Interspecific hybridization in Cucumis L. II. The crossability of species,an investigation of in vivo pollen tube growth and seed set

Summary The crossability of 12 Cucumis species of African and Asiatic origin was studied in a diallel cross, in order to find ways to realise the cross between the common cucumber (C. sativus L.) and its wild relatives which carry resistances against diseases and pests.Self-pollinations and cross-pollinations within species gave normal pollen tube growth and seet set. The different accessions within a species, as a rule reacted alike in interspecific crosses. In crosses between African species different crossing patterns were found, viz. bilateral congruity, bilateral incongruity and unilateral incongruity. Within C. sativus all accessions intercrossed freely, except one, which displayed unilateral incongruity.Good seed was harvested from several crosses and in some cases embryo culture was needed for further development of seeds. No good seeds were obtained from any cross between a species of the African group and C. sativus L.     

Meiotic chromosome and pollen morphological studies of polyploid Cucumis species

summary Meiotic chromosome studies of polyploid Cucumis species revealed the presence of bivalent chromosome configurations in the tetraploid C. aculeatus and C. zeyheri and hexaploid C. figarei, while a maximum of ten quadrivalents were observed in pollen mother cells of a tetraploid species, C. heptadactylus.Allotetraploidy was not accompanied by an increase in the number of pores per pollen grain, but the autotetraploid and hexaploid species showed a relatively high percentage of 4-porate pollen. The pollen fertility of the tetraploid species was normal.Contribution from the Department of Horticulture and Department of Agronomy. Supported by the Colorado State University Experiment Station and published as Scientific Series Paper No 2403.Former Graduate Research Assistant, Department of Horticulture (present address, 1030 Sanders St., Auburn, Alabama 36830 USA) and Professor, Department of Agronomy, respectively.     

In vitro pollen germination and tube growth of tomato (Lycopersicon esculentum Mill.) and its relation with plant growth

Summary In vitro pollen germination at 10°C, 14°C and 22°C of four groups of two pure line tomato varieties was compared with their plant growth at 19°C D/10°C N under controlled environmental conditions. Generally, pollen germination was slow at 10°C but after 6 h the percentage of germination was similar to that at 22°C. Maximum germination was obtained at 14°C already after 4 h. The longest pollen tubes occurred at 22°C. The two varieties within each group differed significantly from each other for percentage of pollen germination. For one group, this difference was greater at 10°C than at 22°C. The varieties in two groups also differed significantly for pollen tube growth. These differences in pollen tube growth rate were greater at 22°C than at 10°C. There was no clear relationship between pollen germination, pollen tube growth and plant growth in any of the four pairs of varieties. The results are discussed with regard to the possibility of pollen selection at low temperature in order to improve the efficiency of breeding for growth at low temperature.     

Estimating pollen fertility in Solanum species and haploids

Summary Three staining methods (acetocarmine, fuchsin and oxidation of benzidine) and germination in vitro and in vivo were applied to estimate pollen fertility in Solanum species and dihaploids. Pollen was divided into six classes based on shape and contents of the grains. With acetocarmine, fuchsin, peroxidase and germination in vitro 4, 3, 2 and 1 classes, respectively, are supposed to be included in the percentage of good pollen as measured by these methods. This percentage therefore, in more than 96% of the cases studied, shows a decrease in the order indicated. Neither aging of pollen at room conditions nor collecting pollen from flowers on 1–9 days after anthesis does influence the percentage of good pollen with acetocarmine and fuchsin, whereas this percentage drops sharply to zero with peroxidase and germination in vitro. The latter two methods apparently measure as good pollen only the grains with living cytoplasm. When pollen is collected at three successive dates from the same flowers the percentage of good pollen drops sharply with all methods used. There is a relation between quantity of pollen per flower and pollen quality (% good), low-quantity pollen containing significantly lower percentages of good pollen than medium- and high-quantity pollen. The latter two are not significantly different in this respect.From calculations of correlation coefficients it is concluded that only germination of pollen in vitro is significantly correlated with berry and seed set and thus gives a reliable estimate of male fertility. This does not hold true for the two staining methods without due reserve. The peroxidase method is not useful for the Solanum material studied.After standardized pollination the average number of haploid pollen grains on diploid stigmata was found to be 1625±127, that of diploid pollen on tetraploid stigmata 2863±98.A. W. B. Janssen 1975.     

A procedure suitable for in vitro pollen selection inBrassica oleracea

Summary Experiments to determine whetherBrassica oleracea pollen could effect fertilisation following incubation in liquid culture medium are reported. Pollen was incubated in vitro, collected by centrifugation and used to pollinate compatible pistils. While retaining viability and the capacity to germinate such pollen was unable to penetrate the stigma papillae. However, incubated pollen produced functional tubes following pollination of styles from which the stigmas had been removed. These tubes grew through the style to the ovary and viable seed was obtained. The potential application of this procedure in pollen selection is discussed.     

An assessment of thein vitro germination capacity of pollen grains of five tea hybrid rose cultivars

Summary Germination trials of pollen grains of the tea hybrid rose cultivars: Ferry Porche, Bronze Masterpiece, Queen Elizabeth, John F. Kennedy and Lady X, were carried outin vitro, to determine whether these cultivars are suitable as pollen donors in a breeding programme.Pollen grains of all five cultivars germinated poorly in a medium containing sucrose only. Addition of boric acid in the medium improved the germination percentages. The highest germinability was observed in Ferry Porche and Lady X on 15% sucrose+50 ppm boric acid, and 20% sucrose+100 ppm boric acid, respectively. Addition of calcium nitrate in the medium reduced germination percentages in all cultivars.The effect of pH of the medium on pollen grain germinability depended on the cultivar and the composition of the medium. On a medium free of boric acid, the pH values between 5.5 and 7.0 caused a significant increase in the germination percentages of pollen grains of Lady X and Queen Elizabeth. The addition of boric acid decreased the responsiveness of the pollen of all cultivars to the pH changes of the medium.The germination capacity of pollen grains of Lady X and Ferry Porche was not affected by incubation temperatures ranging between 20°C and 30°C.On the basis of the above results Lady X and Ferry Porche are considered the most suitable for pollen donors.     

Differential cold sensitivity of pollen grain germination in two Prunus species

Summary Pollen grains from selected cutivars of almond [Prunus dulcis (Mill.) D. A. Webb] and peach (Prunus persica Batsch L.) were exposed to a range of temperatures between 1°C and 34°C on a thermogradient plate. Pollen germination at temperatures below 9°C was conspicuously greater in almond than peach. Miximal germination percentages were attained at about 16°C (almond) and 23°C (peach). The two species did not differ in their capacity for pollen tube elongation over a broad range of temperatures. Maximal pollen tube elongation occurred at temperatures 5°C to 8°C higher than maximal pollen germination. Species affiliation appeared to be of much greater consequence than chilling requirement or bloom date of the sporophyte in predicting gametophytic response to temperature.     

Pollen and pistil morphology, in vitro pollen grain germination and crossing success of Lens cultivars and species

Morphological and in vitro germination characteristics of pollen altogether with pistil and style length were analysed, so as to carry out a morphological comparison between different lentil (Lens culinaris Medik.) cultivars and wild species, as well as to correlate the morphological and functional statistics with data regarding crossing success. Pollen length and width in the different genotypes were found to have a positive and highly significant correlation with pistil and style length and in vitro pollen tube length. A cluster analysis detected two different plant groups, namely, (i) large‐seeded L. culinaris (macrosperma), and (ii) a second group formed by two subgroups, a first subgroup with L. ervoides standing alone and a second subgroup including small‐seeded L. culinaris (microsperma) and the wild L. culinaris ssp. orientalis, L. odemensis, L. nigricans. A high and significant correlation was detected between crossing success and phenotypic similarity (r > 0.98; P < 0.05), therefore implying that phenotypic similarity, based on pollen morphology and in vitro pollen length together with pistil and style length, is a good predictor of hybridization success.     

油松花粉离体萌发及萌发特征研究

为了加强油松有性杂交,为种子园育种研究奠定基础,对油松花粉离体萌发的适宜条件及萌发特征进行了系统研究。结果表明,油松花粉在无蔗糖培养基中仍具一定萌发能力,蔗糖的添加可显著促进花粉萌发,但高浓度蔗糖可抑制花粉管的生长;油松花粉管易形成分枝,蔗糖浓度与分枝花粉频率呈正相关;硼酸的添加在培养初期可有效加速花粉萌发,但过高浓度的硼酸对花粉萌发具抑制作用;Ca2+对花粉萌发的影响较小,不同浓度Ca2+处理均可获得较高的萌发率;培养温度对花粉萌发的促进作用明显,但温度的持续升高将抑制花粉萌发。综合萌发率、花粉管生长及分枝花粉管频率,油松花粉萌发的最适液体培养基组成为：2.00%蔗糖+0.075%硼酸+0.005% Ca2+,26℃下光照培养,72 h时萌发率可达(96.10±0.47)%。     

Effect of mentor pollen on pistil-pollen incongruities among species of Cucumis L.

Summary The merits of mentor pollen in certain interspecific hybridizations in the genus Cucumis L. were evaluated. African wild cucumber species Cucumis metuliferus Naud. and C. africanus L. and Asian species C. sativus var. hardwickii Alef and var. sikkimensis Hook, were reciprocally crossed. Pollen tube growth was arrested halfway down the style in all combinations except in C. sativus × C. africanus. Mentor pollen irradiated with 100 and 200 krad grew through the style and into the ovules in all species. Fruitset was obtained in all cases when mentor pollen was used either alone or in mixtures. Fruits set after self pollination with mentor pollen yielded only seeds without embryos.In several crosses, aided by mentor pollen, a number of relatively large seeds (with enlarged embryosac) were obtained. A sample of these embryosacs contained a globular structure like an embryo. Embryosacs with embryo-like structures were explanted on several different media but no development was obtained.     

Factors affecting haploid production in wheat using the Hordeum bulbosum system. 1. Genotypic and environmental effects on pollen grain germination,pollen tube growth and the frequency of fertilization

Summary A quantitative examination of pollen grain germination and pollen tube growth within the overy wall was made in selected wheat × tetraploid H. bulbosum hybridizations, to investigate the effect of the wheat and the H. bulbosum genotype on these characters. As expected, variation at the known crossability loci had no effect on pollen grain germination. The frequency of pollen tube penetration of the ovary wall was, however, severely reduced when the dominant alleles were present. Pollen tube penetration was nevertheless observed in 3 of the 110 ovaries examined of the non-crossable cultivar Highbury. The H. bulbosum genotype had a much smaller effect on these characters, but significant differences between the clones were observed in the numbers of pollen tubes initially penetrating the ovary wall. Although two H. bulbosum genotypes showed no significant differences in the number of ovaries with pollen tubes at the base of the ovule, significant differences in the frequency of fertilization were observed. The possible cause of this discrepancy is discussed.The frequency of fertilization in crossable wheat × H. bulbosum hybridizations was improved by the application of gibberellic acid within 10 minutes of pollination, and reduced by an increase in the ambient temperature from 20°C to 26°C. Fertilization following the pollination of non-crossable wheat genotypes was not affected by either of these factors.     

The crossing of common wheat (Triticum aestivum L.) with cultivated rye (Secale cereale L.). I. crossability,pollen grain germination and pollen tube growth

Summary The crossing of common wheat (Triticum aestivum L.) with rye (Secale cereale L.), and especially the action of the crossability genes of wheat, was studied using the readily crossable wheat cv Chinese Spring (genotype kr ₁ kr ₁ kr ₂ kr ₂ >), the poorly crossable wheat cv Hope (genotype Kr ₁ Kr ₁ Kr ₂ Kr ₂ ), as well as the disomic substitution line of chromosome 5B of Hope into Chinese Spring (CS/Hope 5B, genotype Kr ₁ Kr ₁ kr ₂ kr ₂ ). By comparing crossability and actual fertilization, the poor crossability with rye of both cv Hope and the CS/Hope 5B substitution line was shown to result from absence of fertilization. Studies of pollen grain germination and pollen tube growth showed that the dominant alleles of the crossability genes manifested themselves through retardation and eventually inhibition of pollen tube growth at the style base and in the ovary wall. In Hope the growth of all pollen tubes was inhibited, whereas in CS/Hope 5B rarely fertilization was achieved. The recessive alleles of the crossability genes do not seem to have an influence on the growth of rye pollen tubes in wheat pistils.     

The site of action of the crossability genes (Kr 1, Kr 2) between Triticum and Secale. I. Pollen germination,pollen tube growth,and number of pollen tubes

Summary Seven genotypes of common wheat (Triticum aestivum L.) were crossed with rye (Secale cereale L.) in order to find the site or sites of action of the crossability genes, Kr ₁ and Kr ₂, of wheat. The data obtained, by fluorescence microscopy, were compared to the controls (wheat x wheat). The results indicate that the crossability genes have little effect on pollen germination and on the time taken for the pollen tubes to reach the microphyle, irrespective of their crossabilities with rye. The number of pollen tubes reaching the microphyle is, however, affected by the Kr-genes, as high crossable genotypes have more pollen tubes than the low crossable ones. There was a high correlation between the mean number of pollen tubes at the micropyle with seed set, which also reflects the crossability. The Kr-genes seem to manifest themselves in the retardation and inhibition of pollen tube growth between the style base and the top of the embryo sac, where the effect is most distinct in the low crossable genotypes.     

Pollen grain germination and pollen tube growth following in vivo and in vitro self and interspecific pollinations in annual Cicer species

Summary Germination of pollen grains and growth of pollen tubes were studied to determine the cause of barreness in crosses among annual Cicer species. In vivo and in vitro time-course studies and fluorescent microscopy revealed no pollination incompatibility among the selfs, crosses and reciprocals of C. arietinum L., C. reticulatum Lad. and C. cuneatum Rich. In general, Cicer pollen grains germinated and grew on styles of Cicer species. Pollen tube growth was characterized by irregularly spaced and intermittent callose deposits. Failure of seed formation in interspecific pollinations may be attributed to the slowness of pollen tube growth or collapse of fertilized ovules. In addition to these causes, shortness of stamens and sparsity of pollen grains were responsible for flower drop in natural selfs. Although the number of pollen tubes entering the micropyle in interspecific pollinations was low, it may be possible to grow the fertilized ovules on an artificial medium to obtain F₁ plants.     

Compatibility and incompatibility in witloof-chicory (Cichorium intybus L.). 1. The influence of temperature and plant age on pollen germination and seed production

Summary For the production of inbred lines and F₁ hybrids in witloof-chicory information is wanted on characteristics such as the incompatibility system. These characteristics can only be studied properly if the influence of temperature and physiological status of the plant on pollen germination and seed production is known. Investigations were carried out with 9 self-incompatible (SI) and 6 self-compatible (SC) clones in glasshouses of the IVT phytotron at constant temperatures of 10, 14, 17, 20, 23 and 26°C. In general, in vivo pollen germination percentages were rather low after self pollination with an optimum for germination around 17–20°C. No seeds were formed at the lowest temperature (10°C) while seed production for SC clones was usually (rather) good at higher temperatures. At 26°C seed production in some clones decreased. Both pollen germination and seed production decreased at the end of the flowering period. There was a rather positive relationship at e.g. 17 and 20°C between pollen germination after selfing and seed production. When no pollen germination was observed, no seed formation occurred. When pollen grains did germinate, seed development would not necessarily occur in all cases. So this relationship only enables negative mass selection for SC.     

Synthesis and preliminary characterization of a new species (amphidiploid) in Cucumis

A successful interspecific hybridization between cucumber (Cucumis sativus L., 2n = 14) and a wild it Cucumis species, C. hystrix Chakr. (2n = 24) was made via embryo rescue. Hybrid plants (2n = 19; 7 from cucumber and 12 from C. hystrix) were sterile, but morphologically uniform. Self-pollination and backcrossing of F₁ hybrid plants to either parent confirmed presence of both male- and female-sterility that were likely caused by lack of homology and improper pairing during meiosis. While the multiple-branching habit, densely brown hairs (on corolla and pistil), orange-yellow corolla, and ovate fruit of F₁ hybrid plants were similar to that of the C. hystrix parent, the appearance of the first pistillate flower was more similar to that of C. sativus parent. The diameter and internode length of the stem, and the shape and size of leaves and flowers were intermediate when compared to the parents. The chromosome number in the hybrid was doubled through somaclonal variation during embryo culture and regeneration process to restore the fertility. Pollen grains were released and fruits with viable seeds matured on fertile, synthetic amphidiploid plants. The results from flow cytometry indicated that, on average, 7.3% of the morphologically unique regenerants had the 4C DNA content of 2.35 pg relative to the 2C DNA content of the original F₁ hybrid at 1.17 pg and, therefore, were likely chromosome-doubled F₁ hybrids (2n = 38). Nutrition alanalysis indicated that the synthetic species had higher protein (0.78%)and mineral (0.35%) content compared to the normal pickling cucumber(0.62% and 0.27%, respectively), and could be considered a new Cucumis crop having a special place in the future agriculture. Preliminarily evaluation indicates that C. hystrix possesses a high level of root-knot nematode resistance, and that this resistance is partially expressed in the interspecific F₁ and chromosome-doubled F₁. This and the fact that the fruit morphology of the fertile amphidiploid differs during the growing season (e.g., short and long fruit) suggest that it could be useful in broadening the germplasm base of cucumber. This revised version was published online in July 2006 with corrections to the Cover Date.     

Viability and storage of bromeliad pollen

Several bromeliad species from two different subfamilies, were used to develop a reliable method to evaluate pollen viability. Pollen germination on a medium containing 20% sucrose, 0.001%H₃BO₃ and 0.5% agar was comparable to germination on a compatible stigma. Maximum germination was reached within 2 to 10 hours depending on the species. Based on this test, six species were considered as being good pollen donors with germination percentages between 49%and 83%. Furthermore, pollen from these species and cultivars could be stored in liquid nitrogen (–196 ^°C) without a considerable loss of viability. For all species, a dehydration period of 4 hours prior to cryopreservation and a rehydration period of 1 hour after cryostorage were essential. Greenhouse humidity influenced anther moisture content and cryostorability. This revised version was published online in July 2006 with corrections to the Cover Date.     

Low temperature tolerance of biomass and pollen germination in potato clones of Andean and European origin

Summary The temperature-related performances of six tetraploid potato clones, two Andean, three European, and one hybrid, were compared by cultivating them in growth chambers under one of two temperature regimes: 10°C day/4°C night or 20°C day/10°C night. Preformance was measured in terms of biomass production and pollen germinability.For dry matter yield at second harvest, the temperature-related performance ratios (performance at 10°C/4°C divided by performance at 20°/10°C) were highest for the Andean clones, intermediate for the Andean/European hybrid clone 201₅×S₁₂, and lowest for the European cultivars. The ability of the European cultivars to maintain their normal rates (i.e. rates at 20/10°C) of biomass production when cultivated at low temperatures varied greatly, with clone S₃ performing most poorly at 10/4°C.For pollen germinability in vitro, performance ratios were highest for the hybrid clone 201₅×S₁₂ and lowest for the European clones.The high tuber yield of hybrid 201₅×S₁₂ at 10/4°C can be attributed to its low-temperature tolerance and it being daylength neutral.     

Effects of high temperatures on mature pollen grains in wild and cultivated maize accessions

Summary The effects of high temperature on mature pollen of various maize lines were investigated. Genotypic differences in pollen reaction to high temperature were revealed. Pollen grains resistant to high temperature (35°C, 26°C) were characterized by higher germination capacity and better ability to develop normal pollen tubes. The studies are of interest to evaluate reproductive system tolerance and conduct gamete selection at the mature pollen grain stage in maize.     

Effect of gamma and ultraviolet irradiation on adventitious regeneration from in vitro cultured pear leaves

Summary Irradiation of in vitro explants and subsequent adventitious regeneration has been tested for 4 commercially important varieties of pear (Pyrus communis) with the aim to create mutants with a reduced susceptibility to fire-blight (Erwinia amylovora). The effect of gamma and ultraviolet irradiation of leaves on adventitious regeneration ability has been studied. The LD50 (50% decrease of regeneration) after gamma irradiation was genotype-dependent and was between 20 and 50 grays. The curves of regeneration showed a threshold dose underneath which none or a very slight decrease was registered. The decrease might result from cumulative events. After an ultraviolet irradiation as low as 62.5 J/m², the leaves became crumbly and rolled up, and their metabolism seemed to be altered. The LD50 was about 125 J/m² for all varieties and the decrease of regeneration was linear. Histological investigations showed leaves with flattened epidermal cells after ultraviolet irradiation and slack spongy parenchyma after gamma irradiation.Abbreviations Gy grays - J joules - LD lethal dose