Microspore embryogenesis induced through in vitro anther culture of almond (Prunus dulcis Mill.)

Anther culture is one of the most widely used methods to induce gametic embryogenesis. The aim of this investigation was to induce microspore embryogenesis in almond (Prunus dulcis Mill.), through this technique. Anthers were cultured at the vacuolated developmental stage, and seven cultivars, two culture media and two temperature treatments were assessed. Although evidence of the microspore induction was observed in all the genotypes and treatments tested (symmetrical nucleus division and multinucleated structures), calli were produced merely by anthers cultured in the medium P and the regeneration of embryos was detected only in anthers of the cultivars Filippo Ceo, Lauranne and Genco, pl…

Nutraceutical Value of Pantelleria Capers (Capparis spinosa L.)

Abstract: Unopened flower buds of Capparis spinosa L. (capers), generally used in the Mediterranean area as food flavoring, are known to be a good source of bioactive compounds. The aim of this work was to evaluate the nutraceutical value of salt-fermented capers collected from different areas of Pantelleria Island (Italy), testing their methylglyoxal and glyoxal trapping capacity and antioxidant activity by 2,2-diphenyl-1-picryl hydrazyl (DPPH), [2,2-azinobis(3-ethylben- zothiazoline-6-sulfonic acid)] diammonium salt (ABTS), and oxygen radical absorbance capacity (ORAC) assays. Hydrophilic extracts were also characterized by high-performance liquid chromatography–electrospray ionization/ma…

Microspore embryogenesis inCitrusand other fruit crops

Conventional methods, involving several generations of selfing, are not applicable to produce homozygous lines in Citrus as well as in the other fruit crops, due to the high heterozygosity of the genomes, the long duration of the generation cycle, the large size, and, often, the self-incompatibility. For this reason, there is no different way to obtain homozygosity in this kind of plants than “gametic embryogenesis” that allows the development of haploids (Hs, plants with gametophytic chromosome number) and doubled haploids (DHs, haploids that have undergone chromosome duplication) from heterozygous parents in a single step. Therefore, gametic embryogenesis is increasingly object of res…

EX VITRO MYCORRHIZATION OF VITRO-DERIVED PLANTLETS OF 'CARRIZO' CITRANGE [C. SINENSIS (L.) OSB. × P. TRIFOLIATA (L.) RAF.]

In this study, vitro-derived plantlets of ‘Carrizo’ citrange were inoculated with Glomus intraradices during the acclimatization phase, from in vitro to in vivo. Plantlets were grown in presence (+M) and in absence of inoculum (-M) and in addition, in sterile and non-sterile substrate. After three months of culture, some vegetative parameters and the level of infection, varied greatly in the different theses. In particular, the highest formation of new branched roots has been observed in inoculated plantlets cultivated in sterile substrate. Good results were observed also in non-inoculated plantlets cultivated in non-sterile substrate, in presence of endemic inoculum. The same trend was…

First stages of microsporere programming to embryogenesis through anther culture in Prunus armeniaca L.

6 páginas, 3 figuras, 2 tablas -- PAGS nros. 152-157

Olive embryo in vitro germination potential: Role of explant configuration and embryo structure among cultivars

The in vitro germination of excised embryos can break dormancy rapidly and shorten the time required to produce seedlings, speeding up olive breeding programmes as well as rootstock production. In this study, the in vitro germination potential of four Sicilian olive cultivars was evaluated during two years of experiments, using explants with three different morphological configurations that represent three different degrees of embryo exposure: (1) intact stoneless seeds containing the embryo, the endosperm and the seed coat (Emb+En+SC), (2) seeds without the seed coat (Emb+En) and (3) naked, isolated embryos (seed coat and endosperm both removed: Emb). Differences were found in the germinat…

Anther culture in Citrus clementina: a way to regenerate tri-haploids

Abstract. Regenerants from anther culture of Citrus clementina Hort. ex Tan. cvv. Nules, SRA 63, and Monreal were obtained in different experiments from 1994 to 2002. Genetic analysis of 37 such regenerants was carried out using 4 microsatellite markers that were heterozygous in the parental genotypes. The results showed that in all cases but one the regenerants carried only one or the other allele of the parental genotype, and were therefore homozygous and produced through a process of gametophytic embryogenesis. Ploidy analysis by flow cytometry of 94 regenerants showed that as many as 82% of them were tri-haploids, rather than haploids or doubled-haploids as expected, with other ploidy …

Encapsulation of black mulberry microcuttings: studies on capsules and synthetic seeds

The encapsulation technology for plant tissue culture can be an efficient tool to optimize the plant management systems and to preserve valuable plant germplasm. In this paper, two experiments were carried out to evaluate the encapsulation efficiency in mulberry (Morus nigra L.). In the first experiment, gel capsules were stored at 4°C for different times (0, 30, 90, 180 and 360 days). The highest percentage of shoot viability (82.5%) and regrowth (72.5%) was scored after 180 days of storage, with an average development of 2.5 shoots/capsule. The second experiment was aimed to point out an efficient protocol to turn the capsules of black mulberry in synthetic seeds ready to develop into who…

Somatic embryogenesis through in vitro anther culture ofCitrus sinensisL. Osbeck ‘Moro’

In many crops, anther culture is the most used method to induce gametic embryogenesis, aimed to regenerate homozygous plants. However, also somatic embryogenesis can be obtained by this method, when somatic tissue is involved in regeneration process. Many factors can affect this procedure, such as genotype, pre-treatments applied to floral buds, pollen developmental stage, donor plant states, culture media composition and growth culture conditions. Anthers of Citrus sinensis L. Osbeck, cv. Moro, were collected at the vacuolate stage, and after a chilling (4°C) pre-treatment of 7 days, were placed on the same medium, evaluating different temperature stresses applied after the placing in cult…

Seasonal variations of antimicrobial activity and chemical composition of essential oils extracted from three Citrus limon L. Burm. cultivars

In order to investigate the seasonal variations of antimicrobial properties and chemical composition of essential oils (EOs), three different cultivars of Citrus limon L. Burm. spp. (Femminello Santa Teresa, Monachello and Femminello Continella) were collected at 6-week intervals, from December 2012 to April 2013, for a total of four harvests. The EOs were extracted from lemon peel by hydro-distillation. The antimicrobial activity, tested by paper disc diffusion method, was evaluated against common food-related pathogenic bacteria (Listeria monocytogenes, Staphylococcus aureus, Salmonella enterica and Enterobacter spp.). EOs were more effective against Gram-positive than Gram-negative bacte…

Preliminary identification of self-incompatibility genotypes of Sicilian almond landraces

Sicily is the main almond producer in Italy. The almond tree is one of the main protagonists of the Sicilian agrarian landscape, especially in the winter, when the bare branches are covered by white or pinkish flowers. Its presence is particularly significant in the province of Agrigento, in Syracuse and in Ragusa areas, where the almond nut is used for fresh consumption and for the preparation of typical sweets. The extremely rich Sicilian almond germplasm can be useful for genetic improvement, since many local landraces possess aromatic characteristics of the nuts of high value. Self-incompatibility in this species is gametophytic and controlled by a multi-allelic S locus, coding for the …

Cryopreservation of Hazelnut (Corylus avellana L.) Axillary Buds from In Vitro Shoots Using the Droplet Vitrification Method

Cryopreservation by droplet vitrification was applied to hazelnut (Corylus avellana L.). axillary buds of the Italian cultivated variety Tonda Gentile Romana, which were collected from in vitro growing shoots, immersed in ice cooled PVS2 or PVS3 for 60 or 90 min, then transferred to a droplet of vitrification solution, placed on a strip of aluminium foil, and plunged into liquid nitrogen (LN). Additionally, the effect on the recovery of the mother plant after cryopreservation was evaluated, following a cold pre-treatment at 4 °C for 3 months. The highest regrowth percentage (56.7%) was obtained after applying PVS3 for 60 min, while the application of PVS2 for the same amount of time reduced…

FIRST STAGES OF MICROSPORE REPROGRAMMING TO EMBRYOGENESIS THROUGH ISOLATED MICROSPORE CULTURE IN LOQUAT

The current experiments were undertaken to develop a method for regenerating doubled haploid (DH) plants of Eriobotrya japonica (Thunb.), Rosaceae, subfamily Maloideae through isolated microspore culture of nine loquat cultivars. Protocols were developed for isolation and culture of loquat microspores. Isolated microspores in vitro cultured started dividing and developing multicellular, globular and irregular structures. After 4 weeks of culture, the responding microspores produced yellowish/white callus. The embryogenic response of microspores was highly dependent on the cultivar and on the medium composition.

Inhibition of foodborne pathogen bacteria by essential oils extracted from citrus fruits cultivated in Sicily

Abstract The antagonistic activity of the essential oils (EOs) extracted by hydrodistillation from the fruit peel of several citrus genotypes (pummelo, grapefruit, orange, kumquat, mandarin and lemon) was evaluated against foodborne pathogen bacteria (43 strains of Listeria monocytogenes , 35 strains of Staphylococcus aureus and 14 strains of Salmonella enterica ). Five commercial EOs were used for comparison. Most of the EOs were more effective against the Gram-positive bacteria rather than Salmonella . EOs of lemon genotypes 14 and 15 showed the best results in terms of number of strains inhibited and width of the inhibition zone. The most susceptible strain of each species ( L. monocytog…

Micropropagation of Citrus spp. by Organogenesis and Somatic Embryogenesis.

Citrus spp., the largest fruit crops produced worldwide, are usually asexually propagated by cuttings or grafting onto seedling rootstocks. Most of Citrus genotypes are characterized by polyembryony due to the occurrence of adventive nucellar embryos, which lead to the production of true-to-type plants by seed germination. Tissue culture and micropropagation, in particular, are valuable alternatives to traditional propagation to obtain a high number of uniform and healthy plants in a short time and in a small space. Moreover, in vitro propagation provides a rapid system to multiply the progeny obtained by breeding programs, allows the use of monoembryonic and seedless genotypes as rootstock…

FRUIT QUALITY EVALUATION OF FOUR LOQUAT CULTIVARS GROWN IN SICILY

Chemical (soluble solids content, titratable acidity, pH), morphological (weight, longitudinal and transversal diameter, thickness of the flesh, number of seeds, peel colour), and sensorial characteristics of four loquat cultivars (‘Algerie’, ‘Claudia’, ‘Nespolone di Trabia’ and ‘Sanfilippara’) from mature, organically-grown trees in Sicily, were analysed. The determinations were carried out by traditional instrumental techniques, by a panel test and by an electronic olfactory system. A panel of 10 trained judges was used to determine intensity of some attributes in the sensory profile of the different cultivars, while the electronic olfactory system was used to identify possible difference…

Induction of pollen embryogenesis in seven cultivars of Prunus armeniaca l.

6 páginas -- PAGS nros. 273-278

Doubled haploid production in fruit crops

The interest of fruit breeders in haploids and doubled haploids (DH), lies in the possibility of shortening the time needed to produce homozygous lines compared to conventional breeding. Haplo-diploidization through gametic embryogenesis allows single-step development of complete homozygous lines from heterozygous parents. In a conventional breeding programme, a pure line is developed after several generations of selfing. With fruit crops, characterized by a long reproductive cycle, a high degree of heterozygosity, large size, and, sometimes, self-incompatibility, there is no way to obtain haploidization through conventional methods. This paper reviews the current status of research on doub…

Two-dimensional electrophoresis analysis of proteins extracted from leaves of homozygous and heterozygous citrus

Microspore Embryogenesis in Almond (Prunus dulcis Mill.)

Almond is the most important tree nut crop in terms of commercial production. Its production is limited to areas characterized by a Mediterranean climate, including regions in the Mediterranean countries, the Central Valley of California, Central Asia, the Himalayan slopes and some equivalent areas in the Southern Hemisphere, including Argentina, Australia and South Africa (Kester et al. 1975).

Viruses infecting different Mediterranean genotypes of Ficus carica and their distribution in different plant organs.

Seven Mediterranean Ficus carica genotypes, i.e. cv. Palazzo, Severoni precoce, Bianca, Pilusedda, Dottato bianco, Bifara and Zidi, were screened for the presence of seven fig-infecting viruses associated with fig mosaic disease (FMD) in order to explore their distribution in different plant organs (leaf, bud and syconium) that will be utilized as a plant source material in different sanitation techniques. RT-PCR assays conducted on reverse-transcribed TNA extracted from leaves, apical buds and syconia (1.5-2cm) of each genotype for the presence of Fig leaf mottle-associated virus 1 (FLMaV-1), Fig mild mottling-associated virus (FMMaV), Fig mosaic virus (FMV), Fig latent virus 1 (FLV-1), Fi…

Monitoring the Shelf-Life of Minimally Processed Fresh-Cut Apple Slices By Physical Chemical Analysis and Electronic Nose

Fresh-cut apples, in slices or in cubes, are minimally processed products, which are currently collecting a great interest by fruit marketers for their promising diffusion. Their shelf life, from a microbiological point of view, has been fixed about 2 or 3 weeks under refrigeration. However in a few days they undergo biochemical degradations with production of off-flavors and texture breakdown. In this work, the change of aromatic fingerprint of apple slices packaged in air and in a modified atmosphere (with 100% N2) and stored at 4°C was measured, by using a commercial electronic nose. The obtained data were also compared with sensory evaluation of judge’s panel. Moreover, quality paramete…

OCCURRENCE OF SPONTANEOUS TRIPLOIDS FROM SMALL SEEDS OF 'HERNANDINA' CLEMENTINE

Micropropagation ofAgeratum houstonianumby nodal segments

Ageratum houstonianum is a bedding and flowering potted plant originated from Central America which is generally propagate by seed. In this report a preliminary in vitro technique for propagation of A. houstonianum was investigated. In vitro germinated seeds were used to establish aseptic shoot cultures of several clones. Seedling stem segments bearing 3-4 nodes were placed on Murashige and Skoog (MS) basal medium plus 20 g L-1 sucrose, 8.0 g L-1 Agar to induce axillary shoot development. Axillary shoots were subcultured into the same medium and nodal segments were sectioned and subcultured to increase the stock of shoot cultures. Shoot cultures of the selected clone AG14 were used to accom…

Gametic embryogenesis and haploid technology as valuable support to plant breeding

Plant breeding is focused on continuously increasing crop production to meet the needs of an ever-growing world population, improving food quality to ensure a long and healthy life and address the problems of global warming and environment pollution, together with the challenges of developing novel sources of biofuels. The breeders' search for novel genetic combinations, with which to select plants with improved traits to satisfy both farmers and consumers, is endless. About half of the dramatic increase in crop yield obtained in the second half of the last century has been achieved thanks to the results of genetic improvement, while the residual advance has been due to the enhanced managem…

STUDY ON CONVERSION OF ENCAPSULATED VITRO-DERIVED PROPAGULES OF 'C35' CITRANGE [C. SINENSIS (L.) OSB. × P. TRIFOLIATE (L.) RAF.]

Fight against Citrus tristeza virus (CTV) implies the use of CTV-resistant rootstocks, such as citranges [C. Sinensis (L.) Osb. × P. Trifoliata (L.) Raf.]. The 'C35' represents a promising tolerant to CTV genotype. The employment of innovative in vitro techniques, such as encapsulation, could open new perspectives for this genotype propagation, particularly in the nursery activity. Preliminary experiments on calcium-alginate encapsulation of microcuttings (unipolar propagules) have already been performed to evaluate the use of tissue culture for the propagation and the preservation of different citrange rootstocks: Carrizo and Troyer. In the present study, the influence of different types a…

Fig virus-free production and survival rate improvement using meristem tip culture techinique associated with the encapsulation technology

Three Mediterranean F. carica genotypes, i.e. cultivars Palazzo, Bifera nera and Catalanisca, initially infected by Fig leaf mottle-associated virus 1 (FLMaV-1), Fig leaf mottle-associated virus 2 (FLMaV-2), Fig mild mottling-associated virus (FMMaV), Fig mosaic virus (FMV), Fig latent virus 1 (FLV-1), Fig Badnavirus 1 (FBV-1) and Fig fleck-associated virus (FFkaV), were subjected to the sanitation technique via Meristem Tip (0.3-0.5 mm in size) Culture (MTC), also associated with the encapsulation technique (MTC-SS), in order to produce virus-free plant material. Encapsulation was tested to overcome the very low survival and regeneration rates, due to the small propagule size. Encouraging …

Gametic and somatic embryogenesis through in vitro anther culture of different Citrus genotypes

Abstract: In vitro tissue culture represents a useful technique for advancing Citrus breeding and propagation. Among in vitro regeneration systems, anther culture is commonly used to produce haploids and doubled haploids for a fast-track producing homozygous lines, in comparison with the traditional self-pollination approach, which involves several generations of selfing. In addition, anthers culture can produce somatic embryos that can also be used for clonal propagation. In this study, two thermal shocks were applied to the anthers of six Citrus genotypes (two clementine and four sweet oranges), just after they were put in culture. The response obtained was different depending on the geno…

Ploidy levels in Citrus clementine affects leaf morphology, stomatal density and water content

The objective of the present study was to understand the relationship among leaf morphology, stomatal characteristics and water relations in triploids generated through anther culture and their counterpart diploid plant of C. clementina. Triploid plants possessed small and narrow leaves as compared to diploid plant as evident by less leaf length, leaf width and leaf area. By contrast, the leaf index was observed to be more in triploids than haploid ones. Flow cytometric analysis re-confirmed the ploidy levels of heterozygous plant Hd as diploid and the ploidy of Th1, Th2, Th3 and Th4 plants as triploids. A positive relation was found between ploidy level and stomatal guard cell length and w…

Characterization of Leaf Essential Oil Composition of Homozygous and HeterozygousCitrus clementinaHort. Extan. and its Ancestors

Clementine is a natural tangor, resulting from an interspecific cross between mandarin and sweet orange. Gametic embryogenesis, allowing the single-step development of complete homozygous line from the heterozygous parents, increases the efficiency of perennial crop breeding programs. Tri-haploids have been regenerated through pollen embryogenesis (specifically, by anther culture) of Citrus clementina Hort. ex Tan., cv. Nules. Two of them (HOMO1 and HOMO2) have been acclimatized and grafted in vivo in 2000. Research regarding the chemical characteristics of plant regenerated by pollen embryogenesis or gynogenesis are useful to study the “gametoclonal variation”, as well as for further appli…

In vitro anther culture of sweet orange (Citrus sinensis L. Osbeck) genotypes and of a C. clementina × C. sinensis 'Hamlin' hybrid

Citrus, and particularly sweet oranges, are very recalcitrant to anther culture. In this paper it was evaluated for the first time the response of 27 genotypes of Citrus sinensis and of one hybrid C. clementina × C. sinensis, to in vitro anther culture. Ten genotypes of sweet oranges showed embryogenic callus induction, mostly blood sweet oranges genotypes, such as Tarocco, Moro and Sanguinelli. In vitro microspore developmental switches from the gamethophytic to the sporophytic pathway were shown by DAPI staining in microspores of these responsive genotypes, after 10 months in culture. However, microsatellite marker analyses showed that these calli were heterozygous. The flow-cytometric an…

Development of new non-dairy beverages from Mediterranean fruit juices fermented with water kefir microorganisms

The aim of this work was to explore the use of several Mediterranean fruit juices as fermentable substrates to develop new non-dairy fermented beverages. Microbiological, chemical and sensory features of kefir-like beverages obtained after the fermentation of juices extracted from fruits cultivated in Sicily (southern Italy) with water kefir microorganisms were investigated. Results indicated that both lactic acid bacteria and yeasts were able to develop in the fruit juices tested, but the highest levels were registered with prickly pear fruit juice. All fruit juices underwent a lactic fermentation, since a lactic acid content was detected in the resulting kefir-like beverages. Except kiwif…

Production of Haploid and Doubled Haploid Lines in Nut Crops: Persian Walnut, Almond, and Hazelnut

This chapter deals with induction of haploidy via parthenogenesis in Persian walnut and via microspore embryogenesis in almond and hazelnut. Haploid induction through in situ parthenogenesis using pollination with irradiated pollen to stimulate the embryogenic development of the egg cell, followed by in vitro culture of the immature haploid embryos. Microspore embryogenesis allows the induction of immature pollen grains (microspores), to move away from the normal gametophytic developmental route in the direction of the sporophytic one, yielding homozygous organisms (embryos in this case). Unlike other fruit crops (such as Citrus), regeneration of entire plants has not yet been obtained in o…

Organogenesis and encapsulation of in vitro-derived propagules of Carrizo citrange [Citrus sinensis (L.) Osb. × Poncirius trifoliata (L.) Raf]

Due to widespread polyembryony, Citrus rootstocks are usually propagated by open-pollinated seed germination, although micropropagation offers many advantages. Encapsulation technology has recently attracted the interest of researchers in the field of plant propagation because it combines the advantages of zygotic or gamic seeds with those of micropropagation. In this study, we examined the encapsulation of Carrizo citrange uninodal microcuttings (3–4 mm long) and evaluated the influence of the calcium alginate coating, a short time storage at cold temperature, and different sowing substrates on the viability and regrowth of the explants. A secondary aim was to develop an efficient protocol…

Free and conjugated polyamine content in Citrus sinensis Osbeck, cultivar Brasiliano N.L.92, a Navel orange, at different maturation stages

Biogenic amines, synthesized during physiological metabolic processes of all living organisms, are present in food. At low concentrations, polyamines are essential for cell renewal and growth, but they can be detrimental when consumed in high amount through the diet as they could support abnormal cell growth pathologies. The daily human diet contains more putrescine than spermidine or spermine, mostly derived from fruits. In general, orange fruits contain high levels of put, a fact that could limit their utilization in the daily diet besides the benefits contributed by their strong antioxidant properties. There is therefore an increasing interest in finding plant foods with low polyamine co…

Effect of polyamines on in vitro anther culture of Citrus clementina Hort. ex Tan

The improvement of the induction rate in Citrus anther culture is important for taking practical advantage of the haploid potential in breeding. The influence of polyamines on anther culture of Citrus clementina, cv Nules, with particular attention to the free, soluble and insoluble-conjugated polyamine levels, has been investigated. Putrescine, spermidine and putrescine plus spermidine, were added to the standard induction medium. Before culture, spermidine was the most abundant among the free polyamines detected in anthers. The exogenous supply of either putrescine or spermidine, either independently or combined, effected greater uptake and accumulation of polyamines. The addition of 2 mM…

Encapsulation of in vitro-derived propagules of two genotypes of Capparis spinosa (L.) from Pantelleria Island

Encapsulation technology, combining the advantages of zygotic or gamic seed with those of micropropagation, has recently attracted the interest of researchers as a new propagation approach, mainly due to the unsatisfying results of the traditional propagation strategies. The encapsulation of uninodal microcuttings (3-4 mm long) from two Sicilian Capparis spinosa (L.) genotypes (from Pantelleria Island: 'Tracino' and 'Scauri') was evaluated, observing the influence of the calcium alginate coating and of three different growth regulators (PGRs): 6-benzylaminopurine (BAP), meta-topolin (MT) and zeatin (ZEA), on viability, regrowth and conversion of the propagules. Caper microcuttings were diss…

Application of MOOSY32 eNose to Assess the Effects of Some Post Harvest Treatments on the Quality of ‘Salustiana’ Orange Juice

A new prototype of Electronic Nose instrument, Multisensory Odor Olfactory System MOOSY32, with a processing method based on a multivariate classification analysis was used to assess different postharvest and storage treatments effects to Salustiana oranges. The analysis method is based on the measurement of the volatile compounds produced under different environmental and operational conditions. The Electronic Nose system revealed that orange juice flavor changes even when juices are analyzed right after each treatment and fruits are stored under refrigerated conditions. The instrument was able to detect even small changes in the aromatic pattern of the juices, confirming that the packing …

Characterization of fruits of four different lemon cultivars, Collected in the northern coast of sicily

The present work is focused on the study of the characteristics of the fruits of four main lemon (Citrus limon L. Burm.) varieties cultivated in Sicily: 'Femminello Comune', 'Monachello', 'Zagara Bianca', and 'Femminello Santa Teresa'. Physical and chemical properties as well as aroma compounds were analysed as quality discrimination factors. Also the effect of the storage conditions was verified. Standard experimental techniques were used to determine: weight, longitudinal and transverse diameters, pH, titratable acidity, total soluble solids, juice percentage and total polyphenols. The volatile component was analysed by i) a gas chromatographer with a mass spectrometer detector and ii) an…

Preliminary Results on the Effect of Magnetic Fields on Anther Culture and Pollen Germination of Citrus clementina Hort. ex Tan.

Exposure to electromagnetic fields seems to modify the biological behaviour of seeds, roots, pollen grains and buds of several plants. This procedure has been also suggested as a tool to improve the utilization of plant species (Firetto et al., 2000, 2001, Petruszewski S., 1996). Since the first haploid embryogenic calli production and plantlet regeneration by anther culture of C. clementina Hort. ex Tan. 'Nules' was reported (Germany et al., 1994), several studies have been carried out to improve both the frequency of microspore embryogenesis and the percentage of plantlet regeneration. To enlarge the number of respondent genotypes and to improve the induction rate (the frequency of pollen…

Effects of tetraploidy on olive floral and fruit biology

Floral biology and fruit development were studied in Leccino Compact (LC), a polyploid olive mutant of cultivar Leccino (L). This mutant, considered a mixoploid with both diploid and tetraploid cells, has thicker leaves and fruit size similar to the diploid cultivar. So far, no information is available on its floral biology. In this study, the ploidy level of the LC fruit epicarp, analyzed by flow cytometry, was determined to be tetraploid. Pollen size distribution confirmed that most flowers were tetraploid. Morphological and histological measurements of various floral structures and fruits were carried out on the two genotypes, and LC showed larger floral structures (i.e. rachis, flower a…

Micropropagation of Sicilian cultivars with an aim to preserve genetic diversity in hazelnut (Corylus avellana L.)

The use of a small number of cultivars in agriculture can lead to a loss of agrobiodiversity. Since in vitro techniques are valuable tools for conserving plant biodiversity, an efficient micropropagation protocol for four Italian hazelnut cultivars, ‘Carrello’, ‘Ghirara’, ‘Minnulara’, and ‘Panottara’, was developed. The highest axillary bud survival was obtained after decontamination with 40 min 1% sodium hypochlorite followed by 40 min 0.1% sodium merthiolate in ‘Minnulara’ and ‘Ghirara’, while the 35þ35 min treatment was the best for ‘Carrello’ and ‘Panottara’. Shoot multiplication was higher in ‘Minnulara’ and ‘Ghirara’ when 6.6 lM N6-benzyladenine was used, even if some hyperhydric shoo…

Microspore Embryogenesis in Citrus

This chapter deals with microspore embryogenesis in Citrus. Microspore embryogenesis allows to induce immature gametes (microspores) and to deviate them, in this case, the male one, from the normal gametophytic developmental route in the direction of the sporophytic one, yielding homozygous organisms (embryos and plants).

Current and Potential Use of Citrus Essential Oils

Since the Middle Ages, citrus essential oils (EOs) have been widely used for their bactericidal, virucidal, fungicidal, antipara- sitical, insecticidal, medicinal and cosmetic proprieties. Also nowadays, they find important applications in pharmaceutical, sanitary, cosmetic, agricultural and food industries. The best method to extract EOs from citrus plant tissue is steam distillation because of a vari- ety of extracted volatile molecules such as terpenes and terpenoids, phenol-derived aromatic components and aliphatic components. In vitro physicochemical assays classify most of them as antioxidants.

Gametic embryogenesis through isolated microspore culture in Corylus avellana L.

Haploid technology is a valuable plant breeding tool for obtaining homozygosity particularly in woody plants. Hazelnut, the world’s sixth ranking nut tree crop is a monoecious, anemophilous species. It is characterized by a sporophytic incompatibility system that prevents production of homozygous plants with conventional methods, involving several self-pollination cycles. In this study, gametic embryogenesis, in particular isolated microspore culture, was tried with five genotypes. Two culture media were tested and four temperature stress treatments were applied to the isolated microspores that were cultivated at the vacuolated developmental stage. To our knowledge, this is the first report…

Gametic embryogenesis through isolated microspore culture in mandarin (Citrus reticulata Blanco), Mandarino Tardivo Di Ciaculli: effect of meta-Topolin and temperature treatments

Haploid technology, allowing the single-step development of complete homozygous genotypes from heterozygous plants through gametic embryogenesis, has already an enormous impact on breeding programs of many important crops. Microspore embryogenesis can be carried out through in vitro culture of anther or isolated microspore. Experiments have been carried out via isolated microspore culture in Citrus reticulata (Blanco), Mandarino Tardivo di Ciaculli, investigating the influence of two culture media, of two plant growth regulator types and of two temperature treatments applied before the culture or just after the culture. After ten months of culture, for all the media tested, different struct…

Morphogenic responsein vitroof epicotyl segments ofCitrus macrophylla

Abstract The influence of explant orientation, contact of the cut surface with the medium, type of growth regulators and distance of epicotyl cuttings from the cotyledonary node (the part of the seedling where the cotyledons are attached) on in vitro morphogenesis of Citrus macrophylla (alemow) was evaluated. Maximum number of shoots was induced from epicotyl segments placed horizontally.

Genetic Structure of Wild and Cultivated Olives in the Central Mediterranean Basin

Background and Aims: Olive cultivars and their wild relatives (oleasters) represent two botanical varieties of Olea europaea subsp. europaea (respectively europaea and sylvestris). Olive cultivars have undergone human selection and their area of diffusion overlaps that of oleasters. Populations of genuine wild olives seem restricted to isolated areas of Mediterranean forests, while most other wild-looking forms of olive may include feral forms that escaped cultivation. Methods: The genetic structure of wild and cultivated olive tree populations was evaluated by amplified fragment length polymorphism (AFLP) markers at a microscale level in one continental and two insular Italian regions. Key…

Development of multicellular pollen of Eriobotrya japonica Lindl. through anther culture

8 pages, 5 figures.-- Printed version published Dec 2006.

Ploidy manipulation and citrus breeding, genetics and genomics

Polyploidy appears to have played a limited role in citrus germplasm evolution. However, today, ploidy manipulation is an important component of citrus breeding strategies. For varieties, the main objective is to develop triploid seedless cultivars. For rootstock, the aim is to cumulate interesting traits in tetraploid hybrids and to improve adaptation to biotic and abiotic stresses. This chapter starts with a review of the recent knowledge acquired on the natural mechanisms of citrus polyploidization and tetraploid meiosis. Chromosome doubling of nucellar cells is frequent in apomictic citrus and results in tetraploid seedling production. Unreduced gametes are also frequently produced, mai…

In vitro response of two Sicilian genotypes of Morus (L.) through axillary bud culture.

A protocol for the in vitro establishment of two Sicilian genotypes of mulberry has been set up. Multiple shoots were initiated and plantlets were produced from buds collected from mature trees of Morus alba L. and Morus nigra L., grown in the field. The influence of two explanting times (September and December) of buds on the in vitro response has been observed. The percentage of sprouting after four weeks of culture was greatly higher with material explanted from the field in September compared with the one collected in December. Morus nigra L. produced more vigorous shoots than Morus alba L.. After the second subculture the number of shoots produced decreased, in both genotypes. No diffe…

Early in vitro development of isolated microspores of four Sicilian olive cultivars.

Nowadays there is a great interest in the advancement of the studies on gametic embryogenesis in olive, as a method for obtaining homozygous material, due to the enormous effort and relevance in its genome sequencing and gene mapping. In this study, in vitro isolated microspore culture of four Sicilian olive cultivars (‘Biancolilla napoletana’, ‘Nocellara del Belice’, ‘Tonda Iblea’ and ‘Verdello’) has been carried out, investigating the influence of two polyamines (putrescine and spermidine) and of two medium compositions on pollen embryogenesis induction. During in vitro development, different structural features have been observed: uninucleated microspores, with no development, binucleate…

Antilisterial effect of citrus essential oils and their performance inedible film formulations

[EN] The antimicrobial activity of eight essential oils (EOs) extracted from the fruit peel of Citrus genotypes (orange, mandarin and lemon) was evaluated against 76 strains of Listeria monocytogenes, previously isolated from different food matrices. EOs showing the most (EO L2 and EO L8) and least (EO O3 and EO M7) effective inhibition activities were chemically characterized by gas chromatography coupled with mass spectrometry (GC/MS) to compare their composition. EO L2 and EO L8 were chosen to determine the MIC and to evaluate the cell viability of the most sensitive strains (L. monocytogenes LM35 and LM69) after 1, 2, 4 and 6 h of exposure. The effectiveness of chitosan (CH) and methylc…

Doubled haploid production in onion (Allium cepa L.): from gynogenesis to chromosome doubling

Bulb onion (Allium cepa L.) is an allogamous diploid (2n = 16) important for its culinary uses, nutritional value, and medicinal benefits. Despite its economic importance, onion yields and bulb quality are declining, emphasizing the need for new and improved strategies for maintaining and enhancing overall crop quality. Development of inbred lines in onion through traditional breeding is often difficult due to its biennial life cycle, inbreeding depression, and comparatively high heterozygosities. Moreover, genetic research in onion has been hampered by large nuclear genome size. In this regard, gynogenic doubled haploids promise several advantages over inbred lines in support of onion bree…

Cytological and molecular characterization of three gametoclones of Citrus clementina

Abstract Background Three gametoclonal plants of Citrus clementina Hort. ex Tan., cv. Nules, designated ESP, FRA, and ITA (derived from three labs in Spain, France, and Italy, respectively), were selected for cytological and molecular characterization in order to elucidate genomic rearrangements provoked by haploidization. The study included comparisons of their ploidy, homozygosity, genome integrity, and gene dosage, using chromosome counting, flow cytometry, SSR marker genotyping, and array-Comparative Genomic Hybridization (array-CGH). Results Chromosome counting and flow cytometry revealed that ESP and FRA were haploid, but ITA was tri-haploid. Homozygous patterns, represented by a sing…

Protocol of Somatic Embryogenesis from Citrus spp. Anther Culture

Microspore Embryogenesis Through Anther Culture in Citrus clementina Hort. ex Tan.

Anther culture is a biotechnological method that allows to obtain, in one step, homozygous plants, very important to plant breeding, due to their numerous applications in mutation research, selection, genome sequencing, genetic analysis, and transformation. To induce the microspores, i.e., the immature male gametes, to switch from the normal gametophytic pathway to the sporophytic one, it is necessary to submit them to a type of stress, such as high or low temperature, starvation, or magnetic field. Stress can be applied to the donor plants and/or the fl oral buds or the anthers or the isolated microspores, before or during the culture. In this chapter, the protocol to induce gametic embryo…

Early embryo achievement through isolated microspore culture in citrus clementina hort. Ex tan., cvs. 'Monreal Rosso' and 'nules'

10 p.-5 fig.-3 tab.

Morphological and anatomical observations of abnormal somatic embryos from anther cultures of Citrus reticulata

A morphological and anatomical study of regenerants obtained from mandarin anther culture was carried out. Beside morphologically normal somatic embryos, abnormal structures were originated in the course of somatic embryogenesis. Anatomical anomalies can be found at several growth stages, causing the formation of slender stems, stubby structures, non-functional leaves. When too long, some structures are subject to shedding, with the formation of various abscission zones. Most of them are subject to degeneration, although many are capable of further, localized, morphogenesis. A thorough knowledge of morphology and anatomy of normal and abnormal regenerants could make possible to select and s…