Monitoring and cellular characterisation of male gametic embryogenesis in Citrus clementina.

Correlation of sequentila floral development and male gametophyte development and preliminary results on anther culture in Opuntia ficus-indica. Acta Physiologiae Plantarum Vol. 27 No. 4B:

STUDY ON ISOLATED MICROSPORE CULTURE IN CITRUS SINENSIS L. OSBECK CV. MORO, A BLOOD ORANGE CULTIVAR

In vitro tissue culture represents a useful support for the advancement of Citrus breeding and propagation. Haploidy technology, that is the single-step development of complete homozygous genotypes from heterozygous parents through gametic embryogenesis, has already a huge impact on many relevant crops, representing an integral part in their breeding programmes (Germanà 2011a; 2011b). In order to increase the number of genotypes responding to gametic embryogenesis, in vitro isolated microspore culture of Citrus sinensis L. Osbeck cv. Moro, has been carried out, investigating the influence of the culture medium composition, and, particularly, of the plant growth regulator types and concentra…

CHARACTERISATION OF MICROSPORE EMBRYOGENESIS IN ANTHER CULTURE OF CITRUS.

Analisi dei trascritti caratterizzanti il processo di maturazione in ficodindia (Opuntia ficus-indica L.)

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

Studio preliminare sulla germinazione in vitro di semi di Argania spinosa (L.)

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…

Effects of high and low intensità magnetic fields on pollen behaviour of Citrus clementina Hort. ex Tan. and Pinus pinea L.

Primeras etapas de la embriogenesis de microsporas en coltivo in vitro de anteras de Citrus clementina Hort. ex Tan.

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.

Anther and isolated microspore culture in the fruit tree species Prunus avium L .

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…

The effects of meta-Topolin and benzyladenine on in vitro organogenesis from epicotyl cuttings of Troyer citrange (Citrus sinensis[L.] Osbeck ×Poncirus trifoliata[L.] Raf.)

Citrus tristeza virus (CTV), worldwide considered as one of the worst disease affecting Citrus plants, produced great changes in citriculture, mainly because it made mandatory the turnover of the sensitive sour orange rootstock with virus resistant ones. Among them, Troyer citrange has become one of the most employed rootstocks, due to its CTV tolerance and to the positive characteristics induced to the scion fruit production. Unfortunately, in Italy, there is a scarcity of seed mother plants. Micropropagation can be a solution for this problem. Among the in vitro techniques allowing the production of good quality true-to-type plants, the regeneration from seedling epicotyl cuttings general…

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

6 páginas -- PAGS nros. 273-278

Preliminary results on anther culture in Opuntia ficus-indica.

ISOLATION AND CHARACTERIZATION OF RNA FRACTIONS INVOLVED IN THE MATURATION PROCESS IN OPUNTIA FICUS-INDICA (L.) MILL.

FIRST EMBRYONIC STAGES OF CITRUS MICROSPORE DERIVED EMBRYOS.

Studio sulla germinabilità in vitro di semi di Fraxinus angustifolia Vahl.

Studio sull'embriogenesi gametica in Olea europaea L.

ISOLATED MICROSPORE CULTURE IN THE FRUIT TREE SPECIES PRUNUS AVIUM, ERIOBOTRYA JAPONICA, OLEA EUROPAEA AND OPUNTIA FICUS-INDICA.

Observations on the effects of high and low intensity magnetic fields on pollen behaviour of Citrus Clementina Hort.ex Tan.and Pinus pinea L.

Gametic embryogenesis in Citrus

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

In Vitro Plant Regeneration from Epicotyls Segments of Carrizo Citrange.

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…

Gametic and somatic embryogenesis through anther culture in Citrus.

Preliminary results on the effect of light quality on anther culture of Citrus clementina.

GAMETIC EMBRYOGENESIS IN CITRUS.

Microspore-derived embryo recovery trough isolated microspore culture in Corylus avellana L.

Studio preliminare sulla coltura in vitro di due genotipi siciliani di Morus.

Observations on anatomical structures of in vitro germination of stoneless and isolated olive embryos.

Development of multi-cellular pollen of Eriobotrya japonica through anther culture.

Caratterizzazione molecolare di un mutante di Olea europaea L. a taglia ridotta.

Studio sul potenziale morfogenico di segmenti di epicotile di Citrus macrophylla Wester.

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…

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…

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…

Anther culture in Citrus:a powerful tool to recover seedless triploids

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…

Coltura in vitro di antere per l'ottenimento di embriogenesi gametica e somatica in Citrus

Study on anther culture in hazelnut (Corylus avellana L.) .

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…

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.

Stabilizzazione e coltura in vitro di due genotipi siciliani di olivo (Olea europaea L.)

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

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

ANTHER CULTURE TECHNIQUE TO OBTAIN GAMETIC AND SOMATIC EMBRYOGENESIS IN CITRUS.

Studio sulla coltura in vitro in Punica granatum L.

Correlation of sequential floral and male gametophyte development in Opuntia ficus-indica.

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…

First stages of microspore reprogramming to embryogenesis through anther culture in Prunus armeniaca L

Correlation of sequential floral development and male gametophyte development and preliminary results on anther culture in Opuntia ficus-indica.

In vitro establishment and culture of several Sicilian olive genotypes (Olea europaea L.)

Analisis genetico de regenerantes de Citrus reticulata Blanco obtenidos por coltivo in vitro de anteras.

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…

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.

Study on the effect of polyamines on in vitro anther culture of Citrus clementina Hort. ex Tan.

Use of gametic embryogenesis for Citrus improvement.

Study on flavonoid contents in fruits of different Citrus genotypes