in diploid plants, followed in order by tetraploids. Expression was similar between haploids and aneuploids at two stages—prior to and after meiosis—but was higher in the haploids during meiosis. When buds were compared within the same ploidy type at different stages, ASY1 expression was obviously higher during meiosis than either before or after. Our study demonstrated the generation and phenotype of a ploidy Chinese cabbage series derived from one haploid. Expression of genes ARGOS and ASY1 were modulated by genome size in this ploidy series, and the regulated patterns of the two genes was different.     

The production and behaviour of hexaploid hybrids between Hordeum vulgare and H. Bulbosum

Summary To produce hexaploid (or other polyploid) hybrids, diploid or tetraploid Hordeum vulgare was crossed with hexaploid or octoploid H. bulbosum, and perennial triploid hybrids between the two species were treated with colchicine. The crosses did not yield viable plants: seedset was low, the seed aborted and embryo culture was unsuccessful. The colchicine treatments geve rise to plants in which hexaploid chromosome numbers were observed. At the hexaploid level chromosomal instability occurred, resulting in chromosome elimination.The colchicine-treated triploid hybrids showed in the first years after the treatment better fertility after open flowering than untreated plants, but the level of fertility remained very low. The offspring consisted of haploid, diploid and approximately triploid plants like H. vulgare, tetraploid and approximately tetraploid plants like H. bulbosum, and plants with hybrid morphology and unstable chromosome number, which were highly sterile. Thus the crossing barrier between H. vulgare and H. bulbosum could not be broken down at higher ploidy level.     

Monohaploids (n=x=12) from autotetraploid Solanum tuberosum (2n=4x=48) through two successive cycles of female parthenogenesis

Summary Viable monohaploids with 12 chromosomes in their somatic cells were obtained from an autotetraploid cultivar of Solanum tuberosum (2n=4x=48) by inducing female parthenogenesis successively in the tetraploid cultivar and in dihaploids from that cultivar. Both dihaploids and monohaploids were induced using the diploid S. phureja clones IvP 35 and IvP 48 as pollinators. The average frequencies of dihaploids and monohaploids in 1973 were 2.0 and 0.14 per berry respectively.Non-homologous chromosome associations (bivalents, trivalents and even quadrivalents) were observed at metaphase I of meiosis in pollen mother cells of the two monohaploids studied.The occurrence of non-homologous associations of chromosomes during meiosis is discussed. In addition the potential significance of monohaploids for basic research andfor breeding of potatoes is considered.     

The production of 2n pollen in rose

Based on the size differences found between haploid and diploid pollen produced by diploid and tetraploid rose cultivars, respectively, 2n pollen producers were identified in a population of 53 diploid hybrids from a cross between a dihaploid rose, derived from the haploidization of a tetraploid modern cultivar and the diploid species R. wichuraiana. Frequency of 2n pollen producers was estimated in 2002, 2003 and 2004. Highly variable frequencies were found i) within population; ii) during years of observation (between years and between different months in the same year). The variation of 2n pollen production could be related to environmental fluctuations. A cytological analysis of male meiosis was carried out in 10 hybrids randomly chosen. Among meiotic abnormalities leading to 2n pollen formation, triads (containing a 2n microspore at one pole and two n microspores at the other) resulting from abnormal spindle geometry were frequently observed. The mode of 2n pollen formation is genetically equivalent to a First Division Restitution (FDR) mechanism. FDR 2n pollen transmits a high percentage of the heterozygosity from the diploid parent – 2n pollen producer-to the tetraploid offspring.     

Interspecific hybridization between Vicia faba (L.) and Vicia narbonensis (L.): Early pod growth and embryo-sac development

Summary Fertilized embryo-sac development and pod growth was studied in one Vicia faba cultivar, one Vicia narbonensis population and their reciprocal crosses. The initial development of endosperm and embryo was at least four days faster in V. narbonensis than in V. faba. Pods and ovules developed also faster in V. narbonensis than in V. faba. The growth rate of the hybrid pods followed the growth rate of the mother species, but was slower than that of the pods from selfed flowers. In the cross V. narbonensis × V. faba the ovules stopped growing 9 days after pollination, while in the reciprocal cross they stopped growing 15 days after pollination. Hybrid embryo-sacs from V. faba × V. narbonensis were aborted before they reached the stage of 256 endosperm nuclei or 200 embryo cells. Selfed V. faba embryo-sacs reached this stage in less than 9 days after pollination. In the reciprocal cross the embryo-sacs were aborted before they reached the stage of 128 endosperm nuclei or 80 embryo cells. Selfed V. narbonensis embryo-sacs reached this stage at the 4th day after pollination. Given that at these stages the embryo has less than 200 cells it was concluded that an in-ovule embryo culture technique should be developed to obtain hybrid plants.     

Histological analysis of embryo-sac formation and detection of meiotic diplospory in cassava (Manihot esculenta Crantz)

Summary Embryo-sac formation in six cultivated Asian cassava clones and one unknown wild type, was histologically investigated to determine apomictic potential in the genus Manihot. All the 6 clones were found to possess one single sexual embryo-sac containing 6–8 nuclei at maturity. However, one clone, Rayong 3 possessed two functional embryo-sacs at 12 hours after controlled pollination (HACP). The larger embryo-sac was 6-nucleate (all 3 nuclei of the egg apparatus, 1 polar and 2 antipodal cells) and located towards the false micropylar or the nucellar beak region. The smaller embryo-sac also contained a total of 6 nuclei and approximately 10 m³ or 1/2 the volume of the former and located towards the chalazal pole. Percentage sexuality estimates indicated a 100% sexual reproduction in embryo-sacs of all the clones analysed. However, apomeiotic relationships indicate a low (0.3%) degree of meiotic diplosporous embryo-sac formation, thus suggesting facultative apomixis in cv. Rayong 3. This is the first embryological identification and evidence of apomeiosis and apomictic potential in cassava.Contribution from Plant Breeding Laboratory, Applied Genetics & Biotechnology Division, Faculty of Agriculture, Miyazaki University, No. 91.     

Crosses between Beta intermedia bunge and B. vulgaris L.

Summary A tetraploid annual male sterile form of Beta vulgaris L. (2n=4x=36) was crossed with the wild beet species Beta intermedia Bunge (2n=36). The resulting F₁-plants were male sterile annuals being two or three times back-crossed to diploid and tetraploid sugar and fodder beets in the next years. Apart from tetraploid material (36 chromosomes) hexaploid (54 chromosomes) and a number of aneuploid plants developed.The results obtained justify the conclusion that, at a tetraploid level the material mostly propagates apomictically after the F₁ generation. The presence of penta-, hexa-, septa-and even octaploid plants might be explained by assuming that no meiosis has taken place in the crossing partners. Triploid plants are sometimes found in the progeny of hexaploid material and may presumably be considered haploids. Moreover some pentaploid plants were found in the progeny of the open pollinated F₁ which after two generations of bagging are still pentaploids although they produce no pollen. This is another clear indication of apomictic reproduction.The tetraploid generation from the cross between the hexaploid material and diploid sugar beets probably contains the best prospects for breeding.     

Interspecific hybridization in sexual diploid Allium senescens var. minor× apomictic tetraploid A. nutans and sexual diploid A. senescens var. minor× apomictic hexaploid A. senescens

Fourteen interspecific hybrids in sexual diploid Allium senescens var. minor× apomictic tetraploid Allium nutans L. crosses, and eight interspecific hybrids in sexual diploid A. senescens var. minor× apomictic hexaploid A. senescens L. crosses were produced. The number of chromosomes was 2n= 24 in interspecific hybrids of diploid × tetraploid, and 2n= 32 in diploid × hexaploid crosses. Triploid and tetraploid interspecific hybrids showed intermediate parental morphological characteristics. Tetraploid interspecific hybrids of A. senescens var. minor×A. senescens crosses formed two groups based on leaf colour and leaf width. Seeds were formed in 11 out of 14 triploid interspecific hybrids under natural conditions. In cytological observations of parthenogenesis, three out of 12 triploid interspecific hybrids and five out of eight interspecific tetraploid hybrids were observed. Parthenogenesis ranged from 26.0% to 86.0% in five tetraploid interspecific hybrids. Non-parthenogenesis to parthenogenesis segregated in a 3:5 ratio in A. senescens var. minor×A. senescens crosses.     

Cytology and reproductive behavior of diploid, tetraploid and hexaploid germplasm accessions of a wild forage grass: Paspalum compressifolium

Summary Fourteen germplasm accessions of Paspalum compressifolium native from southern Brazil were cytologically and embryologically analysed. The study revealed that one accession was diploid (2n=20), twelve were tetraploid (2n=40) and one was hexaploid (2n=60). This is the first report of diploid and hexaploid cytotypes for this species. Studies on microsporogenesis, megasporogenesis, and embryo sac development indicated that the diploid cytotype had regular meiotic behavior and reproduces sexually. Tetraploid cytotype usually had an important proportion of chromosomes that associated as quadrivalents during meiosis and reproduced by mean of aposporous apomixis. The hexaploid cytotype showed irregular meiotic behavior with about one third of the chromosomes associated as multivalents and reproduced by aposporous apomixis. Thus, P. compressifolium could be an agamic complex. Breeding in this complex is possible due to the presence of diploid sexuals which can be treated by colchicine and pollinated by apomicts.     

Nematode Resistance Derived from Wild Beet and its Meiotic Stability in Sugar Beet

Three different karyotypes of sugar beet with resistance against the beet cyst nematode (Heterodera schachtii) have been investigated. These comprised monosomic addition lines (2n = 19) with one complete chromosome from B. patellarris or B. procumbens, one line with a chromosomal fragment added to the normal sugar beet chromosome complement (2n =18 + fragment) and one diploid line (2n = 18). The fragment originated from a B. procumbens chromosome since during meiosis it formed a univalem. It carries the gene for nematode resistance. Meiotic disturbances like univalems. laggards, anaphase I bridges, fragments and micronuclei were observed in all resistant genotypes. These may result in an exclusion of the chromosome fragment carrying the resistance from the rest of the genome. In the diploid resistant line, a chromosome with a translocation could be distinguished from the other B. vulgaris chromosomes. Meiotic irregularities also appeared in diploid resistant types and are one main reason for low transmission of the resistance. Tin-relationship between meiotic stability and the transmission rate of the resistance gene is discussed.