Ultrastructural and molecular study of plastid inheritance in Abies alba and some Abies hybrids

 The ultrastructure of egg cells in Abies alba was examined to elucidate the lack of maternal inheritance of plastids. Before fertilization, maternal plastids are absent in the perinuclar zone containing mainly mitochondria and smooth endoplasmic reticulum. During egg cell development the maternal plastids are transformed into large inclusions which are situated mostly towards the periphery of the egg cell, and finally disintegrate. As a consequence, they do not participate in zygote formation. RFLP analysis of cpDNA of parental trees and their F1 interspecific hybrids (A. alba×A. numidica, A. alba×A. nordmanniana, A. nordmanniana×A. Alba) using HindIII and BamHI showed a paternal mode of cpDNA inheritance. Paternal inheritance has also been found with PCR/RFLP analysis of cpDNA from parental trees and their hybrids (A. alba×A. pinsapo, A. pinsapo×A. alba, A. pinsapo×A. numidica) using ApaI and HaeIII digests, as well as in the crosses of A. cephalonica×A. nordmanniana, A. nordmanniana×A. cephalonica, A. cephalonica×A. numidica using TagI digests. Received: 1 June 1998 / Accepted: 14 August 1998     

Crossability relationships between Korean firs Abies koreana , A. nephrolepis and A. holophylla and some other representatives of the genus Abies

Artificial crossing experiments involving 3 Abies species native to Korean peninsula and 5 other representatives of Abies revealed a high hybridological affinity between Abies koreana and A. nephrolepis. Both these species are reproductively isolated from A. holophylla. All the three Korean species were found to exhibit incompatible relationships with the North American species A. concolor. The species A. holophylla and A. koreana differ also in their abilities to intercross with the Mediterranean firs. The former has been successfully crossed with A. nordmanniana, A. alba and A. cilicica exhibiting 19.1–55.3% crossability, whereas the latter produced filled seeds only with A. nordmanniana reaching 46.4% crossability. A considerable differentiation is postulated to exist between the pair of species A. koreana and A. nephrolepis on the one side and A. holophylla on the other side.     

Doubled haploids of novel trigenomic Brassica derived from various interspecific crosses

To develop doubled haploid (DH) mapping populations of hexaploid Brassica, 10 F₁ hybrids derived from crosses between allohexaploid Brassica parents were evaluated in this study. The allohexaploid Brassica parents were selfed progenies of unique interspecific crosses between Brassica rapa (genome AA) × B. carinata (BBCC), B. nigra (BB) × B. napus (AACC), and a complex cross between B. juncea (AABB), B. napus and B. carinata, with relatively stable chromosome number (2n = 54). Hexaploid status and chromosome behavior during meiosis I in four promising F₁ hybrids were assessed using microscopy and flow cytometry, and progeny were obtained following microspore culture. Hybrids H11-2 and H16-1 demonstrated higher amenability for embryo generation, plantlet regeneration, and frequency of production of DH microspore-derived progeny of hexaploid DNA content (6x) compared to hybrids H08-1 and H24-1. A total of 370 6x DH progeny were selected out of 693 plantlets from H11-2, 241/436 from H16-1, 23/54 from H08-1, and 21/56 from H24-1. DH progenies of hybrids H11-2 and H16-1 were then designated as promising mapping populations of a new hexaploid Brassica species.     

Induction,protein composition and DNA ploidy level of embryogenic calli of silver fir and its hybrids

Somatic embryogenesis was initiated from immature zygotic embryos of intraspecific cross ofAbies alba and interspecific combination ofAbies alba x Abies nordmanniana. The most responsive explants for induction of embryogenic calli were zygotic embryos in precotyledonary stage of development. Biochemically the compared embryogenic lines were uniform irrespective of their morphology or embryogenic potential. The flow cytometric analysis of single embryogenic lines suggests that no changes in ploidy levels occur during induction and culture of embryogenic callus inAbies alba what confirms the convenience of this system for propagation of valuable seed material.     

Morphological and genetic characteristics of genetic tumor in Nicotiana glauca

It is well understood that genetic tumors develop in certain interspecific Nicotiana hybrids. Nicotiana species are divided into “plus” and “minus” groups and crosses between “plus” and “minus” species give rise to tumorous hybrids. However, it has been proposed that parents and hybrids derived from crosses among members within the same group do not produce tumors. In this study, genetic tumors were only obtained in Nicotiana glauca, which exhibited tumor features similar to those of N. glauca × N. langsdorffii. Our results suggest that genetic factors may control tumor formation independent of tumor induction dependent on the specific interspecific cross. Genetic tumor formation exhibited high B-type and D-type cyclin expression levels, indicating tumor cells are characterized by an uncontrolled cell cycle.     

Paternal inheritance of plastids in interspecific hybrids of the genus Actinidia revealed by PCR-amplification of chloroplast DNA fragments

RFLPs (restriction fragment length polymorphisms) of PCR (polymerase chain reaction) -amplified fragments were used to trace the pattern of plastid DNA inheritance in the genus Actinidia. A total of 51 progeny originating from interspecific crosses between three A. arguta cultivars and A. deliciosa, the kiwifruit, and 12 progeny originating from the cross between A. kolomikta and A. chinensis were analysed together with their parents. No reciprocal crosses could be tested since they all failed to set viable seeds. Attempts to rescue immature embryos failed in all cases as well. The A. arguta×A. deliciosa crosses were checked for the RFLP patterns of a sequence encoding part of the Rubisco large subunit (rbcL), using either AluI or MseI, and for a sequence encoding part of the photosystem II D1 protein (psbA), using HinfI. The A. kolomikta×A. chinensis cross was checked for the RFLP patterns of sequences encoding the spacers between trnT and the 5′-trnL exon (a-b spacer DNA) and the trnL 3′ exon and trnF (e-f spacer DNA), respectively. The first spacer revealed a natural polymorphism between the two parent species due to a large deletion occurring in A. kolomikta detectable without further restriction enzyme treatment. The e-f spacer DNA was digested with HinfI. The comparison of the RFLP patterns in the parents and their progeny showed a strictly paternal inheritance of chloroplast DNA in Actinidia, with no exception found in any of the crosses examined. As the reciprocal crosses were not available, we do not know whether paternal inheritance of plastids is restricted to the crosses we analysed or if this is the general rule for plastid inheritance in the genus Actinidia. Actinidia is dioecious and is the first purely outbreeding species for which a paternal plastid inheritance has so far been documented.     

Geographical variation in postzygotic isolation and its genetic basis within and between two Mimulus species

The aim of this study is to investigate the evolution of intrinsic postzygotic isolation within and between populations of Mimulus guttatus and Mimulus nasutus. We made 17 intraspecific and interspecific crosses, across a wide geographical scale. We examined the seed germination success and pollen fertility of reciprocal F₁ and F₂ hybrids and their pure-species parents, and used biometrical genetic tests to distinguish among alternative models of inheritance. Hybrid seed inviability was sporadic in both interspecific and intraspecific crosses. For several crosses, Dobzhansky–Muller incompatibilities involving nuclear genes were implicated, while two interspecific crosses revealed evidence of cytonuclear interactions. Reduced hybrid pollen fertility was found to be greatly influenced by Dobzhansky–Muller incompatibilities in five out of six intraspecific crosses and nine out of 11 interspecific crosses. Cytonuclear incompatibilities reduced hybrid fitness in only one intraspecific and one interspecific cross. This study suggests that intrinsic postzygotic isolation is common in hybrids between these Mimulus species, yet the particular hybrid incompatibilities responsible for effecting this isolation differ among the populations tested. Hence, we conclude that they evolve and spread only at the local scale.     

Halophilic and salts tolerant protoplast cultures of mangrove plants, Sonneratia alba and Avicennia alba

The effects of sea salts, NaCl, KCl, MgCl₂, MgSO₄, and CaCl₂, on the growth of protoplast cultures of two mangrove species, Sonneratia alba and Avicennia alba, were investigated using 96-well culture plates. Plants of these two species naturally grow at the seaward side of a mangrove forest. Cotyledon protoplasts of S. alba showed halophilic nature to NaCl, KCl, and MgCl₂ at low concentrations (10–50 mM) when cultured in Murashige and Skoog’s (MS) medium containing 0.6 M mannitol. CaCl₂ at a concentration higher than 25 mM was inhibitory to cell growth. On the other hand, in protoplast culture of A. alba suspension cells, which were induced from cotyledon tissues, in the modified amino acid (mAA) medium containing 1.2 M sorbitol, tolerance to NaCl, MgCl₂ and MgSO₄ were observed at a wide range of concentrations up to 400 mM. CaCl₂ was always inhibitory for cell divisions in A. alba, but stimulatory for spherical enlargement of cells. However, no difference in cell enlargement was observed among other salts. Similarity and difference in reactivity to salts between protoplasts and suspension cells from our previous studies were discussed in relation to the site of salt tolerance or halophilic adaptation within mangrove cells. For protoplast cultures, the site(s) for response of S. alba and A. alba are located in the cytoplasm and/or the cell membrane.     

Hybridization and introgression in Carex aquatilis and C. paleacea

The diversification and distribution of Abies species throughout the Mediterranean region has led to a complex of species which are difficult to classify. An open question is whether these mainly allopatric taxa have exchanged genetic information via secondary contact. We studied the variation and geographic distribution of paternally inherited chloroplast DNA markers in nine Mediterranean Abies taxa. Markers with high and low mutation rates were applied in order to differentiate among a scenario of secondary genetic contact and a scenario of complete isolation after speciation. The observed molecular variation was analysed using statistical parsimony, geostatistics, and measures of population genetic differentiation. Ancient paternal lineages, represented by markers with low mutation rates, were shared among species. The central and widespread A. alba retained all ancient lineages whereas other species exhibited fewer, down to a single lineage. In contrast, modern lineages, depicted by markers with high mutation rates, were largely separated among species. The western Mediterranean A. pinsapo and A. numidica were clearly separated from each other and from the remaining Abies species. This indicates the absence of secondary contact. The same scenario applies to the eastern Mediterranean Abies species. An exception is the parapatric complex of A. alba, A. cephalonica, and their supposed hybrid A. borisii regis, which exhibited evidence of secondary contact.     

Paternal inheritance of chloroplast DNA and maternal inheritance of mitochondrial DNA in the genus Actinidia

PCR amplification of four chloroplast DNA (cpDNA) and two mitochondrial DNA (mtDNA) regions followed by restriction of the amplified products was used to identify restriction fragment length polymorphisms in 21 Actinidia taxa. Subsequently, the mode of organelle inheritance was investigated in both interspecific and intraspecific controlled crosses made between genotypes showing different cpDNA and/or mtDNA haplotypes. Fifty-six seedlings produced from three interspecific crosses, including in one case the pseudo reciprocal (different genotypes of the same species used as opposite parents), were checked for cpDNA inheritance, and 102 seedlings from the same interspecific crosses and 32 seedlings from two intraspecific crosses within the species A. deliciosa were checked for mtDNA inheritance. In all cases, cpDNA was inherited from the father and mtDNA was inherited from the mother. Maternal inheritance of mtDNA was expected, being the rule in plants, but A. deliciosa is the first genus in angiosperms for which a widespread and strictly paternal inheritance of cpDNA has been reported. Transmission of chloroplastic and mitochondrial genomes through opposite parents provides an exceptional opportunity for studying the paternal and maternal genetic lineages of species in the genus Actinidia.     

An inverse relationship between allelopathic activity and salt tolerance in suspension cultures of three mangrove species,Sonneratia alba,S. caseolaris and S. ovata: development of a bioassay method for allelopathy,the protoplast co-culture method

A bioassay method for allelopathy, the ‘protoplast co-culture method’ was developed to study the relationship between salt tolerance and allelopathy of three mangrove species, Sonneratia alba, S. caseolaris, and S. ovata. Plants of S. alba grow in the seaward-side high salinity region and plants of the latter two species grow in upstream-side regions of a mangrove forest, respectively. Effects of five sea salts (NaCl, KCl, MgCl₂, MgSO₄ and CaCl₂) on the growth of the suspension cells of the latter two species were first investigated by a small-scale method using 24-well culture plates. S. ovata cells showed higher tolerance than S. caseolaris cells to NaCl and other salts, but were not as halophilic as S. alba cells. Protoplasts isolated from suspension cells were co-cultured with lettuce protoplasts in Murashige and Skoog’s (MS) basal medium containing 1 μM 2,4-dichlorophenoxyacetic acid, 0.1 μM benzyladenine, 3 % sucrose and 0.6–0.8 M osmoticum. S. caseolaris protoplasts had a higher inhibitory effect on lettuce protoplast cell divisions than S. alba protoplasts at any lettuce protoplast density, and the effect of S. ovata was intermediate between the two. These results were similar to those obtained from a different in vitro bioassay method for allelopathy, the ‘sandwich method’ with dried leaves. The inverse relationship between allelopathic activity and salt tolerance in suspension cells of Sonneratia mangroves is discussed.     

Genetic diversity in Capsicum germplasm based on microsatellite and random amplified microsatellite polymorphism markers

A sound knowledge of the genetic diversity among germplasm is vital for strategic germplasm collection, maintenance, conservation and utilisation. Genomic simple sequence repeats (SSRs) and random amplified microsatellite polymorphism (RAMPO) markers were used to analyse diversity and relationships among 48 pepper (Capsicum spp.) genotypes originating from nine countries. These genotypes covered 4 species including 13 germplasm accessions, 30 improved lines of 4 domesticated species and 5 landraces derived from natural interspecific crosses. Out of 106 SSR markers, 25 polymorphic SSR markers (24 %) detected a total of 76 alleles (average, 3.04; range, 2–5). The average polymorphic information content (PIC) was 0.69 (range, 0.29–0.92). Seventeen RAMPO markers produced 87 polymorphic fragments with average PIC of 0.63 (range, 0.44–0.81). Dendrograms based on SSRs and RAMPOs generated two clusters. All 38 Capsicum annuum genotypes and an interspecific landrace clustered together, whereas nine non-annuum (three Capsicum frutescens, one Capsicum chinense, one Capsicum baccatum and four interspecific landraces) genotypes clustered separately. Genetic variation within non-annuum genotypes was greater than the C. annuum genotypes. Distinctness of interspecific derivative landraces grown in northeast India was validated; natural crossing between sympatric Capsicum species has been proposed as the mechanism of their origin.     

Maternal inheritance of mitochondrial genomes and complex inheritance of chloroplast genomes in Actinidia Lind.: evidences from interspecific crosses

The inheritance pattern of chloroplast and mitochondria is a critical determinant in studying plant phylogenetics, biogeography and hybridization. To better understand chloroplast and mitochondrial inheritance patterns in Actinidia (traditionally called kiwifruit), we performed 11 artificial interspecific crosses and studied the ploidy levels, morphology, and sequence polymorphisms of chloroplast DNA (cpDNA) and mitochondrial DNA (mtDNA) of parents and progenies. Sequence analysis showed that the mtDNA haplotypes of F1 hybrids entirely matched those of the female parents, indicating strictly maternal inheritance of Actinidia mtDNA. However, the cpDNA haplotypes of F1 hybrids, which were predominantly derived from the male parent (9 crosses), could also originate from the mother (1 cross) or both parents (1 cross), demonstrating paternal, maternal, and biparental inheritance of Actinidia cpDNA. The inheritance patterns of the cpDNA in Actinidia hybrids differed according to the species and genotypes chosen to be the parents, rather than the ploidy levels of the parent selected. The multiple inheritance modes of Actinidia cpDNA contradicted the strictly paternal inheritance patterns observed in previous studies, and provided new insights into the use of cpDNA markers in studies of phylogenetics, biogeography and introgression in Actinidia and other angiosperms.     

Molecular identification and the features of genetic diversity in interspecific hybrids of Amur sturgeon ( Acipenser schrenckii × A. baerii, A. baerii × A. schrenckii, A. schrenckii × A. ruthenus , and A. ruthenus × A. schrenckii ) based on variability of multilocus RAPD markers

The method of polymerase chain reaction with random primers (RAPD PCR) was used to identify the progeny of the crosses between three sturgeon species, Amur sturgeon (Acipenser schrenckii Brandt, 1869), Siberian sturgeon (A. baerii Brandt, 1869), and sterlet (A. ruthenus Linnaeus, 1758). Using ten primers, genetic variation in 70 yearlings, produced in seven individual crosses: Acipenser schrenckii × A. schrenckii, A. baerii × A. baerii, A. ruthenus × A. ruthenus, A. schrenckii × A. baerii, A. baerii × A. schrenckii, A. schrenckii × A. ruthenus, and A. ruthenus × A. schrenckii was described and evaluated. It was demonstrated that the samples composed of hybrids from individual crosses were more variable than the samples of parental species. On the other hand, pooled samples of hybrids from two cross directions were genetically less variable than the pooled samples of their parents. The three main features of the hybrid RAPD profiles identified included: (1) preservation of marker DNA fragments of both parents in one genome; (2) presence of specific DNA fragments, absent from both parents; and (3) dependence of the frequency of some DNA fragments from the cross direction. Multidimensional scaling clearly distinguishes in the space of three coordinates the individuals of original species and the hybrid progeny with differentiation in the groups of direct and backcross hybrids. Analysis of relationships (UPGMA and NJ) pointed to substantial differentiation between the species, as well as between the species and hybrid progeny. Close genetic relationships between direct and backcross hybrids were demonstrated. Multilocus RAPD markers in association with statistical methods are considered to be the useful tool for discrimination of interspecific hybrids of sturgeon. Possible reasons for the differences in the hybrid RAPD profiles are discussed. Original Russian Text ? K.V. Rozhkovan, G.N. Chelomina, E.I. Rachek, 2008, published in Genetika, 2008, Vol. 44, No. 11, pp. 1453–1460.     

LOSS OF HYBRID LETHALITY DURING BACKCROSS PROGRAMS INVOLVING CHLAMYDOMONAS EUGAMETOS AND CHLAMYDOMONAS MOEWUSII (CHLOROPHYCEAE)1

Wild-type strains of the interfertile species Chlamydomonas eugametos (UTEX 9 and 10) and Chlamydomonas moewusii (UTEX 96 and 97) male readily and reciprocally; however, considerable lethality occurs among F₁ hybrid meiotic products. We prepared two hybrid backcross lineages using C. eugametos and C. moewusii. One lineage began with the cross C. eugametos mating-type-plus (mt⁺) × C. moewusii mating-type-minus (mt^?). An F₁ mt⁺ hybrid from this cross was back-crossed to C. moewusii mt^?, and a B₁ mt⁺ hybrid was recovered. The B₁ hybrid was again backcrossed to C. moewusii mt^?, and this process was repeated through the fifth backcross. The other backcross lineage began with the reciprocal cross C. moewusii mt⁺× C. eugametos mt^? and employed C. eugametos as the recurring mt^? parent. This lineage also was continued through the fifth backcross. Meiotic product survival in the reciprocal interspecific crosses was less than 10%. In successive back-cross generations associated with both lineages, this value increased progressively to a maximum of 85–90%, the level observed for the intraspecific crosses. These results are consistent with the hypothesis that multiple genetic differences exist between C. eugametos and C. moewusii and that these are the major source of meiotic product lethality associated with the interspecific crosses. The inheritance of chloroplast genetic markers for resistance to streptomycin (sr-2) and for resistance to erythromycin (er-nM1) was also scored w the interspecific crosses and in the backcrosses. Most hybrid zygospores transmitted the resistance markers of the mt⁺ parent only, or of both parents, with the former zygospore type being more common. Although the intraspecific C. eugametos and C. moewusii crosses differ conspicuously with respect to the fraction of zygospores which transmit chloroplast genetic markers of both parents, the inheritance of chloroplast genetic markers in the interspecific crosses and backcrosses at' scribed here failed to clarify the genetic basis for this difference.     

Transgressive character expression in hybrid zones between the native invasives Tithonia tubaeformis and Tithonia rotundifolia (Asteraceae) in Mexico

Natural hybridization frequently promotes gene introgression among closely related species in sympatric populations, producing complex patterns of morphological variation. Therefore, a detailed understanding of the dynamics of interspecific gene flow and its morphological patterns is of widespread interest. We tested if introgressive hybridization promotes an increase in transgressive characters in comparison with the parental species. A sunflower species complex occurring in Mexico formed by two native invasive species, Tithonia tubaeformis and Tithonia rotundifolia, was analyzed using 46 morphological characters (leaf, flower and fruit) in five hybrid zones (N = 150 individuals) and two pure sites for each parental species (N = 80 individuals). In general, T. tubaeformis differed significantly from T. rotundifolia in all the examined characters, except six foliar and one inflorescence character. Morphological characters support the hypothesis of hybridization in this complex, even though both species remain morphological distinct in mixed stands. Individual hybrids appear to be a mosaic of parent-like (24.8 % of traits), intermediate (26.1 %) and transgressive (37.8 %) phenotypes (the remaining 11.3 % of the traits did not differ significantly from both parental species). Our results suggest that individuals from the same parental species were more similar among themselves than to putative hybrids, indicating occasional hybridization with segregation in hybrid types or backcrossing to parents. Evidence indicates a unidirectional pattern of gene flow toward T. rotundifolia.     

CROSSING STUDIES IN CYPHOMANDRA (SOLANACEAE) AND THEIR SYSTEMATIC AND EVOLUTIONARY SIGNIFICANCE

A crossing program involving eight species of Cyphomandra was instituted to elucidate systematic relationships between the taxa and evolutionary mechanisms operating in the genus. The results show that gametophytic self-incompatibility is widespread in Cyphomandra. Pollen tubes were able to reach the ovules in nearly all interspecific crosses. Most of these crosses failed, however, indicating that postzygotic barriers are important in reproductive isolation. Only five of 44 interspecific combinations attempted have produced viable seeds. F₁ plants have been evaluated from three of the five hybrid combinations: C. betacea × acuminata, C. uniloba × betacea, and C. corymbiflora × diploconos. All three hybrid combinations show reduced fertility as compared to the parents, apparently caused in part by differences in chromosome structure between parent species. Crossing success was congruent with morphological similarity in C. betacea, C. acuminata, and C. uniloba, but not in C. corymbiflora and C. diploconos. External isolating mechanisms such as geography, ecology, phenology, and pollinator specificity may also operate to maintain species integrity in nature.     

Further Evidence for a Polymorphism in Gametic Segregation in the Tetraploid Treefrog HYLA VERSICOLOR Using a Glutamate Oxaloacetic Transaminase Locus

Intra- and interspecific cross combinations between the tetraploid treefrog Hyla versicolor, and between H. versicolor and the diploid treefrog Hyla chrysoscelis were performed. Progeny phenotypes resulting from these crosses were examined electrophoretically using a polymorphic glutamate oxaloacetic transminase (GOT-1) locus, to determine the mechanism of chromosome segregation in H. versicolor, and to test theoretical expectations for isozyme expression in interspecific (2n x 4n or 4n x 2n) hybrids. In some intraspecific tetraploid crosses progeny phenotypes fit a disomic mode of segregation, whereas in other crosses a tetrasomic mode of segregation was the most probable. Additional crosses produced phenotypic ratios that conformed to either a disomic or tetrasomic mode of segregation. These results suggest that a polymorphism, with respect to segregation of gametes, exists in H. versicolor, resulting from differences in chromosome pairings during meiosis I. This polymorphism in gametic segregation occurred in both sexes. Certain crosses, however, produced phenotypic ratios that did not conform to any chromosome segregation model. Progeny phenotypes observed from most interspecific crosses conformed to expected interspecific isozyme staining intensity models. Symmetrical heterozygotes, representing either a single dose for both alternate alleles or double doses for both alternate alleles, were also observed. Such phenotypes are unexpected in triploid progeny. A null allele was postulated to account for the aberrant segregation ratios and phenotypes observed in certain intra- and interspecific crosses.     

Self-compatible sympatric Chamaecrista (Leguminosae-Caesalpinioideae) species present different interspecific isolation mechanisms depending on their phylogenetic proximity

The mating system of seven sympatric taxa of Chamaecrista occurring in the Chapada Diamantina Mountains, northeastern Brazil, was studied to determine the occurrence of self-incompatibility and interspecific genetic isolation mechanisms within the group. Self- and cross-pollination experiments and inter-taxon bidirectional crosses were performed. All of the populations were self-compatible and showed high percentages of spontaneous seed abortion on both self- and cross-pollinated fruits. The inter-incompatibility among the taxa of Chamaecrista is directed by different mechanisms depending on their degrees of phylogenetic proximity. In the crosses between closely related taxa (same clade), seed inviability was observed in the crossing pairs C. desvauxii var. graminea × C. desvauxii var. latistipula, and C. chapadae × C. glaucofilix. Inter-compatibility between species of the same clade occurred in C. blanchetii and C. confertiformis, with the formation of viable seeds. Pre-zygotic gametophytic reproductive isolation was observed among taxa of different clades, without penetration of the pollen tube into the ovule, or post-zygotic isolation through embryo or endosperm inviability, with abortion of the seeds. Inter-incompatibility represents an important factor in reproductive isolation and thus in the maintenance of the genetic identity of sympatric taxa that flower in synchrony and share pollinators.