Evidence for ovarian self-incompatibility as a cause of self-sterility in the relictual woody angiosperm,Pseudowintera axillaris (Winteraceae)

Species within the genus Pseudowintera exhibit high rates of self-sterility. Self-sterility in the genus has been previously posited-but not confirmed-to be the result of late-acting ovarian self-incompatibility (OSI) functioning within nucellar tissue of the ovule to prevent self pollen tubes from entering the embryo sac. Structural and functional aspects of pollen-carpel interactions and early seed development following cross- and self-pollination were investigated in P. axillaris to determine the site, timing and possible mechanisms of self-sterility. No significant differences were observed between pollen tube growth, ovule penetration and double fertilization following cross- and self-pollination. Pollen tubes exhibited phasic growth in an extracellular matrix composed of proteins and carbohydrates, as well as arabinogalactans/arabinogalactan proteins. A uniform failure in embryo sac development prior to division of the zygote was apparent within 15 d following double fertilization by self gametes. Results indicate that SI mechanisms in P. axillaris do not prevent double fertilization from occurring. Instead, mechanisms of self-sterility affect post-zygotic development of the embryo sac. Although self-sterility may be attributed to inbreeding depression, given the post-zygotic nature of failure in embryo sac development, the possibility of late-acting OSI is discussed.     

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

BACKGROUND AND AIMS: 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. METHODS: 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. KEY RESULTS: 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. CONCLUSIONS: 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.     

Do s genes or deleterious recessives control late-acting self-incompatibility in Handroanthus heptaphyllus (Bignoniaceae)? A diallel study with four full-sib progeny arrays

Background and AimsGenetically controlled self-incompatibility (SI) mechanisms constrain selfing and thus have contributed to the evolutionary diversity of flowering plants. In homomorphic gametophytic SI (GSI) and homomorphic sporophytic SI (SSI), genetic control is usually by the single multi-allelic locus S. Both GSI and SSI prevent self pollen tubes reaching the ovary and so are pre-zygotic in action. In contrast, in taxa with late-acting self-incompatibility (LSI), rejection is often post-zygotic, since self pollen tubes grow to the ovary, where fertilization may occur prior to floral abscission. Alternatively, lack of self fruit set could be due to early-acting inbreeding depression (EID). The aim of our study was to investigate mechanisms underlying the lack of selfed fruit set in Handroanthus heptaphyllus in order to assess the likelihood of LSI versus EID.MethodsWe employed four full-sib diallels to study the genetic control of LSI in H. heptaphyllus using a precociously flowering variant. We also used fluorescence microscopy to study the incidence of ovule penetration by pollen tubes in pistils that abscised following pollination or initiated fruits.Key ResultsAll diallels showed reciprocally cross-incompatible full sibs (RCIs), reciprocally cross-compatible full sibs (RCCs) and non-reciprocally compatible full sibs (NRCs) in almost equal proportions. There was no significant difference between the incidences of ovule penetrations in abscised pistils following self- and cross-incompatible pollinations, but those in successful cross-pollinations were around 2-fold greater.ConclusionsA genetic model postulating a single S locus with four S alleles, one of which, in the maternal parent, is dominant to the other three, will produce RCI, RCC and NRC full sib situations each at 33 %, consistent with our diallel results. We favour this simple genetic control over an EID explanation since none of our pollinations, successful or unsuccessful, resulted in partial embryo development, as would be expected under a whole-genome EID effect.     

Pollination and late-acting self-incompatibility in Cyrtanthus breviflorus (Amaryllidaceae): implications for seed production

Background and Aims

Animal pollination is typically an uncertain process that interacts with self-incompatibility status to determine reproductive success. Seed set is often pollen-limited, but species with late-acting self-incompatibility (SI) may be particularly vulnerable, if self-pollen deposition results in ovule discounting. Pollination is examined and the occurrence of late-acting SI and ovule discounting assessed in Cyrtanthus breviflorus.

Methods

The pollination system was characterized by observing floral visitors and assessing nectar production and spectral reflectance of flowers. To assess late-acting SI and ovule discounting, growth of self- and cross-pollen tubes, and seed set following open pollination or hand pollination with varying proportions of self- and cross-pollen, were examined.

Key Results

Native honeybees Apis mellifera scutellata pollinated flowers as they actively collected pollen. Most flowers (≥70 %) did not contain nectar, while the rest produced minute volumes of dilute nectar. The flowers which are yellow to humans are visually conspicuous to bees with a strong contrast between UV-reflecting tepals and UV-absorbing anthers and pollen. Plants were self-incompatible, but self-rejection was late-acting and both self- and cross-pollen tubes penetrated ovules. Seed set of open-pollinated flowers was pollen-limited, despite pollen deposition exceeding ovule number by 6-fold. Open-pollinated seed set was similar to that of the cross + self-pollen treatment, but was less than that of the cross-pollen-only treatment.

Conclusions

Flowers of C. breviflorus are pollinated primarily by pollen-collecting bees and possess a late-acting SI system, previously unknown in this clade of the Amaryllidaceae. Pollinators of C. breviflorus deposit mixtures of cross- and self-pollen and, because SI is late-acting, self-pollen disables ovules, reducing female fertility. This study thus contributes to growing evidence that seed production in plants with late-acting SI systems is frequently limited by pollen quality, even when pollinators are abundant.     

Differential ovule development following self- and cross-pollination: the basis of self-sterility in Narcissus triandrus (Amaryllidaceae)

Self-pollination results in significantly lower seed set than cross-pollination in tristylous Narcissus triandrus. We investigated structural and functional aspects of pollen–pistil interactions and ovule–seed development following cross- and self-pollination to assess the timing and mechanism of self-sterility. Ovule development within an ovary was asynchronous at anthesis. There were no significant differences in pollen tube behavior following cross- vs. self-pollination during the first 6 d of growth, regardless of style morph type. Double fertilization was significantly higher following cross- vs. self-pollination. Aborted embryo development was not detected following either pollination type up to seed maturity. Prior to pollen tube entry, a significantly greater number of ovules ceased to develop following self- vs. cross-pollination. These results indicate that self-sterility in N. triandrus operates prezygotically but does not involve differential pollen tube growth typical of many self-incompatibility (SI) systems. Instead, low seed set following self-pollination is caused by a reduction in ovule availability resulting from embryo sac degeneration. We hypothesize that this is due to the absence of a required stimulus for normal ovule development. If this is correct, current concepts of SI may need to be broadened to include a wider range of pollen–pistil interactions.     

Early Inbreeding Depression and Pollen Competition inCalluna vulgaris(L.) Hull.

We investigated whether partial self-sterility inCalluna vulgarisresultsfrom abortion of selfed offspring owing to inbreeding depressionor a late-acting self-incompatibility mechanism, and whetherself-pollen interferes with normal functioning of cross-pollen.Self-pollination resulted in 75% less seed set than cross-pollination.Self-pollen tubes reached ovaries and penetrated ovules as oftenas those of cross-pollen. Following self-pollination, examinationof the size of undeveloped seeds showed that at least 70% resultedfrom ovule fertilization and arrest of development occurredat various stages. All self-pollinated plants produced seedsand self-fertility varied among plants. These results indicatethat the reduced seed set observed in self-pollination is morelikely the result of inbreeding depression rather than a late-actingself-incompatibility system. The fecundity component of inbreedingdepression was high (0.762). Seed set was reduced by an averageof 40% when self-pollen was mixed with cross-pollen, comparedto pure cross-pollination. Using genetic markers, we found about20% of seeds resulted from self-pollination in mixed-pollinatedfruits.C. vulgarisis likely to experience self-pollination innature and our data suggest this will reduce the number of ovulesthat might otherwise mature after cross-pollination.Copyright1999 Annals of Botany Company Calluna vulgaris(heather), self-pollination, pollen tube, ovule fertilization, early inbreeding depression, pollen interference.     

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.     

THE EFFECTS OF FIVE GENERATIONS OF ENFORCED SELFING ON POTENTIAL MALE AND FEMALE FUNCTION IN MIMULUS GUTTATUS

In prior work we detected no significant inbreeding depression for pollen and ovule production in the highly selfing Mimulus micranthus, but both characters showed high inbreeding depression in the mixed-mating M. guttatus. The goal of this study was to determine if the genetic load for these traits in M. guttatus could be purged in a program of enforced selfing. These characters should have been under much stronger selection in our artificial breeding program than previously reported characters such as biomass and total flower production because, for example, plants unable to produce viable pollen could not contribute to future generations. Purging of genetic load was investigated at the level of both the population and the individual maternal line within two populations of M. guttatus. Mean ovule number, pollen number, and pollen viability declined significantly as plants became more inbred. The mean performance of outcross progeny generated from crosses between pairs of maternal inbred lines always exceeded that of self progeny and was fairly constant for each trait through all five generations. The consistent performance of outcross progeny and the universally negative relationships between performance and degree of inbreeding are interpreted as evidence for the weakness of selection relative to the quick fixation of deleterious alleles due to drift during the inbreeding process. The selective removal (purging) of deleterious alleles from our population would have been revealed by an increase in performance of outcross progeny or an attenuation of the effects of increasing homozygosity. The relationships between the mean of each of these traits and the expected inbreeding coefficient were linear, but one population displayed a significant negative curvilinear relationship between the log of male fertility (a function of pollen number and viability) and the inbreeding coefficient. The generally linear form of the responses to inbreeding were taken as evidence consistent with an additive model of gene action, but the negative curvilinear relationship between male fertility and the inbreeding coefficient suggested reinforcing epistasis. Within both populations there was significant genetic variation among maternal lineages for the response to inbreeding in all traits. Although all inbred lineages declined at least somewhat in performance, several maternal lines maintained levels of performance just below outcross means even after four or five generations of selfing. We suggest that selection among maternal lines will have a greater effect than selecting within lines in lowering the genetic load of populations.     

Inbreeding depression and mixed mating in Leptosiphon jepsonii: a comparison of three populations

BACKGROUND AND AIMS: Inbreeding depression is thought to play a central role in the evolution and maintenance of cross-fertilization. Theory indicates that inbreeding depression can be purged with self-fertilization, resulting in positive feedback for the selection of selfing. Variation among populations of Leptosiphon jepsonii in the timing and rate of self-fertilization provides an opportunity to study the evolution of inbreeding depression and mating systems. In addition, the hypothesis that differences in inbreeding depression for male and female fitness can stabilize mixed mating in L. jepsonii is tested. METHODS: In a growth room experiment, inbreeding depression was measured in three populations with mean outcrossing rates ranging from 0.06 to 0.69. The performance of selfed and outcrossed progeny is compared at five life history stages. To distinguish between self-incompatibility and early inbreeding depression, aborted seeds and unfertilized ovules were counted in selfed and outcrossed fruits. In one population, pollen and ovule production was quantified to estimate inbreeding depression for male and female fitness. KEY RESULTS: Both prezygotic barriers and inbreeding depression limited self seed set in the most outcrossing population. Cumulative inbreeding depression ranged from 0.297 to 0.501, with the lowest value found in the most selfing population. Significant inbreeding depression for early life stages was found only in the more outcrossing populations. Inbreeding depression was not significant for pollen or ovule production. CONCLUSIONS: The results provide modest support for the hypothesized relationship between inbreeding depression and mating systems. The absence of early inbreeding depression in the more selfing populations is consistent with theory on purging. Differences in male and female expression of inbreeding depression do not appear to stabilize mixed mating in L. jepsonii. The current estimates of inbreeding depression for L. jepsonii differ from those of previous studies, underscoring the effects of environmental variation on its expression.     

Embryonic inbreeding depression varies among populations and by mating system in Witheringia solanacea (Solanaceae)

? Premise of the study: Embryonic inbreeding depression is a key influence on mating system evolution and can be difficult to estimate in self-incompatible species. A pollen chase experiment was used to estimate the magnitude of embryonic inbreeding depression in Costa Rican Witheringia solanacea, a species polymorphic for self-incompatibility (SI). In a pollen chase experiment, bud self-pollinations are followed after anthesis by outcross pollinations, with a comparable pair of outcross pollinations used as a control. Lowered seed set for the self-precedence treatment indicates embryonic inbreeding depression. ? Methods: Embryonic inbreeding depression was assayed for self-compatible (SC) individuals and for SI plants from two populations that differ quantitatively in the onset and enzymatic activity of their SI response. Microsatellite markers were used to assay the selfing rate of a sample of surviving progeny from the prior self-pollination treatment. ? Key results: SC individuals showed no evidence of embryonic inbreeding depression. In SI plants, prior self-pollination reduced seed number by 28-70%, depending on population. Microsatellite genotyping revealed that embryonic inbreeding depression was even more severe than estimated by the phenotypic data: for mature fruits resulting from self-pollination precedence, the majority of the progeny were the result of outcross fertilization. ? Conclusions: Lineage-specific purging of recessive lethals has accompanied the evolution of SC in this species. SI populations show contrasting levels of embryonic inbreeding depression, with nearly complete embryonic lethality upon selfing in the Monteverde population. In the face of high embryonic inbreeding depression, an increase in selfing rate can evidently occur only under severe pollen limitation.     

Multiple factors contribute to outcrossing in a tropical emergent Dipterocarpus tempehes, including a new pollen-tube guidance mechanism for self-incompatibility

The self-rejection system of Dipterocarpus tempehes (Dipterocarpaceae), an emergent tree of the lowland tropical forests of Borneo, were studied by means of pollination experiments, fluorescence microscopy of pollen tubes, and monitoring of ovary maturation patterns. Fruit set was higher in cross-pollinated flowers than in control and self-pollinated flowers, indicating the existence of pollen limitation and a self-rejection system. Although the adhesion and the germination of self-pollen and the growth of self-pollen tubes were not inhibited, the proportion of cross-pollen tubes that entered the style was 1.7-2.3 times higher than that of self-pollen tubes, indicating a partial self-incompatibility that inhibits self-pollen tubes from entering the style hollow. These results suggest, for the first time, that self-incompatibility is caused by a defect of pollen-tube guidance. We also suggest a threshold effect in number of pollen tubes or late-acting self-incompatibility to fully explain the drastic and uniform selection against self-pollinated flowers before ovary swelling. After that, maternal selection and/or inbreeding depression caused the abortion and delayed maturation of self-pollinated flowers. The advantages of the self-rejection process composed of partial early-acting self-incompatibility and relatively strong delayed abortion is discussed with respect to the general-flowering phenomenon in lowland dipterocarp forests.     

基于S-核酸酶的自交不亲和性的分子机制

自交不亲和性是一种广泛存在于显花植物中的种内生殖障碍,可以抑制近亲繁殖而促进异交。其中,以茄科、玄参科和蔷薇科为代表的配子体自交不亲和性是最常见的类型。这类自交不亲和性是由单一的多态性S-位点所控制。目前的研究发现这一位点至少包含两个自交不亲和反应特异性决定因子：花柱中的S-核酸酶和花粉中的SLF（S-Locus F-box）蛋白。该文将主要介绍并讨论基于S-核酸酶的自交不亲和性分子机制的研究进展。     

基于S- 核酸酶的自交不亲和性的分子机制

自交不亲和性是一种广泛存在于显花植物中的种内生殖障碍, 可以抑制近亲繁殖而促进异交。其中, 以茄科、玄参科和蔷薇科为代表的配子体自交不亲和性是最常见的类型。这类自交不亲和性是由单一的多态性S-位点所控制。目前的研究发现这一位点至少包含两个自交不亲和反应特异性决定因子: 花柱中的S-核酸酶和花粉中的SLF(S-Locus F-box)蛋白。该文将主要介绍并讨论基于S-核酸酶的自交不亲和性分子机制的研究进展。     

Breeding systems in Clivia (Amaryllidaceae): late‐acting self‐incompatibility and its functional consequences

Late‐acting (ovarian) self‐incompatibility, characterized by minimal or zero seed production following self‐pollen tube growth to the ovules, is expected to show phylogenetic clustering, but can otherwise be difficult to distinguish from early‐acting inbreeding depression. In Amaryllidaceae, late‐acting self‐incompatibility has been proposed for Narcissus (Narcisseae) and Cyrtanthus (Cyrtantheae). Here, we investigate whether it occurs in the horticulturally important genus Clivia (Haemantheae) and test whether species in this genus experience ovule discounting in wild populations. Seed‐set results following controlled hand pollinations revealed that Clivia miniata and C. gardenii are largely self‐sterile. Self‐ and cross‐pollinated flowers of both species had similar proportions of pollen tubes entering the ovary, and those of C. gardenii also did not differ in the proportions of pollen tubes that penetrated ovules, thus ruling out classical gametophytic self‐incompatibility acting in the style, but not early inbreeding depression. Flowers that received equal mixtures of self‐ and cross‐pollen set fewer seeds than those that received cross‐pollen only, but it was unclear whether this effect was a result of ovule discounting or interactions on the stigma. The prevention of self‐pollination by the emasculation of either single flowers or whole inflorescences in wild populations did not affect seed set, suggesting that ovule discounting is not a major natural limitation on seed production. Flowers typically produce one to three large fleshy seeds from approximately 16 available ovules, even when supplementally hand pollinated, suggesting that fecundity is mostly resource limited. The results of this study suggest that Clivia spp. are largely self‐sterile as a result of either a late‐acting self‐incompatibility system or severe early inbreeding depression, but ovule discounting caused by self‐pollination is not a major constraint on fecundity. © 2014 The Linnean Society of London, Botanical Journal of the Linnean Society, 2014, 175 , 155–168.     

The Role of Late-Acting Self-Incompatibility and Early-Acting Inbreeding Depression in Governing Female Fertility in Monkshood,Aconitum kusnezoffii

Reduced seed yields following self-pollination have repeatedly been observed, but the underlying mechanisms remain elusive when self-pollen tubes can readily grow into ovaries, because pre-, post-zygotic late-acting self-incompatibility (LSI), or early-acting inbreeding depression (ID) can induce self-sterility. The main objective of this study was to differentiate these processes in Aconitum kusnezoffii, a plant lacking stigmatic or stylar inhibition of self-pollination. We performed a hand-pollination experiment in a natural population of A. kusnezoffii, compared seed set among five pollination treatments, and evaluated the distribution of seed size and seed set. Embryonic development suggested fertilization following self-pollination. A partial pre-zygotic LSI was suggested to account for the reduced seed set by two lines of evidence. The seed set of chase-pollination treatment significantly exceeded that of self-pollination treatment, and the proportion of unfertilized ovules was the highest following self-pollination. Meanwhile, early-acting ID, rather than post-zygotic LSI, was suggested by the findings that the size of aborted selfed seeds varied continuously and widely; and the selfed seed set both exhibited a continuous distribution and positively correlated with the crossed seed set. These results indicated that the embryos were aborted at different stages due to the expression of many deleterious alleles throughout the genome during seed maturation. No signature of post-zygotic LSI was found. Both partial pre-zygotic LSI and early-acting ID contribute to the reduction in selfed seed set in A. kusnezoffii, with pre-zygotic LSI rejecting part of the self-pollen and early-acting ID aborting part of the self-fertilized seeds.     

鹤顶兰花粉管在子房中的生长途径

运用扫描电镜对鹤顶兰(Phaiustankervilliae(Aiton)Bl.)花粉管在子房内的生长途径进行了观察。结果表明:花粉管在子房中的生长途径可以分为3个阶段:(1)沿子房壁轴向生长阶段,从授粉开始至大孢子母细胞四分体时期,花粉管经过合蕊柱到达子房,经由胎座基部沿子房壁轴向生长;(2)沿子房径向生长阶段,二核胚囊之后,花粉管在胚珠之间穿梭,以径向生长为主;(3)朝珠孔定向生长阶段,胚囊成熟时,花粉管朝珠孔定向生长进入胚囊。实验结果说明花粉管的定向生长受胚珠的分子信号调控。     

Inbreeding depression in two species of Mimulus (Scrophulariaceae) with contrasting mating systems

We examined the effect of self- and cross-pollination on germination success, flowering probability, pollen and ovule production, survivorship, and adult aboveground biomass in two species of Mimulus with contrasting mating systems: the highly seifing M. micranthus and an outcrossing population of M. guttatus. Cross-pollinations were performed both within and between populations in order to examine the scale at which the genetic load is distributed. We found significant inbreeding depression in M. guttatus in four of the six traits, with the highest inbreeding depression observed in biomass (68% and 69% based on within- and between-population crosses, respectively) and lowest in ovule production (21% based on between-population crosses only). M. micranthus displayed significant inbreeding depression in only two of the six traits examined. Again, we observed the highest inbreeding depression in biomass (47–60% based on within- and between-population crosses, respectively), but both traits showing significant differences between self and outcross progeny expressed lower inbreeding depression than in M. guttatus. We detected no significant inbreeding depression for either pollen or ovule production in M. micranthus. An estimate of total inbreeding depression based on the multiplicative effects of all traits was also lower in M. micanthus than∗∗∗ in M. guttatus. Our results are consistent with the expected purging of genetic load in populations with high selfing rates. The absence of inbreeding depression in M. micranthus pollen and ovule production, two traits with strong links to fitness in a selfing annual, further suggests the important role of directional selection in determining the population's genetic load. Comparison of cross-pollinations made within and between populations revealed little evidence of divergence of genetic load among the M. micranthus and M. guttatus populations examined.     

Effects of pollen shortage and self-pollination on seed production of an endangered tree, Magnolia stellata

BACKGROUND AND AIMS: Pollen limitation is a significant determinant of seed production, and can result from both insufficient pollen quantity (pollen shortage) and quality (mainly relating to self-pollination). For animal-pollinated tree species with large floral displays, pollen limitation may be determined by a balance between increased pollen quantity due to increased attractiveness for pollinators, countered by increased self-pollination due to increased geitonogamy. The contributions of pollen shortage and self-pollination on seed production were quantitatively examined in the natural pollination of an insect-pollinated, dichogamous, endangered tree, Magnolia stellata, which has a large, showy floral display. METHODS: Manual self- and cross-pollinations were conducted to determine the effects of selfing on seed production. The outcrossing rate was measured using microsatellite analyses of open-pollinated seeds, and the embryo mortality rate caused by self-pollination was indirectly estimated. The frequency of ovule mortality due to pollen shortage was also inferred using the embryo mortality and ovule survival rates from natural pollination. KEY RESULTS: The average fruit set, seed set per fruit, and ovule survival rate per tree from hand cross-pollination were 1.37, 3.15, and 3.34 times higher than those from hand self-pollination, respectively, indicating that self-pollination causes inbreeding depression for fruit and seed set. The multilocus-outcrossing rate (t(m)) was intermediate, 0.632, and the primary selfing rate was 0.657. This indicates that frequent geitonogamous selfing occurs. The ovule mortality rate due to pollen shortage and the embryo mortality rate due to self-pollination were estimated to be 80.8 % and 45.9 %, respectively. CONCLUSIONS: It is concluded that seed production of M. stellata is strongly limited by both pollen shortage and self-pollination. Inefficient beetle-pollination and the automimicry system via asynchronous flowering might be responsible for the high level of pollen shortage and frequent geitonogamy. This is despite a large, showy floral display and the dichogamous system of the species.     

An uncoupling screen for autonomous embryo mutants in Arabidopsis thaliana

Simple de novo screens in Arabidopsis thaliana have previously identified mutants that affect endosperm development but viable-embryo mutants have not been identified. Our strategy to identify autonomous embryo development was to uncouple embryo and endosperm fertilisation. This involved a male-sterile mutant population being crossed with a distinct pollen parent—the pollen was needed to initiate endosperm development and because it was distinct, the maternal progeny could be selected from the hybrid population. This process was refined over three stages, resulting in a viable approach to screen for autonomous embryo mutants. From 8,000 screened plants, a mutation was isolated in which the integument cells extended from the ovule and proliferated into a second complete twinned ovule. Some embryos from the mutant were normal but others developed fused cotyledons. In addition, a proportion of the progeny lacked paternal genes.     

Inbreeding depression in small populations of self-incompatible plants.

Self-incompatibility (SI) is a widespread mechanism that prevents inbreeding in flowering plants. In many species, SI is controlled by a single locus (the S locus) where numerous alleles are maintained by negative frequency-dependent selection. Inbreeding depression, the decline in fitness of selfed individuals compared to outcrossed ones, is an essential factor in the evolution of SI systems. Conversely, breeding systems influence levels of inbreeding depression. Little is known about the joint effect of SI and drift on inbreeding depression. Here we studied, using a two-locus model, the effect of SI (frequency-dependent selection) on a locus subject to recurrent deleterious mutations causing inbreeding depression. Simulations were performed to assess the effect of population size and linkage between the two loci on the level of inbreeding depression and genetic load. We show that the sheltering of deleterious alleles linked to the S locus strengthens inbreeding depression in small populations. We discuss the implications of our results for the evolution of SI systems.