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P S Testillano - One of the best experts on this subject based on the ideXlab platform.
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stress induced microspore embryogenesis by anther culture of quercus suber l
2018Co-Authors: P S Testillano, Beatriz Pintos, Aranzazu Gomezgaray, Mariacarmen RisuenoAbstract:In vivo, the microspore inside the anther divides and follows the gametophytic program to form the mature pollen grain. In vitro, upon the application of a stress treatment the microspore can be deviated towards a proliferation process leading to embryogenesis, the so-called microspore embryogenesis that can be induced in anther and isolated microspore cultures (Maluszynski et al. 2003).
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auxin biosynthesis accumulation action and transport are involved in stress induced microspore embryogenesis initiation and progression in brassica napus
Plant and Cell Physiology, 2015Co-Authors: Hector Rodriguezsanz, Mariateresa Solis, Mariacarmen Risueno, Mariafernanda Lopez, Aurelio Gomezcadenas, P S TestillanoAbstract:Isolated microspores are reprogrammed in vitro by stress, becoming totipotent cells and producing embryos and plants via a process known as microspore embryogenesis. Despite the abundance of data on auxin involvement in plant development and embryogenesis, no data are available regarding the dynamics of auxin concentration, cellular localization and the expression of biosynthesis genes during microspore embryogenesis. This work involved the analysis of auxin concentration and cellular accumulation; expression of TAA1 and NIT2 encoding enzymes of two auxin biosynthetic pathways; expression of the PIN1-like efflux carrier; and the effects of inhibition of auxin transport and action by N-1-naphthylphthalamic acid (NPA) and α-(p-chlorophenoxy) isobutyric acid (PCIB) during Brassica napus microspore embryogenesis. The results indicated de novo auxin synthesis after stress-induced microspore reprogramming and embryogenesis initiation, accompanying the first cell divisions. The progressive increase of auxin concentration during progression of embryogenesis correlated with the expression patterns of TAA1 and NIT2 genes of auxin biosynthetic pathways. Auxin was evenly distributed in early embryos, whereas in heart/torpedo embryos auxin was accumulated in apical and basal embryo regions. Auxin efflux carrier PIN1-like gene expression was induced in early multicellular embryos and increased at the globular/torpedo embryo stages. Inhibition of polar auxin transport (PAT) and action, by NPA and PCIB, impaired embryo development, indicating that PAT and auxin action are required for microspore embryo progression. NPA also modified auxin embryo accumulation patterns. These findings indicate that endogenous auxin biosynthesis, action and polar transport are required in stress-induced microspore reprogramming, embryogenesis initiation and progression.
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5 azacytidine promotes microspore embryogenesis initiation by decreasing global dna methylation but prevents subsequent embryo development in rapeseed and barley
Frontiers in Plant Science, 2015Co-Authors: Mariateresa Solis, Mariacarmen Risueno, Ahmedabdalla Eltantawy, Vanesa Cano, P S TestillanoAbstract:Microspores are reprogrammed by stress in vitro towards embryogenesis. This process is an important tool in breeding to obtain double-haploid plants. DNA methylation is a major epigenetic modification that changes in differentiation and proliferation. We have shown changes in global DNA methylation during microspore reprogramming. 5-Azacytidine (AzaC) cannot be methylated and leads to DNA hypomethylation. AzaC is a useful demethylating agent to study DNA dynamics, with a potential application in microspore embryogenesis. This work analyzes the effects of short and long AzaC treatments on microspore embryogenesis initiation and progression in two species, the dicot Brassica napus and the monocot Hordeum vulgare. This involved the quantitative analyses of proembryo and embryo production, the quantification of DNA methylation, 5mdC immunofluorescence and confocal microscopy, and the analysis of chromatin organization (condensation/ decondensation) by light and electron microscopy. Four days of AzaC treatments (2.5 µM) increased embryo induction, response associated with a decrease of DNA methylation, modified 5mdC and heterochromatin patterns compared to untreated embryos. By contrast, longer AzaC treatments diminished embryo production. Similar effects were found in both species, indicating that DNA demethylation promotes microspore reprogramming, totipotency acquisition and embryogenesis initiation, while embryo differentiation requires de novo DNA methylation and is prevented by AzaC. This suggests a role for DNA methylation in the repression of microspore reprogramming and possibly totipotency acquisition. Results provide new insights into the role of epigenetic modifications in microspore embryogenesis and suggest a potential benefit of inhibitors, such as AzaC, to improve the process efficiency in biotechnology and breeding programs.
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early embryo achievement through isolated microspore culture in citrus clementina hort ex tan cvs monreal rosso and nules
Frontiers in Plant Science, 2015Co-Authors: Benedetta Chiancone, Ivett Barany, Valeria Gianguzzi, Marines Marli Gniech Karasawa, A Abdelgalel, Daniela Torello Marinoni, Roberto Botta, P S Testillano, Maria GermanaAbstract:Microspore embryogenesis is a method of achieving complete homozygosity from plants. It is particularly useful for woody species, like Citrus, characterized by long juvenility, a high degree of heterozygosity and often self-incompatibility. Anther culture is currently the method of choice for microspore embryogenesis in many crops. However, isolated microspore culture is a better way to investigate the processes at the cellular, physiological, biochemical and molecular levels as it avoids the influence of somatic anther tissue. To exploit the potential of this technique, it is important to separate the key factors affecting the process and, among them, culture medium composition and particularly the plant growth regulators and their concentration, as they can greatly enhance regeneration efficiency. To our knowledge, the ability of meta-Topolin, a naturally occurring aromatic cytokinin, to induce gametic embryogenesis in isolated microspores of Citrus has never been investigated. In this study, the effect of two concentrations of meta-Topolin instead of benzyladenine or zeatin in the culture medium was investigated in isolated microspore culture of two genotypes of Citrus. After eleven months of isolated microspore culture, for both genotypes and for all the four tested media, the microspore reprogramming and their sporophytic development was observed by the presence of multinucleated calli and microspore-derived embryos at different stages. Microsatellite analysis of parental and embryo samples was performed to determine the embryo alleles constitution of early embryos produced in all tested media, confirming their origin from microspores. To our knowledge, this is the first successful report of Citrus microspore embryogenesis with isolated microspore culture in Citrus, and in particular in Citrus clementina Hort. ex Tan, cvs. ‘Monreal Rosso’ and ‘Nules’.
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early embryo achievement through isolated microspore culture in citrus clementina hort ex tan cvs monreal rosso and nules
Frontiers in Plant Science, 2015Co-Authors: Benedetta Chiancone, Ivett Barany, Valeria Gianguzzi, Marines Marli Gniech Karasawa, A Abdelgalel, Daniela Torello Marinoni, Roberto Botta, P S Testillano, Maria GermanaAbstract:Microspore embryogenesis is a method of achieving complete homozygosity from plants. It is particularly useful for woody species, like Citrus, characterized by long juvenility, a high degree of heterozygosity and often self-incompatibility. Anther culture is currently the method of choice for microspore embryogenesis in many crops. However, isolated microspore culture is a better way to investigate the processes at the cellular, physiological, biochemical, and molecular levels as it avoids the influence of somatic anther tissue. To exploit the potential of this technique, it is important to separate the key factors affecting the process and, among them, culture medium composition and particularly the plant growth regulators and their concentration, as they can greatly enhance regeneration efficiency. To our knowledge, the ability of meta-Topolin, a naturally occurring aromatic cytokinin, to induce gametic embryogenesis in isolated microspores of Citrus has never been investigated. In this study, the effect of two concentrations of meta-Topolin instead of benzyladenine or zeatin in the culture medium was investigated in isolated microspore culture of two genotypes of Citrus. After 11 months of isolated microspore culture, for both genotypes and for all the four tested media, the microspore reprogramming and their sporophytic development was observed by the presence of multinucleated calli and microspore-derived embryos at different stages. Microsatellite analysis of parental and embryo samples was performed to determine the embryo alleles constitution of early embryos produced in all tested media, confirming their origin from microspores. To our knowledge, this is the first successful report of Citrus microspore embryogenesis with isolated microspore culture in Citrus, and in particular in Citrus clementina Hort. ex Tan, cvs. 'Monreal Rosso' and 'Nules.'
Stefanie Pöggeler - One of the best experts on this subject based on the ideXlab platform.
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Sordaria macrospora : 25 years as a model organism for studying the molecular mechanisms of fruiting body development
Applied microbiology and biotechnology, 2020Co-Authors: Ines Teichert, Stefanie Pöggeler, Minou NowrousianAbstract:Fruiting bodies are among the most complex multicellular structures formed by fungi, and the molecular mechanisms that regulate their development are far from understood. However, studies with a number of fungal model organisms have started to shed light on this developmental process. One of these model organisms is Sordaria macrospora, a filamentous ascomycete from the order Sordariales. This fungus has been a genetic model organism since the 1950s, but its career as a model organism for molecular genetics really took off in the 1990s, when the establishment of a transformation protocol, a mutant collection, and an indexed cosmid library provided the methods and resources to start revealing the molecular mechanisms of fruiting body development. In the 2000s, "omics" methods were added to the S. macrospora tool box, and by 2020, 58 developmental genes have been identified in this fungus. This review gives a brief overview of major method developments for S. macrospora, and then focuses on recent results characterizing different processes involved in regulating development including several regulatory protein complexes, autophagy, transcriptional and chromatin regulation, and RNA editing. KEY POINTS: •Sordaria macrospora is a model system for analyzing fungal fruiting body development. •More than 100 developmental mutants are available for S. macrospora. •More than 50 developmental genes have been characterized in S. macrospora.
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MAT and Its Role in the Homothallic Ascomycete Sordaria macrospora
Sex in Fungi, 2014Co-Authors: Stefanie PöggelerAbstract:Mating-type genes have been identified not only in heterothallic and pseudohomothallic, but also in homothallic members of the Sordariaceae. This chapter provides a comprehensive and up-to-date overview of the mating-type genes and their roles in the homothallic ascomycete S. macrospora. The homothallic S. macrospora is the most studied species in the genus. Even though single-spore cultures of S. macrospora are self-fertile, they may be crossed. For example, when two S. macrospora strains are paired in a petri dish, each mycelium gives rise to perithecia and homokaryotic asci. Cloning and sequencing of the entire mating-type locus of S. macrospora revealed that sequences homologous to the MATA and MATa idiomorphs of Neurospora crassa are directly linked in the mating-type locus of S. macrospora. To elucidate how reproductive modes might have evolved in different members of the Sordariaceae, phylogenetic relationships of MATA- and MATa-specific mating-type genes from heterothallic and homothallic members of the genera Sordaria and Neurospora have been examined. The phylogenetic analyses suggest that changes in the reproductive strategy may represent a single event in each genus. Transcription of cell-type-specific genes in the ascomycete yeast S. cerevisiae has been shown to rely on the interaction mating-type proteins. Thus, heterodimerization provides a way of regulating transcription factor function by joining different functional domains.
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SmATG7 is required for viability in the homothallic ascomycete Sordaria macrospora.
Fungal genetics and biology : FG & B, 2009Co-Authors: Nicole Nolting, Yasmine Bernhards, Stefanie PöggelerAbstract:In filamentous ascomycetes, autophagy is involved in several developmental processes. Nevertheless, until now little is known about its role in fruiting-body development. We therefore isolated a gene of the homothallic ascomycete Sordaria macrospora with high sequence similarity to the Saccharomyces cerevisiae autophagy-related gene ATG7, encoding a core autophagy regulator. This is the first characterization of an ATG7 homolog in filamentous ascomycetes. A S. cerevisiae complementation assay demonstrated that the S. macrospora Smatg7 gene functionally replaces the yeast homolog. We were not able to generate a homokaryotic knock-out mutant in S. macrospora, suggesting that Smatg7 is required for viability. However, a heterokaryotic DeltaSmatg7/Smatg7 strain and transformants generated by RNA interference showed considerable morphological phenotypes during fruiting-body development. Using real-time PCR, we demonstrated that in the wild type, the transcriptional expression of Smatg7 is markedly up-regulated under amino acid starvation conditions and at late stages during sexual development. Moreover, we showed that transcriptionally down-regulation of Smatg7 disturbs autophagy in S. macrospora.
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functional characterization of an α factor like sordaria macrospora peptide pheromone and analysis of its interaction with its cognate receptor in saccharomyces cerevisiae
Eukaryotic Cell, 2005Co-Authors: Severine Mayrhofer, Stefanie PöggelerAbstract:The homothallic filamentous ascomycete Sordaria macrospora possesses genes which are thought to encode two pheromone precursors and two seven-transmembrane pheromone receptors. The pheromone precursor genes are termed ppg1 and ppg2. The putative products derived from the gene sequence show structural similarity to the α-factor precursors and a-factor precursors of the yeast Saccharomyces cerevisiae. Likewise, sequence similarity has been found between the putative products of the pheromone receptor genes pre2 and pre1 and the S. cerevisiae Ste2p α-factor receptor and Ste3p a-factor receptor, respectively. To investigate whether the α-factor-like pheromone-receptor pair of S. macrospora is functional, a heterologous yeast assay was used. Our results show that the S. macrospora α-factor-like pheromone precursor PPG1 is processed into an active pheromone by yeast MATα cells. The S. macrospora PRE2 protein was demonstrated to be a peptide pheromone receptor. In yeast MATa cells lacking the endogenous Ste2p receptor, the S. macrospora PRE2 receptor facilitated all aspects of the pheromone response. Using a synthetic peptide, we can now predict the sequence of one active form of the S. macrospora peptide pheromone. We proved that S. macrospora wild-type strains secrete an active pheromone into the culture medium and that disruption of the ppg1 gene in S. macrospora prevents pheromone production. However, loss of the ppg1 gene does not affect vegetative growth or fertility. Finally, we established the yeast assay as an easy and useful system for analyzing pheromone production in developmental mutants of S. macrospora.
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Mating-type genes from the homothallic fungus Sordaria macrospora are functionally expressed in a heterothallic ascomycete
Genetics, 1997Co-Authors: Stefanie Pöggeler, Ulrich Kuck, Siegfried Risch, Heinz D. OsiewaczAbstract:Homokaryons from the homothallic ascomycte Sordaria macrospora are able to enter the sexual pathway and to form fertile fruiting bodies. To analyze the molecular basis of homothallism and to elucidate the role of mating-products during fruiting body development, we cloned and sequenced the entire S. macrospora mating-type locus. Comparison of the Sordaria mating-type locus with mating-type idiomorphs from the heterothallic ascomycetes Neurospora crassa and Podospora anserina revealed that sequences from both idiomorphs ( A/a and mat –/ mat +, respectively) are contiguous in S. macrospora . DNA sequencing of the S. macrospora mating-type region allowed the identification of four open reading frames (ORFs), which were termed Smt-a1, SmtA-1, SmtA-2 and SmtA-3 . While Smt-a1, SmtA-1 , and SmtA-2 show strong sequence similarities with the corresponding N. crassa mating-type ORFs, SmtA-3 has a chimeric character. It comprises sequences that are similar to the A and a mating-type idiomorph from N. crassa . To determine functionality of the S. macrospora mating-type genes, we show that all ORFs are transcriptionally expressed. Furthermore, we transformed the S. macrospora mating-type genes into mat – and mat + strains of the closely related heterothallic fungus P. anserina . The transformation experiments show that mating-type genes from S. macrospora induce fruiting body formation in P. anserina .
Ivett Barany - One of the best experts on this subject based on the ideXlab platform.
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early embryo achievement through isolated microspore culture in citrus clementina hort ex tan cvs monreal rosso and nules
Frontiers in Plant Science, 2015Co-Authors: Benedetta Chiancone, Ivett Barany, Valeria Gianguzzi, Marines Marli Gniech Karasawa, A Abdelgalel, Daniela Torello Marinoni, Roberto Botta, P S Testillano, Maria GermanaAbstract:Microspore embryogenesis is a method of achieving complete homozygosity from plants. It is particularly useful for woody species, like Citrus, characterized by long juvenility, a high degree of heterozygosity and often self-incompatibility. Anther culture is currently the method of choice for microspore embryogenesis in many crops. However, isolated microspore culture is a better way to investigate the processes at the cellular, physiological, biochemical and molecular levels as it avoids the influence of somatic anther tissue. To exploit the potential of this technique, it is important to separate the key factors affecting the process and, among them, culture medium composition and particularly the plant growth regulators and their concentration, as they can greatly enhance regeneration efficiency. To our knowledge, the ability of meta-Topolin, a naturally occurring aromatic cytokinin, to induce gametic embryogenesis in isolated microspores of Citrus has never been investigated. In this study, the effect of two concentrations of meta-Topolin instead of benzyladenine or zeatin in the culture medium was investigated in isolated microspore culture of two genotypes of Citrus. After eleven months of isolated microspore culture, for both genotypes and for all the four tested media, the microspore reprogramming and their sporophytic development was observed by the presence of multinucleated calli and microspore-derived embryos at different stages. Microsatellite analysis of parental and embryo samples was performed to determine the embryo alleles constitution of early embryos produced in all tested media, confirming their origin from microspores. To our knowledge, this is the first successful report of Citrus microspore embryogenesis with isolated microspore culture in Citrus, and in particular in Citrus clementina Hort. ex Tan, cvs. ‘Monreal Rosso’ and ‘Nules’.
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early embryo achievement through isolated microspore culture in citrus clementina hort ex tan cvs monreal rosso and nules
Frontiers in Plant Science, 2015Co-Authors: Benedetta Chiancone, Ivett Barany, Valeria Gianguzzi, Marines Marli Gniech Karasawa, A Abdelgalel, Daniela Torello Marinoni, Roberto Botta, P S Testillano, Maria GermanaAbstract:Microspore embryogenesis is a method of achieving complete homozygosity from plants. It is particularly useful for woody species, like Citrus, characterized by long juvenility, a high degree of heterozygosity and often self-incompatibility. Anther culture is currently the method of choice for microspore embryogenesis in many crops. However, isolated microspore culture is a better way to investigate the processes at the cellular, physiological, biochemical, and molecular levels as it avoids the influence of somatic anther tissue. To exploit the potential of this technique, it is important to separate the key factors affecting the process and, among them, culture medium composition and particularly the plant growth regulators and their concentration, as they can greatly enhance regeneration efficiency. To our knowledge, the ability of meta-Topolin, a naturally occurring aromatic cytokinin, to induce gametic embryogenesis in isolated microspores of Citrus has never been investigated. In this study, the effect of two concentrations of meta-Topolin instead of benzyladenine or zeatin in the culture medium was investigated in isolated microspore culture of two genotypes of Citrus. After 11 months of isolated microspore culture, for both genotypes and for all the four tested media, the microspore reprogramming and their sporophytic development was observed by the presence of multinucleated calli and microspore-derived embryos at different stages. Microsatellite analysis of parental and embryo samples was performed to determine the embryo alleles constitution of early embryos produced in all tested media, confirming their origin from microspores. To our knowledge, this is the first successful report of Citrus microspore embryogenesis with isolated microspore culture in Citrus, and in particular in Citrus clementina Hort. ex Tan, cvs. 'Monreal Rosso' and 'Nules.'
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microspore derived embryogenesis in pepper capsicum annuum l subcellular rearrangements through development
Biology of the Cell, 2005Co-Authors: Ivett Barany, Mariacarmen Risueno, Pablo Gonzalezmelendi, Begona Fadon, Judit Mityko, P S TestillanoAbstract:Background information. In vitro-cultured microspores, after an appropriate stress treatment, can switch towards an embryogenic pathway. This process, known as microspore embryogenesis, is an important tool in plant breeding. Basic studies on this process in economically interesting crops, especially in recalcitrant plants, are very limited and the sequence of events is poorly understood. In situ studies are very convenient for an appropriate dissection of microspore embryogenesis, a process in which a mixture of different cell populations (induced and non-induced) develop asynchronically. Results. In the present study, the occurrence of defined subcellular rearrangements has been investigated during early microspore embryogenesis in pepper, an horticultural crop of agronomic interest, in relation to proliferation and differentiation events. Haploid plants of Capsicum annuum L. (var. Yolo Wonder B) have been regenerated from in vitro anther cultures by a heat treatment at 35°C for 8 days. Morphogenesis of microspore-derived embryos has been analysed, at both light and electron microscopy levels, using low-temperature-processed, well-preserved specimens. The comparison with the normal gametophytic development revealed changes in cell organization after embryogenesis induction, and permitted the characterization of the time sequence of a set of structural events, not previously defined in pepper, related to the activation of proliferative activity and differentiation. These changes mainly affected the plastids, the vacuolar compartment, the cell wall and the nucleus. Further differentiation processes mimicked that of the zygotic development. Conclusions. The reported changes can be considered as markers of the microspore embryogenesis. They have increased the understanding of the mechanisms controlling the switch and progression of the microspore embryogenesis, which could help to improve its efficiency and to direct strategies, especially in agronomically interesting crops.
Kamalraj Subban - One of the best experts on this subject based on the ideXlab platform.
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salicylic acid as an effective elicitor for improved taxol production in endophytic fungus pestalotiopsis Microspora
PLOS ONE, 2019Co-Authors: Kamalraj Subban, Ramesh Subramani, Vishnu Priya Madambakkam Srinivasan, Muthumary Johnpaul, Jayabaskaran ChelliahAbstract:Salicylic acid (SA) is an effective elicitor to increase taxol production in Pestalotiopsis Microspora. Addition of SA at the concentration of 300 μM yielded taxol 625.47 μg L-1, 45- fold higher than that of the control. Elicitation of the role of SA in the fungal taxol biosynthetic pathway revealed that SA enhanced reactive oxygen species and lipid peroxidation of unsaturated fatty acids of P. Microspora mycelia. This oxidative process stimulates isoprene biosynthetic pathway by triggering expression of the geranylgeranyl pyrophosphate synthase gene leading to improved biosynthesis of taxol in P. Microspora.
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Effect of different concentrations of SA on catalase and peroxidase activity by P. Microspora.
2019Co-Authors: Kamalraj Subban, Ramesh Subramani, Vishnu Priya Madambakkam Srinivasan, Muthumary Johnpaul, Jayabaskaran ChelliahAbstract:Effect of different concentrations of SA on catalase and peroxidase activity by P. Microspora.
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Effect of different concentrations of SA on total lipids and sterol by P. Microspora.
2019Co-Authors: Kamalraj Subban, Ramesh Subramani, Vishnu Priya Madambakkam Srinivasan, Muthumary Johnpaul, Jayabaskaran ChelliahAbstract:Effect of different concentrations of SA on total lipids and sterol by P. Microspora.
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Effect of different concentrations of SA on glutathione and lipid peroxidase activity by P. Microspora.
2019Co-Authors: Kamalraj Subban, Ramesh Subramani, Vishnu Priya Madambakkam Srinivasan, Muthumary Johnpaul, Jayabaskaran ChelliahAbstract:Effect of different concentrations of SA on glutathione and lipid peroxidase activity by P. Microspora.
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Effect of different concentrations of SA on total protein and superoxide dismutase activity by P. Microspora.
2019Co-Authors: Kamalraj Subban, Ramesh Subramani, Vishnu Priya Madambakkam Srinivasan, Muthumary Johnpaul, Jayabaskaran ChelliahAbstract:Effect of different concentrations of SA on total protein and superoxide dismutase activity by P. Microspora.
Maria Germana - One of the best experts on this subject based on the ideXlab platform.
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early embryo achievement through isolated microspore culture in citrus clementina hort ex tan cvs monreal rosso and nules
Frontiers in Plant Science, 2015Co-Authors: Benedetta Chiancone, Ivett Barany, Valeria Gianguzzi, Marines Marli Gniech Karasawa, A Abdelgalel, Daniela Torello Marinoni, Roberto Botta, P S Testillano, Maria GermanaAbstract:Microspore embryogenesis is a method of achieving complete homozygosity from plants. It is particularly useful for woody species, like Citrus, characterized by long juvenility, a high degree of heterozygosity and often self-incompatibility. Anther culture is currently the method of choice for microspore embryogenesis in many crops. However, isolated microspore culture is a better way to investigate the processes at the cellular, physiological, biochemical and molecular levels as it avoids the influence of somatic anther tissue. To exploit the potential of this technique, it is important to separate the key factors affecting the process and, among them, culture medium composition and particularly the plant growth regulators and their concentration, as they can greatly enhance regeneration efficiency. To our knowledge, the ability of meta-Topolin, a naturally occurring aromatic cytokinin, to induce gametic embryogenesis in isolated microspores of Citrus has never been investigated. In this study, the effect of two concentrations of meta-Topolin instead of benzyladenine or zeatin in the culture medium was investigated in isolated microspore culture of two genotypes of Citrus. After eleven months of isolated microspore culture, for both genotypes and for all the four tested media, the microspore reprogramming and their sporophytic development was observed by the presence of multinucleated calli and microspore-derived embryos at different stages. Microsatellite analysis of parental and embryo samples was performed to determine the embryo alleles constitution of early embryos produced in all tested media, confirming their origin from microspores. To our knowledge, this is the first successful report of Citrus microspore embryogenesis with isolated microspore culture in Citrus, and in particular in Citrus clementina Hort. ex Tan, cvs. ‘Monreal Rosso’ and ‘Nules’.
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early embryo achievement through isolated microspore culture in citrus clementina hort ex tan cvs monreal rosso and nules
Frontiers in Plant Science, 2015Co-Authors: Benedetta Chiancone, Ivett Barany, Valeria Gianguzzi, Marines Marli Gniech Karasawa, A Abdelgalel, Daniela Torello Marinoni, Roberto Botta, P S Testillano, Maria GermanaAbstract:Microspore embryogenesis is a method of achieving complete homozygosity from plants. It is particularly useful for woody species, like Citrus, characterized by long juvenility, a high degree of heterozygosity and often self-incompatibility. Anther culture is currently the method of choice for microspore embryogenesis in many crops. However, isolated microspore culture is a better way to investigate the processes at the cellular, physiological, biochemical, and molecular levels as it avoids the influence of somatic anther tissue. To exploit the potential of this technique, it is important to separate the key factors affecting the process and, among them, culture medium composition and particularly the plant growth regulators and their concentration, as they can greatly enhance regeneration efficiency. To our knowledge, the ability of meta-Topolin, a naturally occurring aromatic cytokinin, to induce gametic embryogenesis in isolated microspores of Citrus has never been investigated. In this study, the effect of two concentrations of meta-Topolin instead of benzyladenine or zeatin in the culture medium was investigated in isolated microspore culture of two genotypes of Citrus. After 11 months of isolated microspore culture, for both genotypes and for all the four tested media, the microspore reprogramming and their sporophytic development was observed by the presence of multinucleated calli and microspore-derived embryos at different stages. Microsatellite analysis of parental and embryo samples was performed to determine the embryo alleles constitution of early embryos produced in all tested media, confirming their origin from microspores. To our knowledge, this is the first successful report of Citrus microspore embryogenesis with isolated microspore culture in Citrus, and in particular in Citrus clementina Hort. ex Tan, cvs. 'Monreal Rosso' and 'Nules.'
