Intergeneric hybridization between Diplotaxis siifolia,a wild species and crop brassicas

Summary Attempts were made to obtain intergeneric hybrids between Diplotaxis siifolia, a wild species, and cultivars of Brassica (B. campestris, B. juncea, and B. napus). The crosses showed unilateral incompatibility. When the wild species was used as female parent, pollen germination and pollen tube growth were normal, but hybrid seeds aborted due to post-fertilization barriers. Reciprocal crosses (cultivars as female parent) showed strong pre-fertilization barriers; although pollen grains showed germination, pollen tubes failed to enter the stigma. Hybrids were realized in two of the crosses, D. siifolia x B. juncea and D. siifolia x B. napus, through ovary culture. The hybrids were multiplied in vitro by multiplication of axillary shoots, or somatic embryogenesis. Detailed studies were carried out on the hybrid D. siifolia x B. juncea. F₁ hybrids had shrivelled anthers and were pollen sterile. Amphiploids of this hybrid showed 60% pollen fertility and produced seeds upon self-pollination as well as backcross pollination with the pollen of B. juncea.     

Isolation of Allium pollen protoplasts

Studies on protoplasts isolation were carried out with mature pollen grains of 29 samples of species of Allium aflatunense, A. cepa, A. fistulosum, A. karataviense, A. longicuspis, A. nutans, A. odorum, A. sativum and A. schoenoprasum. Surface sterilized pollen grains drifted from crushed anthers were incubated in an enzyme solution containing 1% (w/v) cellulase Onozuka R-10, 1% (w/v) Macerozyme R-10, 0,5 mol l^-1 sucrose and the basal salts of Nitsch medium. Protoplasts were released within 3 to 120 min, either from the pollen grain, through a slightly disturbed germination pore (narrow aperture), or through a wider aperture, when the exine surrounding the germination pore was disturbed. For the first time, protoplasts were obtained from 13 genotypes of 6 Allium species, at a rate of 1 to 30% of the digested intact pollen grains, depending on the genotype.     

Wide hybridization in Brassica

Summary Crosses were made to obtain interspecific hybrids between B. fruticulosa (wild species , 2n = 16) × B. campestris (cultivar , 2n = 20). Although many pollen grains germinated and their tubes entered the style, only about 30% of the ovules received pollen tubes. Fertilized ovules aborted at various stages of development. A few hybrid seeds resulted from hand pollinations in the field, and they showed poor germination and seedling establishment. The in vitro culture of ovaries, ovules, and seeds increased the frequency of obtaining hybrid seeds and plants: the most effective method was ovary culture followed by ovule culture. The hybrid nature of the plants was confirmed through morphological, cytological, and electrophoretic studies. A meiotic analysis of F₁ hybrids (2n = 18) showed that they had 0–5 bivalents and were completely pollen sterile. Electrophoretic analysis of leaf esterases and acid phosphatases of F₁ hybrids revealed bands derived from each parent. Induced amphidiploids of F₁ hybrids contained mostly bivalents, and had about 50% fertile pollen.     

Effects of pollen selection on progeny vigor in a Cucurbita pepo x C. texana hybrid

We examined the effects of pollen selection for rapid pollen-tube growth on progeny vigor. First, we crossed a wild gourd (Cucurbita texana) to a cultivated zucchini (Cucurbita pepo cv Black Beauty) to produce an F₁ and then an F₂ generation. Half of the F₁ seeds were produced by depositing small loads of C. texana pollen onto the stigmas of C. pepo. These small pollen loads were insufficient to produce a full complement of seeds and, consequently, both the fast- and the slow-growing pollen tubes were permitted to achieve fertilization. An F₂ generation was then produced by depositing small loads of F₁ pollen onto stigmas of F₁ plants. The F₂ seeds resulting from two generations of small pollen loads are termed the non-selected line because there was little or no selection for pollen-tube growth rate on these plants. The other half of the F₁ and F₂ seeds were produced by depositing large pollen loads (>10 000 pollen grains) onto stigmas and then allowing only the first 1% or so of the pollen tubes that entered the ovary to fertilize the ovules. We did this by excising the styles at the ovary at 12–15 h after pollination. The resulting F₂ seeds are termed the selected line because they were produced by two generations of selection for only the fastest growing pollen tubes. Small pollen loads from the F₂plants, both the selected and the non-selected lines, were then deposited onto stigmas of different C. pepo flowers, and the vigor of the resulting seeds was compared under greenhouse and field conditions. The results showed that the seeds fertilized by pollen from the selected line had greater vegetative vigor as seedlings and greater flower and fruit production as mature plants than the seeds fertilized by pollen from the non-selected line. This study demonstrates that selection for fast pollen-tube growth (selection on the microgametophyte) leads to a correlated increase in sporophyte (progeny) vigor.     

Cytological mechanism of pollen abortion resulting from allelic interaction of F1 pollen sterility locus in rice (Oryza sativa L.)

Pollen abortion is one of the major reasons causing the inter-subspecific F₁ hybrid sterility in rice and is due to allelic interaction of F₁ pollen sterility genes. The microsporogenesis and microgametogenesis of Taichung 65 and its three F₁ hybrids were comparatively studied by using techniques of differential interference contrast microscopy, semi-thin section light microscopy, epifluorescence microscopy and TEM. The results showed that there were differences among the cytological mechanisms of pollen abortion due to allelic interaction at the three F₁ pollen sterility loci. The allelic interaction at S-a locus resulted in microspores unable to extend the protoplasm membrane with the enlargement of the microspore at the middle microspore stage and finally producing empty abortive pollen. The allelic interaction at S-b locus caused asynchronous development of microspores at the middle microspore stage producing stainable abortive pollen. The allelic interaction at S-c locus mainly led to the non-dissolution of the generative cell wall and finally caused the hybrid F₁ mainly producing stainable abortive pollen. Genotypic identification indicated that the abortive pollen were those with S ^j allele.     

Cytogenetic studies on an intergeneric hybrid betweenAgropyron tsukushiense andElymus mollis

Elymus mollis is distributed widely from Korea to Japan, Kamchatka and Alaska, the northern part of U.S.S.R., and Northern and Eastern Canada, Greenland and Iceland. This species is tetraploid (2n=28). A strain of this species collected in Hokkaido was crossed withAgropyron tsukushiense var.transiens collected in Mishima. From this cross, 22 F₁ plants were produced. Crossability calculated from the number of hybrid plants produced and the number of floret pollinated was 30.6%. The shape of the F₁ spikes was of theAgropyron type but the glumes were hairy as were those of theElymus parent. One of the characteristics distinguishingElymus fromAgropyron is the production of two spikelets at almost all nodes of the rachis. This character was not expressed in the F₁ plants. All pollen grains of the F₁ plants were completely abortive. The average chromosome pairing at the MI of the PMCs of the F₁ amounted to 2.03 bivalents and 30.95 univalents. Almost all bivalents ranging from one to seven were rod-shaped connected with interstitial or terminal chiasma. These results indicate a lack of genomic homology between the three genomes ofA. tsukushiense and the two genomes ofE. mollis. Contribution No. 37 from the Plant Germ-plasm Institute, Faculty of Agriculture, Kyoto University, Kyoto, Japan.     

The genomic relationship between Glycine max (L.) Merr. and G. soja Sieb. and Zucc. as revealed by pachytene chromosome analysis

Summary This study was conducted with the objective of determining the genomic relationship between cultivated soybean (Glycine max) and wild soybean (G. soja) of the subgenus Soja, genus Glycine. Observations on cross-ability rate, hybrid viability, meiotic chromosome pairing, and pollen fertility in F ₁ hybrids of G. max × G. soja and reciprocals elucidated that both species hybridized readily and set mature putative hybrid pods, generated vigorous F₁ plants, had a majority of sporocytes that showed 18II + 1IV chromosome association at diakinesis and metaphase I, and had a pollen fertility that ranged from 49.2% to 53.3%. A quadrivalent was often associated with the nucleolus, suggesting that one of the chromosomes involved in the interchange is a satellited chromosome. Thus, G. max and G. soja genetic stocks used in this study have been differentiated by a reciprocal translocation. Pachytene analysis of F₁ hybrids helped construct chromosome maps based on chromosome length and euchromatin and heterochromatin distribution. Chromosomes were numbered in descending order of 1–20. Pachytene chromosomes in soybean showed heterochromatin distribution on either side of the centromeres. Pachytene analysis revealed small structural differences for chromosomes 6 and 11 which were not detected at diakinesis and metaphase I. This study suggests that G. max and G. soja carry similar genomes and validates the previously assigned genome symbol GG.Research supported in part by the Illinois Agricultural Experiment Station and U.S. Department of Agriculture Competitive Research Grant (85-CRCR-1-1616)     

Effect of genome-plastome interaction on meiosis and pollen development in Oenothera species and hybrids

 Oenothera villaricae Dietrich and Oe. picensis Dietrich, complete translocation heterozygotes, are fully interfertile, giving rise to six discrete classes of true-breeding hybrids from a reciprocal cross. Associated with each parent and hybrid is a characteristic abortive non-staining pollen fraction easily distinguished from fully developed pollen under the light microscope. Pollen abortion has been associated with translocation rings in other angiosperm species, and may characterize such systems. The abortive pollen fraction is significantly different between reciprocal Oenothera hybrids, however (P<0.001), indicative of partial cytoplasmic control. Pollen abortion is most severe in the F₁ hybrid generation, and ameliorates with successive generations of hybrid self-fertilization. Three-way analysis of variance shows significant effects on pollen stainability (a measure of the non-abortive fraction) for nucleus, cytoplasm and selfed hybrid generation, individually or in combinations. This result suggests a combined nucleocytoplasmic basis for the pollen abortion. Correlated with the observation of increased pollen abortion in Oenothera hybrids are meiotic findings of broken chromosome rings (chains, univalents), asymmetric anaphase chromosome distributions and trinucleate tetrads. To test the hypothesis that such anomalous meiotic events play a role in the mechanism of pollen abortion, meiotic disjunction frequency was determined for each parent, F₁ and F₉ selfed hybrid accessions. Three-way analysis of variance shows levels of significance comparable to those noted for pollen stainability (P<0.001) for effects of nucleus, cytoplasm and selfed hybrid generation on disjunction frequency. A high degree of correlation (r ²=0.984) is noted between disjunction frequency and pollen stainability. We conclude that the abortive pollen grains are indeed the products of nondisjunctional meiotic events, which themselves are consequences of hybrid nucleocytoplasmic incompatibility. Received: 28 May 1997 / Revision accepted: 23 July 1997     

Airborne‐pollen pool and mating pattern in a hybrid zone between Pinus pumila and P. parviflora var. pentaphylla

The reproductive isolation barriers and the mating patterns among Pinus pumila, P. parviflora var. pentaphylla and their hybrids were examined by flowering phenology and genetic assays of three life stages: airborne‐pollen grains, adults and seeds, in a hybrid zone on Mount Apoi, Hokkaido, Japan. Chloroplast DNA composition of the airborne‐pollen was determined by single‐pollen polymerase chain reaction. Mating patterns were analysed by estimating the molecular hybrid index of the seed parent, their seed embryos and pollen parents. The observation of flowering phenology showed that the flowering of P. pumila precedes that of P. parviflora var. pentaphylla by about 6 to 10 days within the same altitudinal ranges. Although this prezygotic isolation barrier is effective, the genetic assay of airborne‐pollen showed that the two pine species, particularly P. pumila, still have chances to form F₁ hybrid seeds. Both parental species showed a strong assortative mating pattern; F₁ seeds were found in only 1.4% of seeds from P. pumila mother trees and not at all in P. parviflora var. pentaphylla. The assortative mating was concluded as the combined result of flowering time differentiation and cross‐incompatibility. In contrast to the parental species, hybrids were fertilized evenly by the two parental species and themselves. The breakdown of prezygotic barriers (intermediate flowering phenology) and cross‐incompatibility may account for the unselective mating. It is suggested that introgression is ongoing on Mount Apoi through backcrossing between hybrids and parental species, despite strong isolation barriers between the parental species.     

Intrusion of airborne pollen through open windows and doors

The importance of the transport of pollen by air movement into houses was evaluated using six to eight simultaneously collecting rotorod-type samplers, creating either a sampler line from outdoors to inside the room, or a sampler grid inside a room. The number of incoming pollen grains was highly dependent on the outdoor concentration. The highest concentrations inside (1–2 m distance) and outside (1 m) the room were 600 and 3,250 grains/m³, respectively, in the Betula pollen season and 1,980 and 5,080 grains/m³ in the Pinus season. The pollen concentration and the indoor/outdoor (I/O) ratio decreased as the distance from the ventilation opening increased. Inside the room at a distance of 1–2 m 28%, and at a distance of 3–5 m 12%, of the outside concentration was recorded. In the lower part of the opening the mean proportion was 63% and in the upper part of the opening it was 40%. Efficient ventilation with two open windows increased the I/O ratio and enabled the pollen to spread throughout the room. During the Pinus pollen season 3–35% of the outdoor concentration was simultaneously recorded at six locations inside the room with two open windows and only 0.1–3.6% with one open window. At the same point in the room the I/O ratio varied from <1 to 35%, depending on the sampling conditions. Only a minor effect on the I/O ratio was found between small and large ventilation windows and the door, although it was expected that more air and pollen grains would come indoors through a larger opening.     

Interspecific pollen transfer as a mechanism of competition: Consequences of foreign pollen contamination for seed set in the alpine wildflower,Polemonium viscosum

Summary Earlier studies showed that under natural conditions foreign pollen receipt is correlated with loss of seed set in populations of P. viscosum. Here, we report on experimental pollinations using foreign pollen from two co-flowering species that share pollinators with P. viscosum. Experiments were designed to ascertain whether improper pollen transfer causes reduced seed set under controlled conditions and, if so, to determine the stage at which interference occurs. In flowers pre-treated by pollination with either Mertensia or Castilleja pollen, adherence of subsequently deposited conspecific pollen grains was unaffected. However, conspecific grains had significantly lower germination and ovule fertilization success in flowers receiving foreign pollen than in controls. These results suggest that pollinator infidelity in high alpine plant communities can reduce female reproductive success of P. viscosum by inhibiting the performance of conspecific pollen grains. Negative effects of Mertensia and Castilleja pollen were comparable despite large interspecific differences in pollen grain morphology. Accordingly, susceptibility to foreign pollen interference appears to be an intrinsic property of P. viscosum that is not dependent on the unique structural properties of pollen produced by competitive species.     

Cytogenetical changes of offsprings from the induced tetraploid hybrid betweenRanunculus silerifolius (2n = 16) andR. chinensis (2n = 16) (Ranunculaceae)

Cytogenetical studies were carried out on the successive generations of offsprings from the induced tetraploid hybrid (2n = 32) betweenRanunculus silerifolius (2n = 16) andR. chinensis (2n = 16). Aneuploids, 2n = 30 to 35, frequently occurred. In latter subsequent generations the deviation of aneuploids increased, but the proportion of euploids decreased, accompanied by the reduction of fertility of pollen grains and seed sets. F₂ and F₄ PMCs constantly exhibited meiotic abnormality, i.e. formation of quadrivalents and univalents. The speciation process ofR. cantoniensis (2n = 32), which was presumed to arise from tetraploid hybrids between the above two species, is discussed on the basis of the above evidences.Former contributions of this series areOkada & Tamura (1977) andOkada (1984).     

Experimental analyses of the evolutionary processes inTrillium

In the Japanese species ofTrillium examined here, every interspecific cross produced healthy F₁ progeny at the rate of about 50 to 60 per cent of the total inborn ovules except forT. hagae. These figures were not appreciably different from those of self-pollinated or sib-mated examples of these species.Trillium hagae, a putative natural hybrid betweenT. kamtschaticum (2n=10) andT. tschonoskii (2n=20), accepted all foreign sources of pollen grains in manual pollination. These and other crossing experiments suggest that no appreciable barrier has developed that might cause interspecific crossability and that they are habitual inbreeders. Despite theri crossability, a very low rate of actual pollen flow was detected by examining electrophoretic phenotypes of seeds taken from sympatric populations of these species.     

Concordant clines and significant correlation between floral and pollen characters in Asarum heterotropoides var. heterotropoides (Aristolochiaceae)

To examine geographic differentiation in Asarum heterotropoides var. heterotropoides in Hokkaido Is. in the northern Japan, in which two putative cryptic species have been suspected to exist, extensive and detailed morphological research on 794 individuals from 44 populations throughout Hokkaido Is. was performed. Among the characters examined, the angle between and tip shape of the calyx lobes and the supratecta of the pollen grains were significantly correlated and were found in similar geographic clines. Among them, the pollen showed two discrete states in almost distinct distribution. Multidimensional scaling analysis showed that individuals within each of the two pollen types had different trends in flower characters. Consequently, we assumed a cline from south to north on Hokkaido Is. For the causes of the cline, the two hypotheses were proposed, primary geographic differentiation or extensive introgressive hybridization between two distinct geographical species existing in the past.     

凤仙花属棒凤仙亚属(凤仙花科)植物花粉形态的分类学意义

棒凤仙亚属代表着凤仙花属最先分化出的一支,以多年生草本,多于5朵花的总状花序,侧生萼片4枚,3沟花粉,心皮四室且每室具1枚种子,果实棒锤状,种子椭圆形以及三沟花粉其极面观为三角形而区别于其它种类。该亚属主要分布于中国南部,少数种类辐射分布至华中和中南半岛地区。已有报道证实花粉的形态特征对凤仙花属植物的属下分类具有重要意义,但对棒凤仙亚属的种类少有涉及。为了进一步探讨该亚属花粉的形态特征及其分类学价值,该研究应用扫描电镜技术对国产棒凤仙亚属植物的14种以及凤仙花亚属的窄萼凤仙花的花粉进行了观察。结果表明：棒凤仙亚属的花粉以单粒存在,具3条萌发沟,辐射对称,极面观为三角形或三角状圆形,花粉粒外壁具均匀的网状纹饰,网脊平滑或具波状边缘,网眼中具不同密度的颗粒状突起。棒凤仙亚属的花粉形态与水角属的花粉形态相似,表现出了其原始性;其花粉形态特征与花部形态性状等相关性差,对该亚属的组级分类意义较小,但对于该亚属的种间界定具有重要意义;而其花粉的极面观形状、赤道面观形状以及网状纹饰等变异相对稳定,对于界定该亚属具有重要的分类学价值。     

The pollination biology of two species of Helicteres (Malvaceae) with different mechanisms of pollen deposition

Species of Helicteres are pollinated mainly by hummingbirds and bats. Most species pollinated by hummingbirds have a mechanism of depositing pollen on the top of the pollinator's head such as is shown in this work for Helicteres sacarolha. H. brevispira has an unusual mechanism of depositing pollen grains under the tail or on the abdomen of the hummingbirds. The top of the birds’ head may be considered an efficient place to transport pollen grains, because it is plain, easily accessible for deposition and donation of pollen and not easily accessible for grooming, while the tail is movable and inclined. Thus, H. brevispira pollinators may carry or transfer fewer amounts of pollen grains than H. sacarolha pollinators from one flower to another. If a large amount of pollen grains is lost, a higher quantity of flower or pollen has to be produced to guarantee reproductive success. Plants of H. brevispira set higher number of flowers and pollen grains per flower than H. sacarolha and have also higher rates of fruit and flower abortion. Thus, pollination efficiency of H. brevispira may be reached by high pollen and flower production. Flower change mechanism presented in this species may be also involved with the optimization of pollinator feeding and pollination efficiency.     

Some permeability properties of angiosperm pollen grains,pollen tubes and generative cells

Summary The permeability of pollen grains, pollen tubes and generative cells of Helleborus foetidus and Galanthus nivalis has been investigated using four probes spanning a wide range of molecular weights: 4,6-diamidino-2-phenyl indole (DAPI; mol.wt. 350). Evans blue (mol.wt. 960), FITC-dextran (average mol.wt. 19400) and FITC-albumin (average mol.wt. 67000). DAPI penetrated into the vegetative cells of desiccated and hydrated pollen, and also entered growing pollen tubes. In contrast, the generative cells of hydrated pollen and of pollen tubes were highly resistant to penetration, as they were when isolated in osmotically balancing medium. Evans blue failed to enter intact generative cells under any of the conditions tested. The dye ultimately entered the vegetative cells of some pollen grains, but these were non-germinable. Growing pollen tubes invariably resisted penetration. Neither of the high molecular weight conjugates entered germinable pollen grains or intact pollen tubes. The results suggest that it is highly unlikely that DNA fragments of high molecular weight can enter viable pollen, pollen tubes or generative cells under any normal conditions.     

Relationships of pollen size,pistil length and pollen tube growth rates in Rhododendron and their influence on hybridization

Summary Pollen size and pistil length data have been collected for 93 species of Rhododendron (Ericaceae) belonging to a number of different subgeneric taxa. For a sample of eight species in section Vireya, pollen tube growth in the style after selfor interspecific pollination has been quantified. Pollen volume and the time taken for pollen tubes to reach the ovary were both related to pistil length. Pollen-tube growth rates were generally greater for species with longer pistils and larger pollen. Increasing temperature increased the rate of pollen-tube growth. There was no detectable effect of pollen tube density on tube growth rate in the style. After interspecific pollinations tube growth rates in foreign styles could be faster or slower than in self styles. A semisterile individual with two viable pollen grains per tetrad and a plant grafted as scion to a longer-styled stock both showed more rapid pollen-tube growth than expected on the basis of pistil size. Data collected for 26 species in section Vireya showed that where extreme disparity of pollen/pistil size causes failure of interspecific crosses, one or more bridging species with intermediate pollen/pistil size can generally be selected.     

Ambrosia sp. pollen in Switzerland

The presence of Ambrosia sp. pollen is very irregular inSwitzerland. South to the Alps (Ticino), the yearly meanquantities (1993 to 1997) reach 535 pollen grains in Lugano and114 in Locarno. To the West, 205 pollen grains of Ambrosiasp. are observed as yearly mean total in Geneva. In La Chaux-de-Fonds(100 a.s.l.), a yearly mean of 86 pollen is reached, 66 inNeuchâatel, 59 in Payerne and 74 in Basel. In the Swiss RhoneValley, in Visp, this yearly mean reaches 30 pollen ofAmbrosia sp. A little more in the center of the country, inBern, 31 pollen are monitored as yearly mean quantity. In therest of the helvetic territory, in the North and in the East, theyearly mean are below 15 pollen grains. Since 1991–1992, anincrease has been observed essentially in Ticino, in Geneva,Neuchâatel and La Chaux-de-Fonds. The presence of Ambrosiasp. pollen is often noticed in case of strong west or south-westwind flowing, probably coming from the Rhôone Valley and theBresse in France and the Po Valley in Italy.     

Pollen exine ornamentation in the F2 generation of an interspecific hybrid of Chorisia (Bombacoideae,Malvaceae) in relation to inheritance pattern

Pollen morphology in ten plants of F₂ progeny of an interspecific tree hybrid, Chorisia insignis H. B.& K.×C. speciosa St. Hil (Bombacoideae, Malvaceae) has been studied with a view to have an insight in to the effect of hybridization on the pollen morphological features in F₂ generation. The study is a sequel to an earlier study on the pollen morphology of Chorisia species and their F₁ hybrid, in which case the hybrid pollen uniformly exhibited the apertural features of the male parent and exine features of the female parent. In the F₂ progeny the pollen grains display the apertural feature of the male parent in all the plants. However, with regard to exine ornamentation, variability has been observed. Of the ten plants, two plants exhibit the exine features of the male parent showing empty lumina (without bacula), five plants have their pollen exactly like that of the F₁ plant (showing prominent columellar heads in the lumina of the apocolpium region) and the remaining three plants showing columellae in lumina of both apocolpium and mesocolpium region being different from the above types. It has been inferred that the variability in the exine pattern in F₂ pollen indicates that the pollen exine pattern in hybrids, perhaps, is not unequivocally controlled by sporophytic or gametophytic genomes.