Pollination with laser-irradiated pollens breaks cross-incompatibility between zicaitai (Brassica campestris var. purpurea) and ornamental kale (Brassica oleracea var. acephala) to produce hybrids with the aid of ovule culture

The objective of this study was to produce interspecific hybrids between an Ogura-cytoplasmic male sterile (CMS) line of zicaitai (Brassica campestris var. purupurea, 2n = 20) and cultivars of ornamental kale (Brassica oleracea var. acephala, 2n = 18) to develop a CMS system for hybrid seed production. Pollination with pollen grains of ornamental kales irradiated at a power output of 9.0 mW with a He–Ne laser for 3 min could overcome the cross-incompatibility between the species concerned. Intact hybrids could be efficiently produced from ovules cultured on Murashige and Skoog media supplemented by 0.2 mg l⁻¹ 6-benzyladenine. Chromosome number of hybrids was confirmed to be 2n = 19. Hybrids resembled ornamental kales in leaf morphology and in vernalization response. Pollens of hybrids had a sterile appearance. Moreover the hybridity of the putative hybrids was confirmed by RAPD data on a DNA fragment of 820 bp.     

Regeneration and characterization of plants derived from leaf in vitro culture of two sweet orange (Citrus sinensis (L.) Osbeck) cultivars

In the current work attempts were made to investigate culture of leaf explants derived from in vitro seedlings of two sweet orange (Citrus sinensis (L.) Osbeck) cultivars, Bingtangcheng and Valencia. Effects of several factors, including culture medium, lighting condition, explant age and genotype on regeneration response were examined based on three parameters, percentage of explants producing shoots, mean number of shoots per explant and shoot forming capacity. Culture of the explants on shoot-inducing media (SIM) composed of MT salts supplemented with different growth regulators gave rise to disparate shoot regeneration, in which SIM₁ (MT + 0.5 mg L⁻¹ BA + 0.5 mg L⁻¹ Kinetin + 0.1 mg L⁻¹ NAA + 3% sucrose + 0.8% agar, pH 5.8) was shown to be the most effective medium for direct induction of shoots from leaf explants. Highly significant difference in the response of shoot bud regeneration was noted between the two cultivars, with Bingtangcheng being more responsive than Valencia. Culture of explants from fully developed leaves led to better shoot regeneration capacity in comparison to undeveloped ones. However, the two lighting conditions used herein did not cause significant difference in shoot regeneration. Phenotypic observation and randomly amplified polymorphic DNA (RAPD) analysis confirmed that all the regenerated plants from both genotypes were genetically identical to their donor plants, suggesting absence of detectable genetic variation in the regenerated plants. The data presented here demonstrated that direct initiation of plants from leaf explants has been successfully accomplished. To our knowledge, this is the first report on direct regeneration of shoots from leaf explants in Citrus, which will provide an alternative source for citrus genetic manipulation in the future.     

Production of intraspecific hybrids between wild-type and petaloid-sepal cultivars in Habenaria radiata

Orchids are commercially important plants with flowers that are unique and very specialized in shape and color. The flowers consist mostly of sepals, lateral petals, lip (labellum) and column, and are zygomorphic and resupinate. Whereas most orchid species have petaloid tepals in the first and second whorls, Habenaria radiata has a flower with greenish sepals and white lateral petals and lip. ‘Hishou’, one of the cultivars of H. radiata, is a floral homeotic mutant and has a petaloid median sepal and lip-like lateral sepals in the first whorl. Additionally, this cultivar often has non-resupinate flowers whereas wild-type H. radiata flowers are resupinate. In the present study, we investigated the genetic inheritance of these characters in the ‘Hishou’ cultivar by crossing it with wild-type plants. Some intraspecific hybrids, which were confirmed by PCR-RFLP analysis, had flowers with a petaloid median sepal and lip-like lateral sepals in the first whorl, indicating that these were dominant characters. Since the remainder of the intraspecific hybrids had wild-type flowers, these characters must be heterozygous in ‘Hishou’ plants. Although ‘Hishou’ plants had non-resupinate flowers, intraspecific hybrid flowers were resupinate, even though they had the petaloid median sepal and lip-like lateral sepals. This result indicates that non-resupination must be a recessive character. Since sepal-petalization and triple lip characters of ‘Hishou’ inherited dominantly, these characters can be utilized for the breeding of Habenaria species by intra- and interspecific crosses.     

Characterization of tetraploid plants regenerated via protoplast culture of Iris fulva and their crossability with Japanese irises

Characteristics such as flower form, size and color of outer and inner perianths, anthocyanins in outer perianths, size, color and fertility of pollen and self-fertility of diploid and tetraploid lines regenerated via protoplast culture of Iris fulva were examined and compared with those of the diploid wild line. Among these characteristics, flower form, inner and outer perianth sizes of the tetraploid lines were noticeable, because these lines had upward flower forms and bigger flowers than diploid lines. Furthermore, reciprocal crosses between diploid or tetraploid lines of I. fulva and I. ensata and those of I. fulva and I. laevigata were performed. Three seedlings were obtained from the cross of tetraploid I. fulva × diploid I. laevigata through embryo rescue. One of them was identified as the interspecific hybrid between tetraploid I. fulva and I. laevigata by flow cytometoric (FCM), cytological and molecular (RAPD) analyses. This is the first report on production of hybrids from these lines. I. fulva has unique brown flowers, and this trait could be very useful for flower color breeding of I. laevigata which lacks this color. Therefore, the hybrid of I. fulva (4×) × I. laevigata may be the best available gene source for brown color breeding of this species.     

Bioinformatically mined simple sequence repeats in UniGene of Citrus sinensis

Characterization of microsatellites is extremely important for the development of molecular markers. Here, we present the detection and abundance of microsatellites or simple sequence repeats (SSRs) in UniGene sequences of Citrus sinensis. A total of 427 SSRs were mined in 8786 UniGene sequences downloaded from National Center for Biotechnology Information (NCBI). Depending on the repeat units, the length of SSRs ranged from 14 to 21 for mono-, 14 to 48 for di-, 18 to 48 for tri-, 24 to 40 for tetra- and 42 bp for hexa-nucleotide repeats. Average density of SSRs (1SSR/12.92 kb of 5518.71 kb sequences mined) suggests that only 4.43% of sequences contained SSRs. Di-nucleotide repeats were most frequent repeat type (49.41%) followed by tri-nucleotide repeats (41.45%). An attempt was made to design primer pairs for 427 identified SSRs but these were found only for 216 sequences. The positions of SSRs with respect to open reading frame (ORF) detected and annotation of sequences containing SSRs were also carried out to assign function to each of the sequences.     

H2O2 metabolism during sweet orange (Citrus sinensis L. Osb.) ‘Hamlin’ Xanthomonas axonopodis pv. citri interaction

Sweet orange (Citrus sinensis L. Osb.) ‘Hamlin’ is a canker (Xanthomonas axonopodis pv. citri: Xac) susceptible citrus genotype grown commercially worldwide. Canker causes severe economic losses and restricts the marketability of crop for export. Little is known about the role of oxidative stress in canker development. In the present investigation, sweet orange ‘Hamlin’ leaves were artificially inoculated with Xac to determine the impact of Xac infection on hydrogen peroxide (H₂O₂) metabolism. Characteristic symptoms following artificial inoculation were water soaking of the infiltrated zone between 2 and 8 days after inoculation (dai); raised epidermis accompanying tiny yellow colored bacterial colonies at 8 dai; and yellowing and necrosis of the infected zone by 12–16 dai. In planta Xac population increased 1000 fold by 14 dai from an initial population of 7.3 × 10⁶ cfu cm⁻² (0 dai). Peak concentrations of H₂O₂ were observed at 24 h and between 8 and 10 dai and coincided with higher activity of total superoxide dismutase (SOD). Lower levels of H₂O₂ in infected leaves were maintained by Xac induced higher activities of catalase (CAT), ascorbate peroxidase (APOD), and guaiacol peroxidase (POD). It appears Xac altered H₂O₂ metabolism in C. sinensis L. Osb. ‘Hamlin’ to enhance survival and growth.     

Confirmation of Clematis hybrids using molecular markers

The hybrid origin of progeny from crosses of Clematis tubulosa and Clematis brevicaudata was investigated using randomly amplified polymorphic DNA (RAPD) and single nucleotide polymorphisms (SNPs) from sequence analysis of chloroplast rbcL, accD genes, and the C. brevicaudatamatK gene. Plants collected from three and four populations of C. brevicaudata (C. brev) and C. tubulosa (C. tubu), respectively, from Mt. Songshan, Beijing, China were used as parents for hybridization. Morphological characters of pollen, seeds, and leaves were recorded in 2007. DNA from leaf samples of both parents and of C. brev × C. tubu and C. tubu × C. brev were extracted, and used for RAPD and SNPs from sequence data. A dendrogram was constructed by the branching neighbor-joining (IB-NJ) method. Proportionate population scores were generated by the admixture model using the STRUCTURE software. Based on morphological characters, C. brevicaudata was quite uniform. However, variations were detected in C. tubulosa. Hybrids of C. brev × C. tubu and C. tubu × C. brev showed intermediate morphological characters of the parents. Accessions of C. tubu × C. brev were clustered into 2 groups, with the majority of hybrids belonging to group IV b in the RAPD dendrogram, suggesting that this resulted from variations within C. tubulosa. In general, the hybrid origin of all progeny characterized by morphological characters was supported by the RAPD and SNPs data. These results indicate that RAPD results supported by SNPs data will be useful tool to verify hybrids.     

Development of ITS sequence based SCAR markers for discrimination of Paphiopedilum armeniacum, Paphiopedilum micranthum, Paphiopedilum delenatii and their hybrids

Paphiopedilum armeniacum, Paphiopedilum micranthum and Paphiopedilum delenatii are endangered orchid species. These three Paphiopedilum species and their hybrids are difficult to distinguish morphologically. In this study, rDNA-ITS (internal transcribed spacer) sequences were used to design species-specific SCAR (sequence characterized amplified regions) markers to distinguish P. armeniacum, P. micranthum, P. delenatii and their respective hybrids. The developed markers efficiently amplified 600 bp DNA product for P. armeniacum and its hybrids (SCAR-600armF/Pap-ITS2R), 300 bp product for P. delenatii and its hybrids (SCAR-300delF/Pap-ITS2R) and 700 bp product for P. micranthum and its hybrids (SCAR-700micF/Pap-ITS2R). The effectiveness of designed species-specific markers was also confirmed by using multiplex polymerase chain reaction amplification with a combination of developed three SCAR markers.     

Phenotypic variability and genetic structure in plum (Prunus domestica L.), cherry plum (P. cerasifera Ehrh.) and sloe (P. spinosa L.)

In the present study, phenotypic variability of 80 plum (Prunus domestica L.) varieties maintained in the French National Plum Collection was evaluated with 19 quantitative traits. In addition, genetic diversity and genetic structure was studied in three plum species (P. domestica L., Prunus cerasifera Ehrh. and Prunus spinosa L.) using chloroplast DNA (cpDNA) markers and five single sequence repeat (SSR) loci. Based on phenotypic traits, some varieties, such as mirabelle plums, grouped together. Bayesian structure analysis was used to identify different genetic groups, whereby damson plums were clearly distinguished from greengage plums. When examining the three species together, a higher level of cpDNA allelic richness was found in P. cerasifera and in P. spinosa than in P. domestica where only five cpDNA haplotypes were detected in the national plum collection, with one main haplotype that accounted for 80% of the varieties studied. P. domestica cpDNA haplotypes tended to group together with P. cerasifera haplotypes whereas most of P. spinosa haplotypes formed a separate cluster. SSR markers were somewhat able to distinguish the three species. These results provide some clues as to the origin of plum and the various plum varieties. Our results also provide useful information for the management of plum genetic resources.     

The culture of isolated microspores of ornamental kale (Brassica oleracea var. acephala) and the importance of genotype to embryo regeneration

The culture of isolated microspores of kale (Brassica oleracea var. acephala) was studied including the importance of genotype to embryo regeneration, medium composition chiefly the sucrose concentration and the use of colchicine, simultaneously medium renovation. It was initiated using 29 different genotypes as donor plants. Embryos were induced from six of the kale genotypes and these corresponded to the more out-bred genotypes. Embryogenesis was achieved using four different combinations of culture media: (a) microspores initially cultured in NLN medium supplemented with 13% (w/v) sucrose (NLN-13) for 48 h, followed by transfer to fresh NLN-13 medium; (b) microspores cultured for 48 h in NLN-13 medium supplemented with colchicines (50 mg/L) followed by transfer to unsupplemented NLN-13 medium; (c) microspores cultured for 48 h in NLN-16 medium supplemented with colchicines (50 mg/L) followed by transfer to unsupplemented NLN-16 medium; (d) microspores cultured for 48 h in NLN-16 medium supplemented with colchicines (50 mg/L) followed by transfer to unsupplemented NLN-13 medium. The embryos obtained from four of the genotypes developed into plantlets and these regenerated plants have been successfully transplanted to soil.     

Production protocol development for greenhouse cut Linaria, Lupinus, and Papaver flowers

Linaria maroccana Hook. f. Ann., ‘Lace Violet’, Lupinus hartwegii ssp. cruikshankii Lindl. ‘Sunrise’ and Papaver nudicaule L. ‘Meadow Pastels’ seeds were directly sown into 105 cell plug trays and received either ambient light or supplemental high intensity discharge (HID) lighting. For each species, a 2 × 3 × 3 factorial was used with two light intensities during propagation, three transplant stages, and three night temperatures. Seedlings were transplanted at the appearance of 2–3, 5–6, or 8–9 true leaves. Transplanted Linaria and Papaver seedlings were placed at 5/11, 10/16, or 15/21 ± 1 °C night/day temperatures and Lupinus seedlings were placed at 15/24, 18/25, or 20/26 ± 2 °C night/day temperatures. For this study, the optimum production temperature for Linaria was 10/16 °C as the cut stems produced at 15/21 °C were unmarketable and production time was excessively long at 5/11 °C. At 10/16 °C, Linaria seedlings should be transplanted at the 2–3 leaf stage to maximize stem number, stem length and profitability. For Lupinus the optimum temperature was 15/24 °C due to long stems and high profitability per plant. Lupinus seedlings should be transplanted at the 2–3 leaf stage when grown at 15/24 °C to obtain the longest and thickest stems; however, $/m² week was higher for plants transplanted at the 8–9 leaf stage due to less time in finishing production space. For Papaver, the 15/21 °C temperature was optimal as that temperature produced the longest stems in the shortest duration, resulting in the highest $/m² week. At 15/21 °C Papaver plants should be transplanted at the 2–3 leaf stage. Supplemental HID lighting had no effect on any of the species.     

Clonal propagation of Rosa clinophylla Thory. through axillary bud culture

A protocol for clonal propagation of Rosa clinophylla, a rare and endangered species but very important for breeding purposes had been standardized through in vitro axillary bud culture. Although cytokinins alone were able to induce shoot buds, but their proper growth and number could be increased only when they were used in combination with GA₃. However, there was shoot tip necrosis and leaf fall in the proliferated shoots. AgNO₃ at 58.85 μM proved effective to avoid shoot necrosis and yellowing of leaves. Activated charcoal (AC) at 250 mg l⁻¹ was found necessary at all the stages of shoot multiplication as well as rooting. Ninety percent rooting could be achieved in 1/2 MS medium supplemented with 4.92 μM IBA and 250 mg l⁻¹ AC. Rooted plantlets were hardened and transferred to the field successfully with 80% survival rate.     

Chemical compositions and insecticidal effects of essential oils isolated from Achillea gypsicola, Satureja hortensis, Origanum acutidens and Hypericum scabrum against broadbean weevil (Bruchus dentipes)

The essential oils of aerial parts of Achillea gypsicola Hub.-Mor., Hypericum scabrum L., Satureja hortensis L., and Origanum acutidens (Hand.-Mazz.) Letswaart were analyzed in this study by GC and GC–MS and their oils were tested for toxicity against broadbean weevil (Bruchus dentipes). A. gypsicola oil contained camphor (40.17%), 1,8-cineole (22.01%), piperitone (11.29%), borneol (9.50%) and α-terpineol (1.56%) as major components. A total of 74 components were identified by GC–MS in H. scabrum oil, including α-pinene (9.26%), terpinen-4-ol (5.12%), camphor (5.94%), δ-cadinene (4.52%), pulegone (4.45%), γ-muurolene (4.12%), pinocarvone (3.97%) and β-caryophyllene (3.42%) as predominant components. The essential oils of O. acutidens and S. hortensis were characterized by high contents of carvacrol (86.99% and 55.74%), γ-terpinene (0.71% and 20.94%), p-cymene (1.95% and 12.30%), α-terpinene (0.13% and 2.04%) and β-caryophyllene (1.30% and 1.08%). All of the essential oils were toxic to adults of B. dentipes and insect mortality increased with increasing concentration of each oil. The oils (20 μl dose) brought about 100% mortality in 36 h. Although desirable insecticidal activities against the pest were achieved with the oils from all four plant species, S. hortensis and O. acutidens oils were more effective, particularly after 6 h of treatment. The current results concluded that the essential oils, in particular O. acutidens and S. hortensis oils, may be used as potential botanical insecticides against B. dentipes.     

6-Benzylamino purine stimulates in vitro shoot organogenesis in Eucalyptus erythronema, E. stricklandii and their interspecific hybrids

In vitro bud and shoot organogenesis was investigated for the ornamental plants Eucalyptus erythronema var. erythronema, E. stricklandii and their interspecific hybrids cv. ‘Urrbrae Gem’ and ‘Hybrid 2.5’ by using 0.0, 0.1, 0.25, 0.5 or 1.0 μM BAP on apex and leaf explants. Callus developed on all explants and increased with all concentrations of BAP without significant differences between BAP concentrations. Buds formed on apex and leaf explants of E. erythronema and E. cv. ‘Urrbrae Gem’ especially with 1.0 μM BAP, but these buds rarely developed into shoots. Bud clusters formed on E. erythronema and E. cv. ‘Urrbrae Gem’ apex and leaf explants whereas E. stricklandii and ‘Hybrid 2.5’ produced fewer, individual buds on the explant. Shoots regenerated from apex explants of all genotypes with all levels of BAP, whereas few shoots of any genotype regenerated from leaf explants regardless of the number of buds formed. Shoots from apex explants could be multiplied successfully. Light microscopy showed meristems developed within the callus, and at the callus and bud surfaces. However, few shoots developed considering the level of bud and meristem formation. This report is the first for successful shoot organogenesis and multiplication in an ornamental eucalypt.     

Intergeneric hybridizations between Opisthopappus taihangensis and Chrysanthemum lavandulifolium

For the first time, reciprocal intergeneric hybridizations were produced between Opisthopappus taihangensis and Chrysanthemum lavandulifolium without emasculation. About 20% seed set was similarly obtained from reciprocal hybridization. Only 5 and 6 of the viable plants observed from 45 and 78 seedling survived present some conspicuous intermediate characteristics. Phenotypic evaluation among the progenies of the parents and the putative hybrids was performed carefully since an average of 5.3% seed set was produced in the type of self-pollination using pollen from the same flower and >10% seed set was similarly obtained in the types of self-pollination using pollen from different flowers in a plant and flowers in individual plants from different seeds. One individual of each hybrid shared the inflorescence habit with the pollen plant was confirmed further by the sequence of ncpGS. The two hybrids might be used as bridges of breeding of multi-generic hybrids.     

Ethylene influences development and flowering of Ptilotus spp. in vitro and ex vitro

The genus Ptilotus has immense potential for ornamental horticulture but its commercial development has been hindered by propagation limitations. Poor seed quality and germination are reported. Cutting propagation is limited by cutting supply as the juvenile phase of Ptilotus is short. Micropropagation has been used in an attempt to overcome these difficulties but explants become floral in vitro and this causes plantlets to elongate. Ethephon has been used to control flowering of stock plants of many ornamental species. This study investigated the effect of ethephon applied to young (3-week-old, deflasked from tissue culture) and mature (1-year-old) Ptilotus plants in a greenhouse. A system of applying gaseous ethylene at 0, 100, 200 and 300 mg l⁻¹ to the headspace of in vitro plantlets in glass jars was developed and the response of in vitro plantlets to ethylene studied. One-year-old Ptilotus plants were treated with 500 mg l⁻¹ ethephon 2 days before pruning or 1 or 2 weeks after pruning. Ethephon application 2 days before pruning decreased the number of inflorescences and increased the number of shoots (compared to the control) but was phytotoxic. Ethephon applications of 150 or 300 mg l⁻¹ applied weekly or fortnightly to 3-week-old plants deflasked from tissue culture reduced plant height and number of inflorescences and at low concentrations increased the number of new shoots. A fortnightly application at 150 mg l⁻¹ is recommended. Previous reports on the effects of ethylene on inflorescence production on plantlets in vitro are limited. Our study showed that exposure of in vitro plantlets of P. nobilis to ethylene gas at 100 mg l⁻¹ for 1 h significantly increased the number of shoots and plant height but this did not occur for plantlets of P. spicatus. Plantlets of P. spicatus exposed to transient ethylene at 200 and 300 mg l⁻¹ showed significantly greater rooting (52.4%) than the control (13.6%).     

Interspecific relationships of Lycoris (Amaryllidaceae) inferred from inter-simple sequence repeat data

Inter-simple sequence repeat (ISSR) markers were used to evaluate interspecific relationships of Lycoris. Twenty-four samples were included in the study representing a total of 20 among species and varieties. Thirteen primers produced 228 discernible DNA fragments, 205, or 89.91%, of which were polymorphic, indicating a high level of interspecific genetic variation in Lycoris. Our UPGMA cluster analysis recognized four major groups of species, which were consistent with morphological and karyotype observations. The first group included species with telocentric and metacentric chromosomes, while the second consisted of species with a haploid genome of 11 subtelocentric chromosomes. The third group contained species with a mixture of subtelocentric, telocentric and metacentric chromosomes. The last group included the Japanese and Korean species. Lycoris anhuiensis was clustered within accessions of Lycoris longituba, suggesting that it could be recognized as a variety of L. longituba. Our ISSR data also suggested that L. straminea may be a hybrid between Lycoris chinensis and Lycoris radiata var. pumila, and that L. caldwellii, an allotriploid, may be of a hybrid origin of L. chinensis and L. sprengeri.     

Application of treated wastewater for cultivation of roses (Rosa hybrida) in soil-less culture

No information is available today concerning the effect of irrigation with secondary treated sewage water on growth, production or quality of roses or other cut flowers. In the present study we investigated the effect of irrigation with treated sewage water on roses cultivated in two soil-less medium, perlite, an inert mineral medium and Choir (coconut fibers), an organic medium of high ion absorption capacity. During 12 months of exposure to the treated water, the visible appearance of the plants, their growth, the quantity and size of the flowering stems and their postharvest performance were not affected by the irrigation treatments. Contents of macroelements in the leaf tissues were unaffected by the irrigation with the secondary treated sewage water. At the same time, Cl contents increased 47% in perlite and 73% in Choir grown plants reaching levels characteristic of exposure to moderate salinity. Mn, Cu and B contents increased as well under cultivation in both perlite and Choir under irrigation with treated sewage water. On the other hand, contents of Fe, Zn, Mo and Al, were similar in all treatments. In all treatments contents of all the examined micro and macroelements were within the range accepted for proper plant function.     

Development of seedless and Mal Secco tolerant mutant lemons through budwood irradiation

Mal secco (caused by Phoma tracheiphila (Petri) Kantsch. and Gik.) is the most destructive fungal disease of lemon plantations worldwide and seedless lemons would be preferred by most consumers. Five dosage levels, 0, 3, 5, 7, and 9 kiloradian (krad), of cobalt (⁶⁰Co) gamma irradiation were applied to budstick of ‘Kutdiken’ lemon (Citrus limon (L.) Burm. f.) clone KT-2A. Mutations were stabilized in three vegetative generations. Three hundred fifty-eight and 478 M1V3 (mutation one and vegetation three) plants were evaluated for seed number and mal secco tolerance in the field and the greenhouse, respectively. LD₅₀ was around 5 krad gamma irradiation for ‘Kutdiken’ lemon. The seed number varied from 0 to 34 per fruit. The level of mal secco tolerance also varied significantly among the plants from 1.0 (no symptom) to 4.3 (high level of disease occurance). The stable seedless and mal secco tolerant plants were obtained from 5 and 7 krad irradiation: the three mutants from 5 krad irradiation gave more lemon-like fruits, while 7 krad irradiation caused altered tree morphology and early maturation of fruits. This study shows considerable potential for lemon cultivar improvement aiming to obtain seedless and mal secco tolerant lemons.     

Flower emasculation accelerates ovule degeneration and reduces fruit set in sweet cherry

Flower emasculation is commonly used to make flowers unattractive to pollinating insects and to carry out controlled pollinations. In sweet cherry, we have observed recurrent low fruit set after flower emasculation and compatible pollination without apparent causes. This led us to evaluate its effect on the progress of the reproductive phase and on fruit set in this species. Flower emasculation reduced by more than a half the fruit set obtained in crosses made during two consecutive years. This effect could be traced back to the first week after anthesis where weight increase of pistils from emasculated flowers was smaller and ovule degeneration was accelerated compared to pistils from non-emasculated flowers. Pollen tubes, which behaved similarly at the stigma-style level in emasculated and non-emasculated flowers, lost their directionality in the area close to the degenerated ovule in the ovary. While flower emasculation is valid to evaluate pollen tube performance in the style and to determine incompatibility relationships, the lower fruit set registered after emasculation alerts on its use in fruit set experiments and breeding programs.