Pollen Tube Penetration Research Articles

Marking live conifer pollen for long-distance dispersal experiments

Long-distance dispersal (LDD) theory requires a method for marking live LDD pollen. Such a method must complement more intensive sampling methods inclusive of molecular cytogenetics, proteomics and genomics. We developed a new method for marking live Pinus taeda pollen using two dyes, rhodamine 123 and aniline blue, dissolved in a sucrose solution. Marked and unmarked pollen were compared with respect to in vitro germination, storage, terminal velocity, and in vivo pollen tube penetration of ovules. We found that: (1) both types of marked pollen retained their capacity for germination, (2) both types of marked pollen had similar aerodynamic properties when compared to unmarked pollen controls, (3) marked pollen retained its germination capacity for 48 h, and (4) of the marked pollen, only the aniline-marked pollen penetrated ovules during pollination. Germination declined rapidly for both types of marked pollen after 48 h and before 37 days at -20°C storage, while unmarked pollen lots retained 93% germination at all stages. This method for marking live P. taeda pollen is feasible for tracing LDD pollen only if released and deposited within 48 h of dye treatment.

Sistema reprodutivo do Ipê-Branco: Tabebuia roseo-alba (Ridley) Sandwith (Bignoniaceae)

Estudos sobre sistemas reprodutivos têm indicado o predomínio da autoincompatibilidade de ação tardia (AIT) em Bignoniaceae, embora poucas espécies tenham sido investigadas e ocorram outros tipos de sistemas reprodutivos na família. O presente estudo objetivou determinar o sistema reprodutivo de T. roseo-alba através de experimentos de polinizações controladas, análise histológica dos eventos posteriores à polinização, verificação do desenvolvimento in situ dos tubos polínicos e testes de germinação de sementes. Apesar de os tubos polínicos penetrarem e fecundarem a maioria dos óvulos em pistilos autopolinizados, o aborto de 100% dos mesmos foi verificado e, embora sua abscisão tenha ocorrido entre o quarto e o sexto dia após o início da antese, observou-se um ligeiro crescimento dos óvulos e do ovário precedendo a abscisão, porém inferior ao crescimento nos pistilos submetidos à polinização cruzada. A endospermogênese inicial e a formação do tubo proembriônico também foram mais lentas nos pistilos autopolinizados. A longevidade dos pistilos autopolinizados foi maior que a de pistilos não polinizados, e a taxa de germinação de sementes foi de 93%, sendo todas as sementes monoembriônicas. Os resultados demonstram que T. roseo-alba é espécie auto-estéril, destituída de poliembrionia e que apresenta AIT pós-zigótica.

Pollen tube reuses intracellular components of nucellar cells undergoing programmed cell death in Pinus densiflora

Through the process known as programmed cell death (PCD), nucelli of Pinus densiflora serve as the transmitting tissue for growth of the pollen tube. We sought to clarify the processes of degradation of nucellar cell components and their transport to the pollen tube during PCD in response to pollen tube penetration of such nucelli. Stimulated by pollination, synthesis of large amounts of starch grains occurred in cells in a wide region of the nucellus, but as the pollen tube penetrated the nucellus, starch grains were degraded in amyloplasts of nucellar cells. In cells undergoing PCD, electron-dense vacuoles with high membrane contrast appeared, assumed a variety of autophagic structures, expanded, and ultimately collapsed and disappeared. Vesicles and electron-dense amorphous materials were released inside the thickened walls of cells undergoing PCD, and those vesicles and materials reaching the pollen tube after passing through the extracellular matrix were taken into the tube by endocytosis. These results show that in PCD of nucellar cells, intracellular materials are degraded in amyloplasts and vacuoles, and some of the degraded material is supplied to the pollen tube by vesicular transport to support tube growth.

Sistema reprodutivo e polinização de Senna multijuga (Fabaceae) em Mata Atlântica Montana

Resumo O conhecimento da biologia reprodutiva e polinização de árvores neotropicais vêm aumentando nas últimas décadas. No entanto, ainda é incipiente para a Mata Atlântica e muitas questões persistem, por exemplo, sobre a prevalência de autoincompatibilidade. A biologia floral, o sistema reprodutivo e os visitantes florais de Senna multijuga foram estudados, com o objetivo de verificar a ocorrência de autoincompatibilidade e determinar seus polinizadores. A espécie apresenta características florais típicas do gênero, como flores assimétricas, enantiostilia, heteranteria e deiscência poricida. A antese é diurna e a viabilidade do pólen não diferiu entre os morfos de anteras. Os polinizadores foram abelhas dos gêneros Bombus, Epicharis, Centris e Xylocopa e abelhas menores atuaram como pilhadores de pólen. A reprodução sexual em S. multijuga depende da transferência de pólen pelos polinizadores, pois não foram formados frutos por apomixia ou autopolinização. Este último resultado pode ser devido à depressão por endogamia ou a um mecanismo de autoincompatibilidade de ação tardia, uma vez que houve aborto uniforme de pistilos e penetração de tubos polínicos nos óvulos após autopolinização. Diferença na fecundidade após polinização manual e em condições naturais indica que a limitação polínica afeta o sucesso reprodutivo feminino, assim como a disponibilidade de recursos.

Pollen‐Pistil Interactions in North American and Chinese Cypripedium L. (Orchidaceae)

Fluorescence microscopy is used to compare frequencies of pollen tube penetration in in situ populations of Cypripedium bardolphianum W.W. Smith et Farrer, Cypripedium flavum W.W. Smith, Cypripedium montanum Dougl. ex Lindl., Cypripedium parviflorum Salisbury var. pubescens (Wildenow) O.W. Knight, Cypripedium reginae Walter, and Cypripedium tibeticum Schltr. The average natural (insect‐mediated) pollination rates measured over five seasons are wide ranging among the six species (0.08–0.74). However, the pollination rate of hand‐manipulated populations (self and/or cross) is significantly greater than the rate of insect‐mediated pollinations in all species studied. A few pollen tubes in both self‐ and cross‐pollinations display aberrant growth in the styles and/or ovaries, but their numbers are too small to suggest evidence of self‐incompatibility. Pollen tubes germinate and grow up to the bases of styles within 48 h in C. bardolphianum, C. flavum, and C. tibeticum. Pollen tubes remain at the bases of the ...

Evaluation of self-(in)compatibility in the almond (Prunus amygdalus batsch) genotype population from the Slankamen hill, Serbia

Due to the importance of obtaining almond cultivars adapted to the agroecological conditions of Serbia, in the period 2005-2006 pollen viability and self-(in)compatibility in 19 almond genotypes selected from the seedling population on Slankamen hill, were studied. All analyzed almond genotypes had good (50-70%) or high (over 70%) pollen germination. The study of self-(in)compatibility was done by monitoring of the fruit set in the field and observing self-pollen growth by fluorescence microscopy. Self-incompatibility was confirmed in all the 19 genotypes by both methods. Pollen tube penetration was stopped mostly at the upper third of the style of all genotypes, with characteristic irregularities.

Studies on the early development of zygotic and synergid embryo and endosperm in polyembryonic rice ApIII

A morphological, cytological and embryological investigation was conducted on the early development of embryo and endosperm in polyembryonic rice ( Oryza sativa L.) ApIII. We found that the percentage of single-, twin- and triple-seedlings from mature caryopses was 82.4–85.6%, 11.3–14.6% and 2.0–3.8%, respectively. From twin-seedlings one of the plants and in the triple-seedlings usually two seedlings were very weak having a mortality of about 50–60% and over 95%, respectively, and most of the dead seedlings were haploid. In addition, among 674 young caryopses the percentage of single-, double- and triple-embryo sacs was 77.60%, 18.10% and 4.30%, respectively. The development of both embryo and endosperm was observed in the embryo sacs of 653 young caryopses, while embryo development occurred in absence of endosperm in the embryo sacs of 21 ones. Furthermore, after pollen tube penetration, frequently one, rarely two, additional embryos arise via apogamy from the typical specialized synergid of normal egg apparatus with a collapsed cap-neck region that had a damaged filiform apparatus at the micropylar end. Finally, the formation of a strikingly degenerated endosperm cell region is a specialized functional structure that serves for the active transport of metabolites from endosperm to the developing embryo. Taken together, these results suggest that synergid apogamy seems to be the unique reason of polyembryony, poor growth and high mortality of many seedlings may be due to haploidy, and high genetic stability of polyembryony occurs in rice ApIII.

Class B β-expansins are needed for pollen separation and stigma penetration

Group 1 grass pollen allergens comprise a distinctive clade within the beta-expansin family of cell wall-loosening proteins and are divided by sequence divergence into two phylogenetically separable classes (A and B). They have been proposed to loosen the walls of the stigma and style. Supporting this idea, we recently showed that a transposon insertion in one of the maize group-1 allergen genes reduces the ability of pollen to effect fertilization under conditions of pollen competition. In this work, we provide additional information on the phenotype of this mutant, showing that pollen deficient in beta-expansin gene expression tended to form large aggregates, leading to poor pollen dispersal on anther dehiscence, and that emerging pollen tubes had difficulties entering the silk. In addition, a silencing construct was created to reduce expression of all the class B genes with results that are consistent with those seen with the transposon insertional line, including reduced transgene transmission through the pollen. Our results provide a more detailed understanding of the role of group 1 allergens (pollen beta-expansins) in maize pollen development, pollen dispersal, pollen tube penetration into the style, and pollen tube growth through the transmitting tract.

Development of the floral shoot and pre-anthesis cleistogamy inHydrobryopsis sessilis(Podostemaceae)

The reproductive biology of Hydrobryopsis sessilis (Podostemaceae, subfamily Podostemoideae), a reduced, threatened, aquatic angiosperm endemic to the Western Ghats of India, was examined. This is the first report on the transition from the vegetative to the reproductive phase in this plant, describing floral ontogeny, pollination and the breeding system. The cytohistological zonation of the apical meristem of the reproductive thallus is identical to that of the apical meristem of the vegetative thallus. The floral shoots do not replace vegetative shoots (i.e. the vegetative shoots never bear flowers), but form at new sites at the tip of the flattened plant body. Each floral shoot meristem is tiny, deep-seated and concave and arises endogenously following lysigeny. The floral shoot meristem gives rise to four to six bracts in a distichous manner. The development of spathe, stamens and carpels is described. The ab initio dorsiventrality of the carpels and the occurrence of endothelium in the ovules are reported. The mature stigmas and anthers lie close to each other. The pollen germinates within undehisced anthers and the pollen tubes enter the stigmas in the unopened floral bud, leading to pre-anthesis, complete, constitutional cleistogamy under water. The seed set is 63.2%. A significant finding is the penetration of several pollen tubes into the filaments of stamens in 16% of the flower buds, indicating a trend towards cryptic self-fertilization. The Indian Podostemoideae appear to show a shift from xenogamy or geitonogamy or autogamy in a chasmogamous flower to complete autogamy in a cleistogamous flower. The floral modifications leading to cleistogamy in H. sessilis have been identified.

Influences of Cross Pollination on Pollen Tube Growth and Fruit Set in Zuili Plums (Prunus salicina)

Zuili plum (Prunus salicina L.) trees usually set fruit poorly, although they produce high quality fruit. To elucidate the causes of the poor fruit set, pollen tube growth into pistils and fruit set percentage were investigated after cross-, self- and open-pollination. Ovule development in Zuili pistils was also investigated. Pollen tube penetration into the ovules via the obturator and micropyle was best when Zuili pistils were pollinated by cv. Black Amber (P. domestica) pollen grains, although cross-pollinations with Hongxinli and Miili (P. salicina) pollen were more effective than self- and open-pollination. The fruit set percentage was also highest in pistils pollinated with Black Amber pollen grains. Morphological observation of Zuili pistils revealed that the trees produce "double pistils", developing two ovaries from a basal pistil, at a rate as high as 28%. In such abnormal pistils, most ovules were lacking an embryo sac or were entirely degenerated. The percentage of normally developed ovules was 24.3% and 8.9% in normal and double pistils, respectively. From these results, we conclude that the main causes of poor fruit set of Zuili plums are a lack of effective cross-pollination and the production of high percentages of double pistils in which normally developed ovules are scarcely formed.

Plant cell separation and adhesion. Annual Plant Reviews, Volume 25.

Plant cell separation and adhesion. Annual Plant Reviews, Volume 25.

Recent Proliferation and Translocation of Pollen Group 1 Allergen Genes in the Maize Genome

The dominant allergenic components of grass pollen are known by immunologists as group 1 allergens. These constitute a set of closely related proteins from the beta-expansin family and have been shown to have cell wall-loosening activity. Group 1 allergens may facilitate the penetration of pollen tubes through the grass stigma and style. In maize (Zea mays), group 1 allergens are divided into two classes, A and B. We have identified 15 genes encoding group 1 allergens in maize, 11 genes in class A and four genes in class B, as well as seven pseudogenes. The genes in class A can be divided by sequence relatedness into two complexes, whereas the genes in class B constitute a single complex. Most of the genes identified are represented in pollen-specific expressed sequence tag libraries and are under purifying selection, despite the presence of multiple copies that are nearly identical. Group 1 allergen genes are clustered in at least six different genomic locations. The single class B location and one of the class A locations show synteny with the rice (Oryza sativa) regions where orthologous genes are found. Both classes are expressed at high levels in mature pollen but at low levels in immature flowers. The set of genes encoding maize group 1 allergens is more complex than originally anticipated. If this situation is common in grasses, it may account for the large number of protein variants, or group 1 isoallergens, identified previously in turf grass pollen by immunologists.

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.

Pollen Tube Growth in Cross Combinations between Torenia fournieri and Fourteen Related Species

The precise guidance of pollen tubes is important for successful fertilization. Based on abnormalities in interspecific and intergeneric crosses, we investigated pollen tube guidance in Torenia fournieri, which is a model plant for studies on plant fertilization due to its protruding embryo sac. Critical abnormalities of pollen tube growth have never been recorded in interspecific crosses between either T. baillonii or T. concolor, and hybrids could be obtained. In contrast, abnormalities appeared in intergeneric crosses with twelve other species. The abnormalities could be observed for five phases of pollen tube guidance revealed in Arabidopsis mutants as follows: germination of the pollen grains, penetration of pollen tube through stigma cells, growth in the transmitting tract, arrival at the placenta and release of sperm into the micropyle. In the self-pollination of T. fournieri, in vivo, the pollen tubes grew straight from the placenta to the protruding embryo sac, omitting funiculus guidance. In addition, observation of an intergeneric cross between T. fournieri (female) and Mimulus hybridas revealed that 64% of the pollen tubes grew straight to the micropyles of ovules rather than to synergid cells, suggesting that the micropyle in T. fournieri transmits the attraction signal. In addition, the signal attracted pollen tubes of M. hybridas to the micropyle, unlike those of T. fournieri.

Effective pollination period in durian (Durio zibethinus Murr.) and the factors regulating it

The effective pollination period (EPP) of durian was determined by both delayed and bud pollination, during which reproductive factors affecting the EPP, e.g., stigma receptivity, pollen tube growth in the style, and ovule longevity were studied histologically. This study was conducted in three distinct locations in Thailand, namely, the Chantaburi Horticultural Research Center and two private orchards in Chantaburi and Trat provinces. Results from artificial pollination revealed that at anthesis, the durian flower is receptive and has a high fruit set ratio. A mean fruit set of 50% was obtained at anthesis in the private orchard in Chantaburi province. However, the EPP of durian was found to be very short, lasting for only one night; the fruit set from pollination on the morning after anthesis ranged from 0% to 3.4%. No fruit set occurred following pollination 24 or more hours after anthesis. When compared with the flowers of other fruit species, the durian flower has a unique feature in that it blooms overnight; the following morning, there is abscission of all parts of the flower, except the gynoecium. Thus, EPP appears to be synchronized with flower longevity. On the other hand, the durian flower was receptive several hours before anthesis. The results of chemical tests, including the hydrogen peroxide test and Perex-Test®, for the evaluation of stigma receptivity appeared to be in agreement with the EPP. However, fluorescent microscopy revealed that pollen could germinate even in the stigmas pollinated 48 h after anthesis, but the number of pollen tubes at the top of the style rapidly decreased from 34.6 (at anthesis) to 0.5 (48 h after anthesis). A correlation test demonstrated a higher correlation coefficient between the fruit set and the number of pollen than that between the fruit set and the result of Perex-Test®. This indicated that pollen tube penetration or elongation in the style was inhibited, probably due to the deterioration of nutritional support from the pistil to the pollen tubes; this can be a limiting factor of the EPP in durian.

The Effect of Temperature on Pollen Germination, Pollen Tube Growth, and Stigmatic Receptivity in Peach

Temperature is a major climatic factor that limits geographical distribution of plant species, and the reproductive phase has proven to be one of the most temperature-vulnerable stages. Here, we have used peach to evaluate the effect of temperature on some processes of the progamic phase, from pollination to the arrival of pollen tubes in the ovary. Within the range of temperatures studied, 20 degrees C in the laboratory and, on average, 5.7 degrees C in the field, the results show an accelerating effect of increasing temperature on pollen germination and pollen tube growth kinetics, as well as an increase in the number of pollen tubes that reach the style base. For the last two parameters, although the range of temperature registered in the field was much lower, the results obtained in the laboratory paralleled those obtained in the field. Increasing temperatures drastically reduced stigmatic receptivity. Reduction was sequential, with stigmas first losing the capacity to sustain pollen tube penetration to the transmitting tissue, then their capacity to offer support for pollen germination and, finally, their capacity to support pollen grain adhesion. Within a species-specific range of temperature, this apparent opposite effect of temperature on the male and female side could provide plants with the plasticity to withstand changing environmental effects, ensuring a good level of fertilization.

IMPROVEMENT OF BREEDING EFFICIENCY FOR INTERSPECIFIC HYBRIDIZATION OF LILIES IN KOREA

Interspecific hybridization of lily in Korea is focused on increasing breeding efficiency especially under Korean climate conditions. Cultivars derived from L. formolongi were propagated by seed and were sensitive to day length. These cultivars differ from flowering time with the Asiatic and Oriental hybrids. Therefore, to breed interspecific hybrids by using L. formolongi, the use of stored viable pollen is a prerequisite. Lily pollen was stored at four different temperature conditions for one year and pollen viability was investigated using three different methods. As storage temperatures increased, pollen viability was drastically reduced. Viability of lily pollen stored was maintained effectively at temperatures lower than -20°C. FDA test was the most effective and convenient method to ascertain pollen viability. To overcome pre-fertilization barriers in interspecific hybridization of lilies, pollen tube growth in the stigma or style, and the penetration of pollen tubes into the ovules were observed. OA (Oriental Asiatic), OL (Oriental Longiflorum), FA (Formolongi Asiatic), and FO (Formolongi Oriental) hybrids were obtained through embryo rescue and stigmatic and CSM pollination, while AO (Asiatic Oriental), AL (Asiatic Longiflorum), LA (Longiflorum Asiatic) and LO (Longiflorum Oriental) hybrids were not obtained through these methods. Backcrosses were used in order to reduce scattered petals in interspecific hybrids. It was possible to obtain BC107 hybrids between FA (Fomolongi Asiatic) and Asiatic hybrids as male parent. But, it was difficult to obtain BC1 hybrids between FA hybrid and L. formolongi as female parent. Normal pollination was better than CSM (Cut style pollination method) to get BC1 hybrids. In order to overcome the sterility of interspecific hybrids pollen, low concentration of oryzalin was more effective than high concentration of colchicine to obtain tetraploid. The fertility of pollen increased by about 40% in tetraploid level. Fusarium oxysporum screening test was carried out. ‘Hae-wool’, ‘Sinavro’ and FA 96-16 were classified as highly disease resistant and ‘Supia’ and ‘Doran’ as moderately resistant to Fusarium bulb-rot. The FO-hybrid lilies from NHRI were susceptible.

Paternal cytoplasmic transmission in Chinese pine (Pinus tabulaeformis)

In this paper, the stages of normal sexual reproduction between pollen tube penetration of the archegonium and early embryo formation in Pinus tabulaeformis are described, emphasizing the transmission of parental cytoplasm, especially the DNA-containing organelles--plastids and mitochondria. The pollen tube growing in the nucellus contained an irregular tube nucleus followed by a pair of sperm cells. The tube cytoplasm contained abundant organelles, including starch-containing plastids and mitochondria. The two sperm cells differed in their volume of cytoplasm. The leading sperm, with more cytoplasm, contained abundant plastids and mitochondria, while the trailing one, with a thin layer of cytoplasm, had very few organelles. The mature egg cell contained a great number of mitochondria, whereas it lacked normal plastids. At fertilization, the pollen tube penetrated into the egg cell at the micropylar end and released all of its contents, including the two sperms. One of the sperm nuclei fused with the egg nucleus, whereas the other one was retained by the receptive vacuole. Very few plastids and mitochondria of male origin were observed around the fusing sperm and egg nuclei, while the retained sperm nucleus was surrounded by a large amount of male cytoplasm. The discharged tube cytoplasm occupied a large micropylar area in the egg cell. In the free nuclear proembryo, organelles of maternal and paternal origins intermingled in the neocytoplasm around the free nuclei. Most of the mitochondria had the same features as those of the egg cell, but some appeared to be from sperm cells and tube cytoplasm. Plastids were obviously of male origin, with an appearance similar to those of the sperm or tube cells. After cellularization of the proembryo, maternal mitochondria became more abundant than the paternal ones and the plastids enlarged and began to accumulate starch. The results reveal the cytological mechanism for paternal inheritance of plastids and biparental inheritance of mitochondria in Chinese pine.

Oleosin-like proteins are not present on the surface of tobacco pollen

Pollen-stigma interactions on wet- and dry-type stigmas involve similar processes: the hydration of the pollen, followed by pollen tube growth and penetration of the stigma. Furthermore, in some species, identical molecules, namely lipids, are used to achieve this. In addition to lipids, oleosin-like proteins of the pollen coat of dry-type stigma plants have been shown to be involved in pollen-stigma interactions. However, little information is present about the proteins on the surface of pollen of wet-type stigma plants, in particular that of the Solanaceae. To analyze proteins from the surface of pollen of Nicotiana tabacum (tobacco), a solanaceous plant, we used an antiserum raised against Brassica pollen coat, a dry-type stigma plant of the Brassicaceae. In addition we used a molecular approach to identify tobacco homologues of oleosin-like genes. Our results show that no proteins similar to Brassica oleracea pollen coat proteins are present on the surface of tobacco pollen, and that oleosin-like genes are not expressed in tobacco anthers or stigmas.

Recent Progresses on Self-incompatibility Research in Brassica Species

Many hermaphrodite plant species have evolved mechanisms to prevent self-fertilization. One such mechanism is self-incompatibility (SI), which is defined as the inability of a fertile hermaphrodite plant to produce zygotes after self-pollination. SI prevents self-fertilization by rejecting pollen from plants with the same genotype. The SI system in Brassica is controlled sporophytically by multiple alleles at a single locus, designated as S, and involves cell-cell communication between male and female. When the S phenotype of the pollen is the same as that of the stigma, pollen germination and/or pollen tube penetration are disturbed on the papilla cells. On the female side, two genes (SLG and SRK) located at the S locus, are involved in the SI reaction. SLG is a secreted glycoprotein expressed abundantly in the papilla cell, and SRK is a membrane-spanning receptor-like serine/threonine kinase whose extracellular domain is highly similar to SLG. Gain-of-function experiments have demonstrated that SRK solely determines S haplotype-specificity of the stigma, while SLG enhances the recognition reaction of SI. The sequence analysis of the S locus genomic region of Brassica campestris (syn. rapa) has led to the identification of an anther-specific gene, designated as SP11, which encodes a small cysteine-rich basic protein. Pollination bioassay and gain-of-function experiments have indicated that SP11 is the male S determinant. When the sequence of SP11 was aligned, six cysteine residues were found to be completely conserved among alleles. These conserved cysteine residues could be important for the tertiary structure of SP11. Recent biochemical analysis has suggested that SP11 operates as a sole ligand to activate its cognate SRK specifically. Because the activity of the S allele is controlled sporophytically, dominance relationships influence the ultimate phenotype of both the stigma and pollen. Molecular analysis has demonstrated that the dominance relationships between S alleles in the stigma were determined by SRK itself, but not by the relative expression level. In contrast, in the pollen, the expression of SP11 from the recessive S allele was specifically suppressed in the S heterozygote, suggesting that the dominance relationships in pollen were determined by the expression level of SP11.