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 gamma irradiation on in vitro pollen germination of different Cucumis species

Summary Pollen grains of 14 Cucumis accessions were irradiated with 0, 1, 2, 3 or 4 kGy acute gamma rays and germinated in vitro directly afterwards. Pollen germination was significantly reduced by increasing irradiation dose for all species, except C. melo var. agrestis. Pollen tube growth was generally reduced likewise. Pollen of two C. anguria subspecies was most sensitive to irradiation. Sensitivity of the pollen with respect to pollen tube growth in relation to irradiation dose was inversely related to total DNA amount per nucleus. In vitro germination was not related to DNA amount per nucleus. Results show that the examined Cucumis species, especially C. melo var. agrestis, are sufficiently resistant to irradiation to be used as donor species for in vivo egg cell transformation of the cucumber.     

Hybrid Tea-rose pollen. I. Germination and storage

Summary Germination and storage trials were carried out with pollen of several rose varieties. The pollen grains germinated well in a 15% sucrose solution with 40 ppm boric acid. Staining the pollen with a 0.1% tetrazolium solution and standardizing the degree of colour at which the pollen grains are counted as viable, provided a good viability estimate, simpler to carry out than in vitro germination. Germination capacity and staining ability of the pollen were greatly impeded-about halved-by dehydration during storage in desiccators at low humidity. This effect could be corrected by humidifying the pollen beforehand for about one hour, though this pre-treatment increased the percentage of germinated pollen grains more than the percentage stained. There was no difference between the two percentages in fresh or in deep-frozen pollen.Pollen stored at 1°C and high relative humidity soon lost its germination capacity: between 0 and 20% humidity a considerable proportion of the pollen remained viable for 9 months and longer. Storage for the same period in vacuum-sealed glass tubes at –24°C maintained viability as well or better and would probably prolong it further. Some of the cold-stored pollen induced a reasonable seed set after one year, a low seed set was obtained even after two years of storage at 1°C and low humidity.     

Chromosome association and pollen fertility in some interspecific hybrids of Lilium

Summary Meiotic observations in PMCs were made in eight kinds of interspecific hybrids in Lilium. Three hybrids, 6134-S × L. cernuum. L. regale × L. leichtlinii maximowiczii and L. auratum platyphyllum × L. henryi showed respectively the mean chromosome association frequencies of 1.4_II, 0.8_II and 1.5_II per cell at MI which were the lowest values hitherto reported in Lilium hybrids. In L. longiflorum × L. henryi, 33.3° of the cells had 1 or 2 multivalents of three to five chromosomes in addition to uni- and bivalents. 12° bivalents and 12 univalents were invariably observed in the triploid hybrid L. longiflorum × L. cernuum. The hybrids between the species belonging to the different sections of the genus generally showed high pollen sterility, with some exceptions. A more or less remote genomic homology was found between the different sections in Lilium.     

Comparison of different pollen viability assays to evaluate pollen fertility of potato dihaploids

Summary The main obstacle in breeding potato (Solanum tuberosum L.) dihaploids is the severe limitation of male fertility. To determine pollen viability assays that correlate well to fertility in crosses, results of five different pollen viability assays were compared by correlation analysis with fruit and seed set characters in test crosses, and to pollen tube growth in situ (PL-test). The methods used were: staining the pollen cells with carmino acetic acid (CAA-test); in vitro pollen germination (PG-test); and detection of pollen staining rates after incubation with fluoresceine diacetate (FDA-test), 2-3-5-triphenyle tetrazolium chloride (TTC-test), and 5-bromo-4-chloro-3-indolyle--galactoside (X-Gal-test).The results of test crosses and pollen tube growth in situ correlated with the results of all other assays with the following ranking, from highest to lowest: enzyme activity assays (X-Gal-test, FDA-test, TTC-test), in vitro pollen germination (PG-test), and pollen staining by CAA. The newly developed X-Gal-test for monitoring -galactosidase activity showed the least variation of all assays investigated. Thus, this highly reproducible simple procedure is recommended for male fertility screening.Abbreviations B/F Berries obtained per 100 flowers - CAA Carmino acetic acid - FDA Fluoresceine diacetate - PG Pollen germination rate in vitro - PL Pollen tube growth in situ - S/B Seeds per berry - S/F Seeds per pollinated flower - TTC 2-3-5-triphenyle tetrazolium chloride - X-Gal 5-bromo-4-chloro-3-indolyle--galactoside     

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.     

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.     

Pollen fertility among BC2 offspring of Impatiens interspecific hybrids of New Guinea and Indonesian ancestry

BC_T2 seedlings, derived from the cross Tangeglow (I. hawkeri Bull. × I. aurantiaca Teysm.) × 7851-1 (I. hawkeri Bull. × I. platypetala Lindl.) with 7851-1 as recurrent parent, were tested for presence of pollen fertility. Of 59 BC₂ seedlings, 11 were capable of in vitro pollen germination and also of effecting fertilization and subsequent seed set as pollen parent when crossed with Tangeglow as the seed-parent tester. The Spearman's rank correlation coefficient [r_s] of + 0.63 indicated a moderately good correlation of seedling ranking, based on pollen germination percent in vitro and seed set in vivo. Pollen germination of the 11 pollen-fertile BC₂ seedlings varied from a mean low of 4% to a mean high of 41%. The 3 highest pollen-germinating BC₂ seedlings also had the highest seed sets, but siblings showed a wide range in pollen-germinating ability. The best pollen germination for a seedling was 8% in the BC₁ and 41% in the BC₂. Pollen fertility in the BC₂ is discussed in relation to using interspecific hybridization in an Impatiens breeding program. This revised version was published online in August 2006 with corrections to the Cover Date.     

Freeze preservation of gladiolus pollen

Viability and fertility profiles of cryopreserved gladiolus pollen from 7 cultivars have shown that it is possible to use cryogenic methods for conservation and management of the haploid gene pool in this species. There was no decline in pollen viability (in vitro) levels after 1 and 10 years of cryogenic storage. Field pollinations with cryogenic stored pollen induced capsule and seed set in varying capacities. Long-term cryogenic storage of gladiolus pollen could enhance breeding efficiency through better management of the haploid gene pool resources. Pollen parents could be made available throughout the breeding programme, ensuring guaranteed supply at the time of peak stigma receptivity. A pollen cryobank facility established for this species would increase genetic diversity conservation at the haploid stage.IIHR Contribution No. 112/93     

Cryopreservation of Delphinium pollen at –30 °C

Pollen of garden varieties and species of Delphinium that was cryopreserved at –30 ^°C after 3 hours of air drying and storage on silica gel, had high germination and pollen tube growth in vitroeven after 180-day storage. On the other hand, pollen stored at 25 ^°C showed a marked decrease in the germination rate within 10 days. The best in vitro germination of Delphinium pollen was on a 1% water agar containing 15% sucrose and 0.005 to 0.01% boric acid and at 15 to 20 ^°C. Field pollination with the cryopreserved pollen showed higher fruit and seed set than pollination with pollen stored at 25 ^°C, and was not significantly different from fresh pollen. This revised version was published online in August 2006 with corrections to the Cover Date.     

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.     

The viability and storage of strawberry pollen

Investigations were carried out in 1990 and 1991 to estimate the pollen viability of five strawberry genotypes and their suitability for storage. Pollen viability was assessed by acetocarmine staining, in vitro germination, and in vivo assays. Pollen was stored in darkness at ?18°C for 1 year, and pollen viability was estimated every 4 months during storage. The highest percentage of stained and in vitro germinated pollen grains, respectively, was shown by fresh pollen of cv. ‘Dukat’ (91 and 45%). The lowest values of these characteristics were observed in pollen of cv. ‘Paula’ (48% and 20%, respectively). The best response to storage at ?18°C occurred in pollen of the breeding clone ‘B-302’ and the cv. ‘Redgauntlet’. ‘Paula’ pollen responded least favourably to this. Pollen of the cvs. ‘Dukaf’. ‘Senga Sengana’, and clone ‘B-302’ after storage for 4 months, still had sufficient capacity for fruit set after cross-pollinations.     

Doubled haploids of Coffea canephora: development,fertility and agronomic characteristics

Summary Doubled haploids (DH) of Coffea canephora Pierre were developed using haploid embryos which occur spontaneously in association with polyembryony. The frequencies of polyembryonic seeds and haploid embryos varied according to the parental genotypes. However, production of a large number of DH seemed possible from all genotypes. More than 750 DHs produced from various genotypes were grown under field conditions and evaluated for different characters of agronomic importance. Approximately half of DH genotypes did not survive, suggesting a strong, negative effect of homozygosity. Inbreeding depression is particularly severe on general vigor and reproductive aspects. For several characters studied such as leaf shape, leaf rust resistance and hundred bean weight, considerable genetic variations were observed within and between groups of DHs constituted by the DHs produced from the same clone. Despite their low vigor and reduced fertility, the DHs of C. canephora offer new possibilities in genetic research and coffee breeding.     

Embryo development and fruit-set in pear induced by untreated and irradiated pollen

Summary Flowers of the pear cultivars Conference, Doyenné du Comice and Gieser Wildeman were pollinated with untreated and 50-krad-irradiated pollen. Fruitlets were sectioned at regular intervals up to 6 weeks after pollination. Tubes of the irradiated pollen grew slower, but within two weeks nearly as many embryos were initiated as in the case of untreated pollen. Thereafter, the proportion of degenerated embryos increased, although the endosperm sometimes persisted. Six weeks after pollination only about 21% of the seeds induced by the irradiated pollen still contained normal embryos. At or before that time in-vitro culture of the immature seeds will be necessary because few embryos survive until harvest. The irradiated pollen stimulated a rather large parthenocarpic fruit-set, presumably associated with endosperm rather than embryo development during the first 4–6 weeks after pollination. The possibilities of haploid production are discussed.     

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.     

Fertility restoration of hybrids between Solanum melongena L. and S. aethiopicum L. Gilo Group by chromosome doubling and cytoplasmic effect on pollen fertility

Reproductive fertility traits were studied in the reciprocal hybrids of the eggplant(Solanum melongena L.) and S. aethiopicum L. Gilo Group, and in synthetic amphidiploids to discover whether fertility in these reciprocal hybrids was restored by chromosome doubling. Isozyme and RAPD analyses confirmed hybridity of the hybrids and amphidiploids. Analyses of chloroplast and mitochondrial DNAs confirmed that the cytoplasm of each of the hybrids and amphidiploids was from the maternal parent. Pollen sterility of S. melongena × S. aethiopicum Gilo Group [F₁ (Mel × Aet)] was restored by chromosome doubling, while the reciprocal hybrid S. aethiopicum Gilo Group ×S. melongena [F₁ (Aet × Mel)]and its amphidiploid did not produce any pollen grains; their microspores degenerated without being released from tetrads. Hence the cytoplasm of S. aethiopicum Gilo Group seems to beresponsible for their pollen-non-formation type sterility of the hybrid. Both the F₁ hybrids did not set any fruits by either selfing or backcrossing, while their amphidiploids set fruits after pollinating with pollen from the amphidiploid of F₁ (Mel × Aet). Seeds obtained from both the amphidiploids germinated normally. Chromosome doubling has been effective in restoring fertility of the hybrids. This revised version was published online in July 2006 with corrections to the Cover Date.     

Cytology and pollen grain fertility in populations of Astragalus adsurgens Pall. indigenous to Yunwu Mountain in the Loess Plateau,north-west China

Summary The wild type populations of Astragalus adsurgens Pall. growing on the northern slopes of Yunwu Mountain were investigated as a part of the program on the reclamation of the Loess Plateau, north-west China.In 1990, plants were sampled from two populations indigenous to Yunwu Mountain and Liupan Mountain, and two populations cultivated in experimental plots. Pollen grain fertility of the plants showed a wide variation from 20 to 99%. In 1991, cytological analysis of chromosome behaviour during meiosis was carried out in order to clarify the causal factors of pollen grain sterility. In anaphase-1, a chromatid bridge and an acentric fragment as well as two chromatid bridges were observed in high frequencies. In addition, more than two chromatid bridges per nucleus existed in considerably high frequencies.These observations clearly indicate that inversions are dominant in populations of wild A. adsurgens indigenous to Yunwu Mountain. xxThis study was supported by a Grant-in-Aid for scientific research from the Ministry of Education, Science and Culture, Japan (No. 01102023).     

The effect of chromosome doubling on fertility,meiotic behaviour and crossability of Solanum etuberosum × S. pinnatisectum

Summary Doubling the chromosome number of sterile F₁ hybrids of S. etuberosum × S. pinnatisectum resulted in fertile allotetraploid hybrids (code 4x-EP) with nearly normal chromosome pairing at meiosis. Many seeds were obtained from selfing and sibbing, but seed set varied considerably. The 4x-EP hybrids were successfully crosses as females with S. stoloniferum and S. polytrichon and as males with S. verrucosum. The crosses with S. stoloniferum and S. polytrichon were carried out reciprocally and unilateral incompatibility was detected. Crosses of 4x-EP hybrids as males onto S. phureja and S. tuberosum cv. Gineke produced parthenocarpic berries only. Nearly 1700 flowers of colchicine-doubled S. pinnatisectum × S. bulbocastanum and the trispecific hybrid (S. acaule × S. bulbocastanum) × S. phureja did not yield a single berry after pollination with a pollen mixture from 20 4x-EP plants.It is concluded that the use of the bridging species S. pinnatisectum and either S. stoloniferum, S. polytrichon or S. verrucosum may enable gene transfer from the non-tuberous Etuberosa species to potato cultivars.     

Breeding tomato for pollen tolerance to low temperatures by gametophytic selection

Haploid selection for traits related to pollen cold tolerance in tomato was performed in segregating populations derived from a Lycopersicon esculentum × L. pennellii hybrid. BC₁ populations were obtained by combining normal and low temperature treatments on two stages of pollen development: pollen formation, and germination and pollen tube growth. F₁ hybrids were cultivated under low and normal temperatures and their pollen was used to pollinate L. esculentum plants at low and normal temperatures. The four BC₁ populations obtained were tested for the quality and quantity of pollen produced at low temperatures. The population obtained by cold treatment at both stages had a significantly improved pollen germination ability at low temperatures. The two other coldselected BC₁ populations showed no differences compared with the unselected BC population. A second cycle of pollen selection, corresponding to BC₂, was applied in order to test its persistence in the subsequent generations and the possibility to further improve the character. This second cycle showed no improvement although some plants retained the high pollen germination ability at low temperatures that was observed in the first cycle. Hence, gametophytic selection of some characters related with tomato pollen performance may be feasible, at least for the first cycle of selection.