Self-pollen Tubes Research Articles

Relative success of self and outcross pollen after mixed- and single-donor pollinations in Eucalyptus grandis

A previous observation that self-pollen tubes traversed the style at a slower rate than cross-pollen tubes in Eucalyptus grandis and E. urophylla suggested the presence of cryptic self-incompatibility (CSI) in these species. The aim of the present study was, with the help of molecular markers, to examine the siring ability of self- and cross-pollen in a single clone of E. grandis, after both mixed- and single-donor pollinations, in order to confirm the presence of CSI. Single-donor cross-pollinations set a significantly higher number of seeds per flower pollinated compared to those performed with self-pollen, while there were no significant differences between the open control and single-donor self-pollinations. Molecular markers revealed that 100% of the progeny from mixed-donor pollinations were outcrossed, confirming the competitive advantage of cross-pollen. In addition, there was a significant change in the self:outcross seed ratio between single- and mixed-donor pollinations, suggesting that the observed deficit of selfed seeds in mixed-donor fruits could be the result of differential pollen tube growth. From the extremely low seed yields following single-donor self-pollinations, it is clear that an additional incompatibility mechanism is operating in E. grandis, and this is suggested to be late-acting self-incompatibility, acting before fertilisation.

CgSL2, an S-like RNase gene in ‘Zigui shatian’ pummelo (Citrus grandis Osbeck), is involved in ovary senescence

'Zigui shatian' pummelo (Citrus grandis Osbeck) is one nature mutant from 'Shatian' pummelo, which showed self-compatibility, because self-pollen tubes were not arrested in the style, moreover abnormal post-zygotic development in ovary caused seed abortion in the cultivar. Herein we constructed a cDNA library from flowers of 'Zigui shatian' pummelo and identified one RNase gene fragment. The full length of cDNA sequence of this gene, with an open reading frame of 834bp, was isolated by 5'-RACE method. The gene, named as CgSL2, contained five conserved regions and two histidine residues essential for RNase activity. Phylogenetic analysis indicated that CgSL2 was mostly similar to AhSL28, an S-like RNase from Antirrhinum. Southern hybridization verified CgSL2 existed in the genome as multiple copies. qRT-PCR and RT-PCR analysis showed that the expression of CgSL2 was not tissue-specific. The expression of CgSL2 was down-regulated during senescence of stem, petal, style and stamen, whereas up-regulated during ovary senescence. Further in situ hybridization of CgSL2 in the ovary during the balloon stage to anthesis stage also showed that it dramatically increased in mature flower, consistent with qRT-PCR and RT-PCR results. These findings suggested that CgSL2 might play an important role during ovary senescence.

The Skp1-like protein SSK1 is required for cross-pollen compatibility inS-RNase-based self-incompatibility

The self-incompatibility (SI) response occurs widely in flowering plants as a means of preventing self-fertilization. In these self/non-self discrimination systems, plant pistils reject self or genetically related pollen. In the Solanaceae, Plantaginaceae and Rosaceae, pistil-secreted S-RNases enter the pollen tube and function as cytotoxins to specifically arrest self-pollen tube growth. Recent studies have revealed that the S-locus F-box (SLF) protein controls the pollen expression of SI in these families. However, the precise role of SLF remains largely unknown. Here we report that PhSSK1 (Petunia hybrida SLF-interacting Skp1-like1), an equivalent of AhSSK1 of Antirrhinum hispanicum, is expressed specifically in pollen and acts as an adaptor in an SCF(Skp1-Cullin1-F-box)(SLF) complex, indicating that this pollen-specific SSK1-SLF interaction occurs in both Petunia and Antirrhinum, two species from the Solanaceae and Plantaginaceae, respectively. Substantial reduction of PhSSK1 in pollen reduced cross-pollen compatibility (CPC) in the S-RNase-based SI response, suggesting that the pollen S determinant contributes to inhibiting rather than protecting the S-RNase activity, at least in solanaceous plants. Furthermore, our results provide an example that a specific Skp1-like protein other than the known conserved ones can be recruited into a canonical SCF complex as an adaptor.

Reproductive biology of Melaleuca alternifolia (Myrtaceae) 2. Incompatibility and pollen transfer in relation to the breeding system

The onset of stigma receptivity in Melaleuca alternifolia (Maiden & Betche) Cheel was evaluated by observing pollen-tube growth and seed set following controlled pollination. Pollen-tube numbers in the style, following controlled pollinations, increased from Day 1 to Day 6, then declining rapidly. The stigma was most receptive during Days 3–6, and still receptive at low levels as early as shortly after anthesis and as late as 10 days after pollination. The present study found that individuals of M. alternifolia differed in their degree of expression of self-incompatibility. Artificial self-pollination, with emasculation, in several families resulted in complete self-incompatibility, with no capsule retention. The microscopic observation of pollen-tube development revealed a mechanism of self-incompatibility in M. alternifolia. A self-incompatibility system operates in the style, although a few self-pollen grains are capable of germinating and producing pollen tubes. It also appears that late-acting self-incompatibility mechanisms discriminate against self-pollen tubes when they descend to the ovary. Artificial cross-pollination of selected parents produced seed with greater germination capacity and seedlings that grew faster than the corresponding open-pollinated seed and seedlings from the same parent. Freeze-dried pollen stored at −18°C maintained viability (22%) over 1 year of storage. This finding will allow greater flexibility in undertaking controlled pollinations, because stored pollen can be substituted for fresh pollen when insufficient quantities are available from new-season flowers. A wide variety of insects was observed visiting the flowers of M. alternifolia, and capsule set was high even in bags that excluded flower visitors greater than 2 mm. Thrips species seem likely to be important pollinators of this species because they are small and were abundant inside and outside of exclusion bags, although several other insect species such as bees, flies and wasps were also identified as frequent floral visitors.

Self‐sterility in two Cytisus species (Leguminosae, Papilionoideae) due to early‐acting inbreeding depression

In most angiosperms, the endosperm develops before the embryo, but with harmony between the two structures until final seed formation. In an embryological study, we show that inbreeding depression causes disharmony in development of the two structures in two Leguminosae shrubs, Cytisus multiflorus and C. striatus. Our main objective was to test the causes of self-sterility in the two species by comparing the embryological development of the self seeds with that of cross seeds. In developing selfed seeds of C. multiflorus, the embryo reaches at most the globular stage and never forms mature seeds, while in C. striatus a few mature selfed seeds are formed. In both species, the main cause of abortion of developing selfed seeds is diminished endosperm development (low values of the ratio of endosperm to embryo), which triggers collapse of the endosperm and embryo. The results indicate that self-sterility in C. striatus is postzygotic because of strong, early inbreeding depression, while in C. multiflorus there exists a mixed pre- and postzygotic mechanism; the prezygotic mechanism causes rejection of some self-pollen tubes in the style/ovary, and the early inbreeding depression triggers abortion of fertilized ovules that escaped that action.

Biologia reprodutiva da "catingueira" (Caesalpinia pyramidalis Tul., Leguminosae-Caesalpinioideae), uma espécie endêmica da Caatinga

The Leguminosae comprises 293 species in the Caatinga, and only eight were studied for pollination and, or breeding system. We analysed the floral biology, pollinators and breeding system of Caesalpinia pyramidalis Tul. The pollen tube growth was observed eight, 24 and 48 hours after hand pollinations. There were 5.7 ± 0.9 ovules/flower, 66.9 ± 47.8 flowers and 2.1 ± 1.2 fruits per inflorescence and 2.88 ± 1.44 seeds/fruit. Nectar production was about 1.0 µL during the first day of anthesis, 0.5 µL in the second day, and there was no production in the third day. C. pyramidalis is self-incompatible but self-pollen tubes grew down to the embryo sac. Species of Xylocopa and Centris are important pollinators of C. pyramidalis. During the visits, the bees promote mostly geitonogamy which favours the loss of fruits, and generates a low fruit/flower ratio (0.03). However, high seed/ovule ratio (0.50) shows highest investment in seeds by cross-pollination, reducing the effects of geitonogamy.

Comparison of S-RNase, RNase T 1, T 2, and A effects on growth inhibition and RNA degradation of in vitro-cultured pear pollen

To obtain the basic information on fruit set regulation, effects of several RNases including S-RNase on pollen tube growth and RNA degradation in the tube were studied in the pear. Purified S-RNase from the Japanese pear ‘Kosui’ (S 4S 5) predominantly inhibited the growth of ‘Kosui’ pollen tubes (self) in vitro at 0.28 unit μL −1, but it inhibited ‘Chojuro’ (S 2S 3) pollen (cross) only slightly. The same unit of RNase T 1 (EC 3.1.27.3) clearly inhibited the pollen tube growth, but the action was significantly weaker than that of the S-RNase against the self-pollen. Inhibitory effect of RNase T 2 (EC 3.1.27.1) and RNase A (EC 3.1.27.5) was only slight. The proteins other than the S-RNase extracted from pear style did not have any inhibitory action, though they possessed RNase activity 3.8 times higher than S-RNase. Thus, RNases tested here could not substitute for the S-RNase in specific inhibition against the self-pollen tube growth. Total RNA degradation by each RNase occurred in the pollen tubes as following order; S-RNase (self) ≥T 1 > T 2 ≥ A > S-RNase (cross). Degradation degree of 28S and 18S rRNA was as follows; S-RNase (self) > A > T 1 > T 2 > S-RNase (cross). The degradation of 5.8S and 5S rRNA was; S-RNase (self) > S-RNase (cross) > A > T 2 > T 1. The degree of rRNA degradation was, thus, not always in parallel with the degree of pollen growth inhibition. The S-RNase may degrade not only rRNA but also mRNA essential for pollen tube growth, and may be specifically adapted to inhibit the growth of self-pollen tubes. Therefore, controlling S-RNase amount in the style will produce self-thinning cultivars efficiently, which are unnecessary not only for hand-pollination but fruit-thinning practices in the pear. Practically, cultivar with weak self-incompatibility and small amount of S-RNase, such as ‘Okusankichi’, may be an expecting candidate for breeding self-thinning cultivars.

Treatment of isolated pistils with protease inhibitors overcomes the self-incompatibility response in buckwheat

Isolated pistils of distylous buckwheat (Fagopyrum esculentum Moench) were treated with protease inhibitors (PMSF, pepstatin A, and antipain). Pistils were cross- or self- pollinated, and growth of pollen tubes was observed under a fluorescence microscope. Treatments with all inhibitors suppressed inhibition of self-pollen tube growth, suggesting that activity of proteases is involved in rejection of self-pollen during the SI response.

(239) Evaluation of Self-compatibility in Clethra alnifolia

Clethra alnifolia, which is commonly known as summersweet, is an attractive deciduous shrub that produces fragrant flower in mid-summer. Breeding efforts are hampered by a lack of information on the reproductive behavior of this native species. The objective of this study was to evaluate self-compatibility in C. alnifolia. Pollen germination and pollen tube growth in styles were examined following self- and cross-pollinations using fluorescence microscopy. Seed set and germination were compared following self- and cross-pollinations. While self-pollen tubes appeared to grow slightly slower than cross-pollen tubes, there was no indication of a self-incompatibility system acting at the stigmatic or stylar level in C. alnifolia. Self-pollinations of `Hummingbird' and `Ruby Spice' produced fewer seeds than did cross-pollinations of these cultivars. Germination of all seed obtained from this study was too poor to allow a comparison of germination rates of the self- and cross-pollinated seed. However, because a few self-progeny were obtained, emasculation is recommended when making controlled pollinations. The presence of a late-acting self-incompatibility system or early acting inbreeding depression was proposed as being responsible for the lower seed set following self-pollination.

Reproductive Biology of Clethra alnifolia

Breeding efforts in Clethra alnifolia L., an ornamental shrub native to the Eastern U.S., are hindered by a lack of information on the reproductive behavior of this species. The objective of this study was to evaluate self-compatibility, time of stigma receptivity, and the relationship between time of pollen shed and stigma receptivity in C. alnifolia. Stigma receptivity and changes in floral morphology were monitored over a 7-day period beginning at flower opening. Pollen germination and pollen tube growth in styles were examined following self- and cross-pollinations using fluorescence microscopy. Seed set and germination were compared following self- and cross-pollinations. Anthers began to dehisce in `Hummingbird' and `Ruby Spice' the day after flowers opened, but stigmas did not become fully receptive to pollen until 2 days later. An increase in the length of pistils was observed following flower opening. Maximum elongation of pistils occurred at approximately the same time stigmas became receptive and could be utilized as an indicator of receptivity. While self-pollen tubes appeared to grow slightly slower than cross-pollen tubes, there was no indication of a self-incompatibility system acting at the stigmatic or stylar level in C. alnifolia. Self-pollinations of `Hummingbird' and `Ruby Spice' produced fewer seeds than did cross-pollinations of these cultivars. Germination of all seed obtained from this study was too poor to allow a comparison of germination rates of the self- and cross-pollinated seed. However, because a few self-progeny were obtained, emasculation is recommended when making controlled pollinations. The presence of a late acting self-incompatibility system or early-acting inbreeding depression was proposed as being responsible for the lower seed set following self-pollination.

Recent advances in identification of male specificity determinant and its function in S-RNase-mediated gametophytic self-incompatibility

S-RNase-mediated gametophytic self-incompatibility (GSI) is controlled by a multillalelic S-locus at which two separate genes, the female (pistil) and male (pollen) specificity determinants, are tightly linked. This review described both the identification of pollen specific F-box genes, SLF/SFBs, in Antirrhinum, Petunia and Prunus species and the demonstration of SLF/SFB as pollen determinant together with their functions in GSI response. Recent studies of how the pollen determinant functions in pollination reaction revealed that pollen determinant interacted with S-RNases in a non-allele-specific manner. It targeted all of the non-self S-RNases for ubiquitination through a functional SCF complex and subsequent degradation via 26S proteasome pathway in compatible reaction. It allows pollen tube to reach into the embryo sac and to finish double fertilization. In incompatible response, the intact self S-RNases were left to function as a cytotoxin that degrades self-pollen tube RNA, resulting in the cessation of pollen tube growth.

Variation in the self‐incompatibility response within and among populations of the tropical shrub Witheringia solanacea (Solanaceae)

Breakdown of genetically enforced self-incompatibility (SI), an extremely common and important evolutionary transition in plants, has conventionally been conceived as a qualitative rather than a quantitative change. We evaluated qualitative and quantitative variation in SI for four populations of Witheringia solanacea in Costa Rica, examining growth of self-pollen tubes in pollinations of buds and mature flowers. We also measured levels of RNase production in styles to determine whether enzyme production was correlated with differences in self-rejection. The two small populations contained both self-compatible (SC) individuals and obligate outcrossers (female or SI). Plants in the two large populations were uniformly SI as revealed by pollen tube growth, although several of these individuals sporadically set seed autogamously. Stylar RNase activity did not differ significantly between bud and mature flowers, but self-pollen tube growth did differ, suggesting that a gene product in addition to S-RNase is responsible for developmental onset of SI. Population-level differences in RNase activity were consistent with differences in the strength of the rejection response in bud pollinations, suggesting that a threshold level of S-RNase, in combination with other factors, is necessary for SI. Our results support a growing body of evidence that not only qualitative variation in SI, but also quantitative variation may be functionally significant.

Self‐sterility in Ipomopsis aggregata (Polemoniaceae) is due to prezygotic ovule degeneration

Based on previous studies, extreme (>99%) self-sterility in scarlet gilia (Ipomopsis aggregata) appears to be involved in late-acting ovarian self-incompatibility (OSI). Here, we confirm this suggestion by comparing structural events that follow from cross- vs. self-pollinations of I. aggregata. Growth of cross- and self-pollen tubes in the style at 11 h and growth in the ovary at 24 h was equivalent. Nonetheless, by 24 h, cross-pollen effected a significantly higher percentage of both ovule penetration and fertilization. Ovules in self-pollinated flowers showed pronounced changes, including an absence of embryo sac expansion and reduced starch in the integument, by 11 h post-pollination, well before pollen tube entry into the ovary. In addition, the integumentary tapetum and adjacent 1-3 cell layers exhibited abnormal cell division, pronounced deposition of thick, pectin-rich cell walls, and cellular collapse. Ovules and embryo sacs from cross-pollinated flowers rarely showed such features. Developmental changes in ovules from self-pollinated flowers eventually resulted in integument and embryo sac collapse, a process not observed in ovules of unpollinated flowers. We suggest that OSI involves long-distance signaling between self-pollen or self-pollen tubes and carpel tissue that reduces availability of receptive ovules for fertilization before pollen tubes arrive in the ovary.

STIGMATIC RECEPTIVITY AND POLLEN TUBE GROWTH IN RABBITEYE BLUEBERRY

Effective pollination period (EPP) is the number of days during which pollination is effective to produce a fruit. The EPP is determined by ovule longevity, pollen tube growth rate and length of stigmatic receptivity. The objectives of this research were to establish the EPP of rabbiteye blueberry and to further the understanding of its limiting parameters. The experiments were conducted in growth chambers using blueberry plants of the cultivars Brightwell and Tifblue. Emasculated flowers were hand-pollinated at 0, 2, 4, 6, and 8 days after anthesis (DAA). Ripe fruit were harvested to record percentage fruit set. Stigmatic receptivity was evaluated as the number of germinated tetrads on the stigma 24 hours after pollination. Under day/night temperatures of 23/10 °C, the EPP was 7 days. Stigmatic receptivity was lowest on the day of anthesis and increased as flowers aged. Stigmatic receptivity was not positively correlated to fruit set, therefore, this parameter was not the most limiting factor of the EPP. Observations of pistils pollinated 3 DAA indicated that the fastest growing pollen tubes reached the bottom of the style 2 to 3 days after pollination. Self-pollination resulted in normal pollen tube growth in the style and inside the ovary. Self-pollen tubes were seen penetrating the micropile.

Reproductive biology of Boswellia serrata, the source of salai guggul, an important gum-resin

Detailed studies were carried out on the phenology, floral biology, pollination ecology and breeding system of Boswellia serrata Roxb. (Burseraceae) the source of ‘salai guggul’. The trees remain leafless during the entire period of flowering and fruiting. The inflorescence is a terminal raceme and produces up to 90 bisexual, actinomorphic flowers. On average a flower produces 10 044 ± 1259 starch-filled pollen grains. About 85% of the fresh pollen grains are viable; the pollen to ovule ratio is 3348 : 1. The stigma is of the wet papillate type. The style is hollow with three flattened stylar canals filled with a secretion product. The stylar canals are bordered by a layer of glandular canal cells. The inner tangential wall of the canal cells shows cellulose thickenings. The ovary is trilocular and bears three ovules, one in each locule. Flowers offer nectar and pollen as rewards to floral visitors. The giant Asian honey bee (Apis dorsata) and A. cerana var. indica(Indian honey bee) are the effective pollinators. The species is self-incompatible and the selfed pollen tubes are inhibited soon after their entry into the stigma. Self-pollen tubes develop a characteristic ‘isthmus’ as a result of enlargement of the tube soon after emergence through the narrow germ pore. Cross-pollinated flowers allowed normal pollen germination and pollen tube growth, and resulted in fruit- and seed-set. Under open pollination fruit-set was only about 10%. Although manual cross-pollinations increased fruit set, it was only up to about 20%. Low fruit set appears to be the result of inadequate cross-pollination and other constraints, presumably limitation of available nutrients. © 2005 The Linnean Society of London, Botanical Journal of the Linnean Society, 2005, 147, 73–82.

Outcrossing in interspecific hybrids between Eucalyptus spathulata and E. platypus

Outcrossing was investigated in interspecific hybrids between self-fertile Eucalyptus platypus Blakely and partially self-sterile E. spathulata Hook., which shows both pre- and post-zygotic timing. Four hybrid trees were used for the study, two with E. spathulata and two with E. platypus as female parent. Each hybrid had a similar number of locules to each other and to the E. platypus parent, and an intermediate number of ovules per flower compared with the parent species. Controlled hand-pollinations were carried out, in which both self- and cross-pollen from the other hybrid tree with the same female parent species was applied to flowers on each of the four trees, and observations were made 10days, 4weeks and 8weeks after pollination and at seed maturity. In all hybrids, mean seeds per capsule was consistently higher following cross-pollination than following self-pollination. All hybrids showed a reduction in pollen tube number between the top and base of the style when examined by fluorescence microscopy. One tree had significantly fewer cross- than self-pollen tubes at the base of the style, but a similar number of ovules was penetrated by pollen tubes following both treatments. In the other three, there was no difference between cross- and self-pollination in pollen tubes in the style. In three of the four trees there was no difference in ovule penetration following self- or cross-pollination, but in the other, more crossed than selfed ovules were penetrated. Light-microscopy observation of ovules indicated that ovule abortion following fertilisation accounted for the reduced numbers of seeds following self-pollination and to a lesser extent following cross-pollination. All four hybrid trees, irrespective of female parent, were partially self-sterile and resembled the partially self-sterile E. spathulata rather than the self-fertile E. platypus. While the timing of outcrossing control of E. spathulata was both pre- and post-zygotic, only one hybrid was similar, with the other three showing post-zygotic control.

The RHV region of S-RNase in the European pear (Pyrus communis) is not required for the determination of specific pollen rejection

In the gametophytic self-incompatibility system, growth of self-pollen tubes in the style is inhibited in a haplotype-specific manner by S-RNase. The mechanism by which S-RNase confers its specificity is unknown. However, a hypervariable region (RHV in Rosaceae and HVa-HVb in Solanaceae) that differs among the many cloned S-RNase alleles has been proposed to be involved in conferring the S-haplotype specificity of the S-RNase. Region swapping experiments between S-RNases and crystallography of the enzyme support this assumption. However, the deduced amino acid sequences of Sn-RNase and Si-RNase alleles from the European pear (Pyrus communis) were recently found to have an identical RHV. In the present study it is shown that Sn-RNase does not prevent fertilization by Si-pollen haplotype, thus presenting a case in which RHV is not required for the determination of specific pollen rejection by S-RNase, and implying that other regions in the enzyme may be sufficient for this specificity.

The Causes of self-sterility in natural populations of the relictual angiosperm, Illicium floridanum (Illiciaceae).

Illicium floridanum, a species belonging to the basal extant angiosperm taxon Illiciaceae, reportedly exhibits self-incompatibility (SI). To date, the site and timing of SI within the carpel of this species remains unidentified. Thus, the objective of this research was to determine the cellular and temporal aspects of SI in I. floridanum. Following controlled application of cross- and self-pollen in natural populations of I. floridanum, embryo sac development and temporal aspects of stigma receptivity, as well as pollen tube growth, fertilization, and embryo and endosperm development, were investigated with the aid of light and fluorescence microscopy. Flowers of I. floridanum exhibited complete dichogamy whereby stigmas only supported cross- and self-pollen tube growth prior to anther dehiscence. In contrast to earlier reports of SI in this species, a prezygotic SI resulting in rejection of self-pollen tube growth at the stigma was absent and there were no significant differences between cross- versus self-pollen germination and pollen tube growth within the style and ovary during the first 5 d after pollination. Structural development of the four-celled embryo sac was not differentially influenced by pollen type as noted to occur in other angiosperms with late-acting ovarian SI. The ovule micropyle and embryo sac were penetrated equally by cross- and self-pollen tubes. In addition, there were no statistically significant differences in cross- versus self-fertilization. A resting zygote and multicellular endosperm at a variety of developmental stages was present by 30 d after application of cross- or self-pollen. In the clear absence of a prezygotic SI that was previously reported to result in differential self-pollen tube growth at the stigma, self- sterility in I. floridanum is likely due to early-acting inbreeding depression, although late-acting post-zygotic ovarian SI cannot be ruled out.

Effects of self-, chase and mixed self/cross-pollinations on pistil longevity and fruit set in Ceiba species (Bombacaceae) with late-acting self-incompatibility.

Late-acting self-incompatibility (LSI), in which selfed flowers fail to form fruits despite apparently successful growth of the pollen tubes to the ovules, is a contentious and still poorly understood phenomenon. Some studies have indicated pollen tube-pistil interactions, and major gene control. Others favour an early acting inbreeding depression explanation. Experimental pollinations, including selfs (in a subsample of which the style was cut before pollen tubes reached the ovary), chase self/cross-pollinations, crosses, and mixed self/cross-pollinations were used to study floral/pistil longevity and effect on fruit set and seed yield in two Ceiba species known to have LSI. Self-pollinations, including those with a cut style, had extended floral longevity compared with unpollinated flowers. Chase pollinations in which cross-pollen was applied up to 3 h after selfing set fruits, but with reduced seed set compared with crosses. Those with cross-pollen applied at 4 and 8 h after self-pollination all failed to set fruits. Flowers subjected to 1 : 1 and 2 : 1 self/cross-pollinations all produced fruits but again with a significantly lower seed set compared with crosses. Extended floral longevity initiated with self-pollen tubes growing in the style indicates some kind of pollen tube-pistil interaction. Fruit set only in chase pollinations up to 3 h implies that self-pollen tubes either grow more slowly in the style or penetrate ovules more slowly on arrival at the ovary compared with cross-tubes. This agrees with previous observations indicating that the incidence of penetrated ovules is initially lower in selfed compared with crossed pistils. However, the low seed yield from mixed pollinations indicates that self- and cross-pollen tubes arrive at the ovary and penetrate ovules more or less simultaneously. Possible explanations for these discordant results are discussed.

Identification of the pollen determinant of S-RNase-mediated self-incompatibility.

Many flowering plants have adopted self-incompatibility mechanisms to prevent inbreeding and promote out-crosses. In the Solanaceae, Rosaceae and Scrophulariaceae, two separate genes at the highly polymorphic S-locus control self-incompatibility interactions: the S-RNase gene encodes the pistil determinant and the previously unidentified S-gene encodes the pollen determinant. S-RNases interact with pollen S-allele products to inhibit the growth of self-pollen tubes in the style. Pollen-expressed F-box genes showing allelic sequence polymorphism have recently been identified near to the S-RNase gene in members of the Rosaceae and Scrophulariaceae; but until now have not been directly shown to encode the pollen determinant. Here we report the identification and characterization of PiSLF, an S-locus F-box gene of Petunia inflata (Solanaceae). We show that transformation of S1S1, S1S2 and S2S3 plants with the S2-allele of PiSLF causes breakdown of their pollen function in self-incompatibility. This breakdown of pollen function is consistent with 'competitive interaction', in which pollen carrying two different pollen S-alleles fails to function in self-incompatibility. We conclude that PiSLF encodes the pollen self-incompatibility determinant.