Non-germinating Pollen Research Articles

Physiological Assessment of Temperature Stress Tolerance in Selectively Fertilzed Coconut Hybrids

In vitro pollen germination study is one of the important technologies for understanding the functions of pollens as well as in many areas of pollen biotechnology particularly in pollen selection. Ability to separate germinated and non-germinated pollen offers selective treatments and can be used for identifying stress tolerant and sensitive alleles carried by their pollen grains. The present study was carried out in Kerasree and Keraganga selectively fertilized (S.F) coconut hybrids produced through pollen selection. Pollens were subjected to germination in specific media maintained at different levels of water stress induced by PEG 6000 and the critical water potential for pollen germination was identified. The water stress tolerant pollen grains at critical level were selected and used for fertilization. Evaluation of temperature stress tolerance of these two selectively fertilized hybrids along with their normal hybrids and west coast tall (WCT) was carried out by assessing their critical temperature stress for pollen germination. Both Kerasree selectively fertilized and Keraganga selectively fertilized hybrids recorded highest critical temperature of 420C for pollen germination with germination percentage of 20.17 and 22.77 respectively compared to their normal hybrids (380C) and WCT (400C).

A simple method for measuring pollen germination rate using machine learning.

The pollen germination rate decreases under various abiotic stresses, such as high-temperature stress, and it is one of the causes of inhibition of plant reproduction. Thus, measuring pollen germination rate is vital for understanding the reproductive ability of plants. However, measuring the pollen germination rate requires much labor when counting pollen. Therefore, we used the Yolov5 machine learning package in order to perform transfer learning and constructed a model that can detect germinated and non-germinated pollen separately. Pollen images of the chili pepper, Capsicum annuum, were used to create this model. Using images with a width of 640 pixels for training constructed a more accurate model than using images with a width of 320 pixels. This model could estimate the pollen germination rate of the F2 population of C. chinense previously studied with high accuracy. In addition, significantly associated gene regions previously detected in genome-wide association studies in this F2 population could again be detected using the pollen germination rate predicted by this model as a trait. Moreover, the model detected rose, tomato, radish, and strawberry pollen grains with similar accuracy to chili pepper. The pollen germination rate could be estimated even for plants other than chili pepper, probably because pollen images were similar among different plant species. We obtained a model that can identify genes related to pollen germination rate through genetic analyses in many plants.

Flower stages, germination and viability of pollen grains of Annona squamosa L. in tropical conditions

The success in the application of artificial pollination techniques, aspects of floral biology should receive special attention, especially regarding the viability studies of pollen grains. In this sense, two experiments were carried out, aiming to determine the floral stages: floral opening (anthesis), female, male and senescence stages of A. squamosa L flowers under tropical climate conditions, and to evaluate the germination and viability of pollen grains submitted to different storage conditions. In the first experiment, observations and data collection began when flowers were still closed. Readings were taken every two hours for 46 hours until all the flowers reached the senescence stage. For the second, the experimental design was completely randomized, in a 2 x 7 factorial scheme, with 2 storage conditions (environment at 27 ± 2°C and refrigerated at 5 ± 2°C) and 7 pollen grain conditioning times (0, 4, 8, 12, 16, 20 and 24 hours), with 4 repetitions for the analysis of pollen grain germination and 3 repetitions for the analysis of pollen grain viability, and each repetition consisted of one blade. The variables evaluated were percentage of viable and non-viable pollen grains and percentage of germinated and non-germinated pollen grains. For tropical climatic conditions, the flower anthesis (female stage) of the sugar-apple begins at 00:00 h, extending until 12:00h on the same day. The flowers of the tree reach a functionally staminate stage (male stage) from 6 a.m. the next day. Pollen grains stored at room conditions (27 ± 2°C) remain viable for up to 24 hours after collection, showing that storage in a cold environment at low temperatures (5 ± 2°C) is not necessary. Seed pollen grains have a germination percentage of 51.25% when stored under ambient conditions (27 ± 2°C) for up to 4 hours after collection.

Maize Pollen Coat Xylanase Facilitates Pollen Tube Penetration into Silk during Sexual Reproduction

Cell wall hydrolases are well documented to be present on pollen, but their roles on the stigma during sexual reproduction have not been previously demonstrated. We explored the function of the tapetum-synthesized xylanase, ZmXYN1, on maize (Zea mays L.) pollen. Transgenic lines (xyl-less) containing little or no xylanase in the pollen coat were generated with use of an antisense construct of the xylanase gene-coding region driven by the XYN1 gene promoter. Xyl-less and wild-type plants had similar vegetative growth. Electron microscopy revealed no appreciable morphological difference in anther cells and pollen between xyl-less lines and the wild type, whereas immunofluorescence microscopy and biochemical analyses indicated an absence of xylanase on xyl-less pollen. Xyl-less pollen germinated as efficiently as wild-type pollen in vitro in a liquid medium but less so on gel media of increasing solidity or on silk, which is indicative of partial impaired water uptake. Once germinated in vitro or on silk, the xyl-less and wild-type pollen tubes elongated at comparable rates. Tubes of germinated xyl-less pollen on silk did not penetrate into the silk as efficiently as tubes of wild-type pollen, and this lower efficiency could be overcome by the addition of xylanase to the silk. For wild-type pollen, coat xylanase activity on oat spelled xylan in vitro and tube penetration into silk were inhibited by xylose but not glucose. The overall findings indicate that maize pollen coat xylanase facilitates pollen tube penetration into silk via enzymatic xylan hydrolysis.

FLOW CYTOMETRIC DETECTION OF UNREDUCED POLLEN IN BEGONIA

Unreduced gametes, formed due to a disruption in meiosis, are considered to play an important role in the formation of polyploids in nature. A screening method for the detection of unreduced gametes in ornamentals is however needed. Begonia was used as a model plant for the detection of unreduced pollen. Chemical, enzymatic, mechanical and osmotic assays have been used for the isolation of nuclei from Begonia pollen. The relative DNA content of pollen nuclei was measured using flow cytometry. Two methods, on germinated and non-germinated pollen, were successfully applied. Flow cytometric analysis showed two peaks in pollen, associated with the vegetative (1C) and generative (2C) nuclei of normal (reduced) pollen. Two of seven tested Begonia genotypes showed a peak at the 4C level, associated with the presence of unreduced gametes. Formation of unreduced gametes was associated with abnormalities in pollen shape or pollen size.

Suppression of heterotrimeric G-protein β-subunit affects anther shape, pollen development and inflorescence architecture in tobacco

The role of the heterotrimeric G-protein beta-subunit in plant development was studied in transgenic tobacco (Nicotiana tabacum L.) plants with reduced beta-subunit levels due to the antisense expression of the beta-subunit mRNA. The antisense plants had aberrant anther shape and produced non-germinating pollen. The anthers were sporadically transformed to petals, whereas other floral organs were not affected. The pollen grains were smaller than the wild-type pollen and had abnormal cell walls. The architecture of mature antisense plants was altered. The plants had long branched panicles and short stems. These data suggest that the beta-subunit of the plant heterotrimeric G-proteins is involved in the regulation of the reproductive phase of the tobacco life cycle, particularly in stamen development and pollen maturation.

Treatment of maize pollen to reduce nuclease activity

Recently it has been reported that maize (Zea mays L.) pollen can be stored up to several hours in a hypertonic aqueous medium at 0°C without losing its germinability (Broglia and Brunori 1994). We found that both release and activity of pollen nucleases are diminished in a cold hypertonic aqueous medium. Nucleases can be washed off while preserving germination ability and thus preserving the possibility of passive uptake of exogeneous DNA into germinating pollen grains. Alternatively, active DNA transfer into non-germinating pollen grains in the storage medium itself may be facilitated, due to the very reduced nuclease activity in this medium.

The localization of nonspecific esterase and cholinesterase activity in germinating pollen and in pollen tube ofVicia faba L. The effect of actinomycin D and cycloheximide

Two types of esterases, nonspecific esterase and cholin esterase have been distinguished in germinating pollen grains and in pollen tubes ofVicia faba using cytochemical methods. The localization of each of them was different. In the hydrated non-germinating pollen grain the nonspecific esterase was present in the cytoplasm and in the intine. The cholinesterase was localized mainly in the sexine and on the outside of the plasma membrane. A particularly large agglomeration of this enzyme was found in the aperture. During germination both types of extracellular esterases were released into the medium. In the pollen tube the activity of the enzymes studied was connected with the wall of its tip. The localization of both types of esterases in the germinating pollen grain and the growing tube were independent of actinomycin D. Cycloheximid prevented the cholinesterase localization at the pollen tube tip.

Dioecism of the genusCallicarpa (Verbenaceae) in the Bonin (Ogasawara) Islands

The morphology and function of the flowers of three species of the genusCallicarpa, C. glabra, C. nisnimurae andC. subpubescens, endemic to the Bonin Islands, revealed that all three species were dioecious. Male plants had short-styled flowers with sterile ovaries, while female plants had long-styled flowers with non-germinating inaperturate pollen grains. Such dioecism has never been reported from the family Verbenaceae. This curious sexual system with non-germinating pollen grains may have been maintained as a reward to insect pollinators.

Pollen Viability, Pollen Shedding, and Combining Ability for Tassel Heat Tolerance in Maize1

High temperatures during maize (Zea mays L.) pollination limit kernel number. Knowledge is limited on the effect of heat stress on both pollen viability and tassel shedding characteristics or of possible heat tolerance sources and testers. The purpose of this research was to determine the tassel response to heat stress, as measured by in vitro pollen viability and shedding characteristics, and the combining ability for these traits using a diallel of four heat‐tolerant and three heat‐intolerant inbreds. Tassels from field‐grown plants were excised at the beginning of anthesis, immediately placed in sufficient water to cover the base, and transferred to growth chambers. The growth chambers were maintained at light period temperatures of 29 and 38°C and dark period temperatures of 24 and 32°C, respectively. In vitro germination, anther emergence, and tassel color were determined after 24 h in the growth chamber. Due to bursting, low levels of germination were obtained, but the percentage of nongerminated pollen increased greatly at the high temperature (65% mean increase). The increase in nongerminated pollen varied among the genotypes, and all except B73 × Mo17 and N28 × Mo17 had statistically significant increases at the high temperature. Mo17 had the most desirable general combining ability (GCA) effect at the high temperature for nongerminated pollen. Specific combining ability effects were significant for nongerminated pollen and accounted for 51% of the genotype and genotype × temperature sum of squares. Anther emergence of the central tassel spike decreased differentially among the hybrids at the high temperature. Mo17 had the most desirable GCA effect for anther emergence. Anther emergence and pollen viability did not always respond the same to high temperature. Results from this study indicate pollen shed and viability must be considered in tassel heat tolerance and hybrids involving Mo17 had the most heat tolerant tassels.

Histochemical studies of alpha and beta-glucosidases in the germinating pollen grains ofPortulaca grandiflora

The paper deals with the distribution of alpha and beta-glucosidases in the germinating pollen grains ofPortulaca grandiflora. Both these enzymes are localized in the pollen wall and the cytoplasmic granules. The latter are distributed throughout the pollen cytoplasm, pollen tube and stigma hair. In non-germinating pollen grains, enzymes are concentrated in the pollen wall. Stigma hair sheaths are completely free from enzymes. The functional significance of these enzymes in the hydrolysis of phenolic glycosides and polysaccharides is discussed.

Histoenzymological compartmentation of β-glucuronidase in the germinating pollen grains ofPortulaca grandiflora

The contribution deals with the distribution of β-glucuronidase in the germinating pollen grains ofPortulaca grandiflora. In non-germinating pollen grains the enzyme is localized in the pollen wall; the cytoplasmic activity is subdued. With the initiation of germination, the activity of enzyme increases and the positive granules are richly packed in the pollen grains and pollen tubes. The stigma hairs also have such an activity. The functions of the enzyme in the metabolism of germinating pollen grains are discussed.

Properties of uridine diphosphoglucose dehydrogenase from pollen of Lilium longiflorum

UDP-glucose dehydrogenase was partially purified from germinating lily pollen ( Lilium longiflorum). The enzyme was associated with the particulate fraction in extracts of nongerminated pollen but could be solubilized when this fraction was treated with the detergent Tween 80. The enzyme was inhibited by UDP-galacturonic acid, UDP-glucuronic acid, and UDP-xylose. The latter was a powerful inhibitor when concentration of the substrate UDP-glucose was low and caused the normally hyperbolic UDP-glucose saturation curve to become sigmoid. These data were interpreted as indicating cooperative effects, UDP-glucose dehydrogenase being viewed as a regulatory enzyme.

Effect of pH on <i>in vitro</i> Pollen Germination of Fourteen Cultivated and Wild Species of Cucurbit

Context: The pH of the in vitro germination medium is the important factor controlling pollen germination and pollen tube development in different plant speciesObjectives: The aim of the present study was to assess the effect of pH levels of germination medium on the germinability 14 species of cucurbit.Materials and Methods: In vitro pollen germination of 14 cucurbit species was examined at five different levels of pH using Brewbaker and Kwack's medium. The pH of the medium was adjusted to 7.0, 7.5, 8.0, 8.5 and 9.0 by using a digital pH meter with the help of 0.1 N HCl and 0.1 N NaOH. In vitro pollen germination was tested using sitting drop culture technique. Five sitting drop cultures of pollen grains of five different pH for each species were raised on five different microslides. Then the slides were placed in a humidity chamber and incubated at 22 ± 2 °C for 1 to 2 hrs. All the cultures were fixed by adding a drop of a 1-2% acetocarmine before start scoring and the germinated and non-germinated pollen grains were scored using a microscope.Results: The pollen germination rate was higher in increased pH level in most of the cucurbit species studied and at 9.0 pH the maximum germination was observed in pumpkin (98%), snake gourd (98%), ridge gourd (96%), pointed gourd (89%), cucumber (98%), bitter gourd (96%), sweet gourd (98%) and sponge gourd (93%). The highest percentage of germination was observed in bottle gourd (98%) cucumber short (96%), watermelon (98%), muskmelon (97%) and ivy gourd (87%) at 8.5 pH level. The only cucurbit species wax gourd showed highest germination (97%) at the lower pH level of 7.5. Analysis of variance for the 14 cucurbits at five different pH levels showed that the differences between the germination rates of pollen grains of 14 cucurbit species at different pH level were highly significant (F=12.49, P<0.001, at 13/52 df). Conclusion: In most of the investigated cucurbit species pH level 8.5 to 9.0 was found optimum for the highest number of in vitro pollen germination, while wax gourd responded maximum pollen germination at pH 7.5.Key words: cucurbit; pollen; germination; pH.DOI: 10.3329/jbs.v17i0.7120J. bio-sci. 17: 129-133, 2009

Pollen Germination in Different Genotypes of Gladiolus

Fresh pollen of 16 promising genotypes of gladiolus was tested for their pollen viability. Modified cellophane method was employed to assess the pollen viability. Pollen germination media consisted of 15% sucrose supplemented with 300 ppm calcium nitrate, 200 ppm magnesium sulphate, 100 ppm potassium nitrate and 100 ppm boric acid. Highly significant variations were observed for percentage of pollen germination, non-germinated pollen and sterile pollen. The highest pollen germination (76.41%) was in genotype Hybrid selection 88-10-22, and did not differ significantly from Gladiolus callianthus (75.41%), Sapna (75.10 %), Hybrid selection 86-32-11, (73.28 %), Kum Kum (69.41%), Poonam (69.22 %), Hybrid selection 87-22-1 (67.87 %), Hybrid selection 87-1-1 (67.61 %), Psittacinus hybrid (64.64%) and Darshan (63.97%). The lowest non- germinated pollen (10.47%) was in genotype Gladiolus callianthus, and insignificantly followed by Hybrid selection 88-10-22 with 18.77 % and Hybrid selection 87-22-1 with 18.95 %. The lowest percentage of sterile pollen was noticed in genotype Sapna (2.82%) followed by Poonam with 4.00 % Hybrid selection 88-10-22 with 4.82% and Hybrid selection 82-11-27 with 5.22%. Key words: Gladiolus; germination; pollen DOI: 10.3126/njst.v11i0.4089Nepal Journal of Science and Technology 11 (2010) 47-50