Microtubules in maize leaf protoplasts in relation to donor tissue and in vitro culture

Wang, H.; Cutler, Adrian J.; Saleem, Mahmood; Fowke, Larry C.

doi:10.1007/bf01352920

Cited by 9 publications

(6 citation statements)

References 12 publications

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“…Regenerated cell walls were stained with the fluorescent dyes Calcofluor White, which stains both β‐(1→3)‐ and β‐(1→4)‐glucans (Nagata and Takebe 1970), or aniline blue, which stains β‐(1→3)‐glucans (Eschrich and Currier 1964). Nuclei of fixed [with 8% (w/v) paraformaldehyde] protoplasts were stained with the fluorescent dye Hoechst according to Wang et al (1989). Fluorescence was examined with an Axioplan Zeiss microscope.…”

Section: Methodsmentioning

confidence: 99%

Protoplast isolation and culture from carob (Ceratonia siliqua) hypocotyls: ability of regenerated protoplasts to produce mannose‐containing polysaccharides

Sotiriou¹,

Fry²,

Spyropoulos³

2007

Physiologia Plantarum

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The aim of this study was to isolate protoplasts from carob (Ceratonia siliqua L.) embryonic tissues with the ability to regenerate cell walls, divide and synthesize galactomannan, a valuable polysaccharide for industry. Protoplasts isolated from carob hypocotyl hooks regenerated cell walls within 24 h. The first divisions of the regenerated cells were observed after 2 days of culture. The highest percentage that successfully divided was achieved when the seedlings were grown under diffuse light, the hypocotyl hooks were plasmolysed for 1 h before incubation in the protoplast isolation solution and the protoplasts were cultured under diffuse light. After 9 days of culture, cell clusters, consisting of eight cells, had been produced, which underwent further mitotic divisions and which were expected to lead to callus formation. Polysaccharide and oligosaccharide synthesis during protoplast regeneration was studied by radiolabelling with exogenous d‐[U‐14C]glucose, d‐[U‐14C]mannose or d‐[2‐3H]mannose, which gave rise to uniform, moderately specific and highly specific labelling, respectively. As revealed by the radioactivity distribution in cell wall monosaccharides, the regenerants deposited new wall polymers that differed markedly from those being synthesized by the hypocotyls from which the protoplasts had been isolated. The regenerants deposited large amounts of callose and smaller amounts of galactose‐, arabinose‐ and mannose‐containing polymers. The latter included glucuronomannan, as demonstrated by a new method involving partial acid hydrolysis followed by β‐glucuronidase (EC 3.2.1.31) digestion. The regenerating protoplasts also released soluble extracellular carbohydrates: polysaccharides which appeared to be mainly acidic arabinogalactans, and oligosaccharides which were mainly neutral and contained glucose, galactose and mannose. We conclude that regenerating carob protoplasts are a useful system for studying carbohydrate secretion, including mannose‐rich poly‐ and oligosaccharides.

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Section: Methodsmentioning

confidence: 99%

Protoplast isolation and culture from carob (Ceratonia siliqua) hypocotyls: ability of regenerated protoplasts to produce mannose‐containing polysaccharides

Sotiriou¹,

Fry²,

Spyropoulos³

2007

Physiologia Plantarum

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“…The high percentage of nuclei with DNA synthesis suggests that these cells are not irreversibly differentiated. Dedifferentiation in terms of microtubule reappearance has also been observed in these protoplasts [31]. Experiments are underway to compare DNA synthesis in mesophyll protoplasts isolated from different parts of maize leaves.…”

Section: Dna Synthesis In Maize Leaf Protoplastsmentioning

confidence: 99%

DNA replication in maize leaf protoplasts

Wang

Cutler

Saleem

et al. 1989

Plant Cell Tiss Organ Cult

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Maize leaf protoplasts were investigated for their metabolic competence and capacity to synthesize DNA. When protoplasts were incubated at elevated temperatures, they exhibited a heat shock response with specific proteins being preferentially synthesized. This indicated that the protoplasts were fully metabolically functional and capable of responding to environmental stimuli. Significant DNA synthesis was observed in these protoplasts after incorporation of 3H-thymidine into chromatin by trichloroacetic acid precipitation and by incorporation of 5-bromo-2-deoxyuridine (BrdU), an analog of thymidine, detected by immunofluorescence. The immunocytochemical method revealed that about 50% of nuclei in the maize leaf protoplasts were labelled after 3 days of culture and that most of these nuclei were labelled as intensely as normal mitotic cells. Aphidicolin, an inhibitor of DNA polymerase-a, decreased the percentage of labelled nuclei, demonstrating that the labelling was substantially due to replicative DNA synthesis. However, chromosome condensation was not observed. It is proposed that these protoplasts are capable of DNA synthesis, but incapable of nuclear division. Effects of media additives on the number of nuclei entering S phase in these protoplasts were also assessed by the immunocytocbemical method. Inclusion of 80 mM Ca 2+ in the enzyme solution increased protoplast yield and also appeared beneficial to DNA synthesis. The antioxidant, n-propyl gallate, which was used to stabilize the protoplasts, delayed the onset of DNA synthesis. Arginine and spermidine produced a slight increase in DNA synthesis.Abbreviations: BrdU--5-bromo-2-deoxyuridine; DMSO--dimethyl sulfoxide; n-PG--npropyl gallate; PBS--phosphate-buffered saline

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“…H προσοχή τα τελευταία χρόνια έχει στραφεί στην επίδραση παραγόντων των ενζυμικών παρασκευασμάτων (Ishii, 1988) ή παραγόντων ελευθερούμενων από τα κυτταρικά τοιχώματα κατά τη διάρκεια της απομόνωσης (Hahne and Lorz, 1988). Εχει διαπιστωθεί οξείδωση λιπιδίων (lipid peroxidation) σε πρωτοπλάστες μεσοφύλλου σιτηρών (Cutler et al, 1989) και παρουσία αναγμένων μορφών οξυγόνου κατά τη διάρκεια της απομόνωσης πρωτοπλαστών από αδιαφοροποίητα κύτταρα ρυζιού (Ishii, 1987(Ishii, , 1988. Οι αναγμένες μορφές οξυγόνου είναι το αίτιο για αλυσσιδωτές αντιδράσεις οξείδωσης των μεμβρανικών λιπιδίων (Gutteridge and Halliwell, 1990) αλλά και πολλών άλλων βιολογικών μορίων σε όλα τα διαμερίσματα του κυττάρου (Scandalios, 1993 (Ishii, 1988, Sharon et al, 1993.…”

Section: εισαγωγη -βιβλιογραφικη επισκοπησηunclassified

“…Εχει ευρεθεί ότι ο χρόνος επώασης με τα ενζυμικά παρασκευάσματα της απομόνωσης των πρωτοπλαστών επηρεάζει την οργάνωση του δικτύου των μικροσωληνίσκων σε πρωτοπλάστες Vitis (Lee et ai, 1989). Επίσης, σε μή αναγεννόμενους πρωτοπλάστες, το δίκτυο των μικροσωληνίσκων είναι πολύ αραιό (Hahne and Hoffman, 1984, 1985, Meijer and Simmonds, 1988, Wang et al, 1989. Υποστηρίζεται έτσι από τους ίδιους ερευνητές ότι, ένα πυκνό και καλά οργανωμένο δίκτυο μικροσωληνίσκων, είναι απαραίτητη προϋπόθεση για τη διαδικασία της μίτωσης, ενώ η επανασύσταση του νέου κυτταρικού τοιχώματος επιδρά στη σταθεροποίηση των μικροσωληνίσκων με την βοήθεια διαμεμβρανικών πρωτεϊνών (Simmonds, 1992).…”

Section: εισαγωγη -βιβλιογραφικη επισκοπησηunclassified

“…Among the factors assessed that may contribute to recalcitrance of protoplasts is oxidative stress (Ishii, 1988;Cutler et al, 1989;. Wounding, cell wall hydrolytic enzymes, and change of cultured protoplasts and cells from autotrophic to heterotrophic metabolism in the presence of altered hormonal status are some of the factors that induce oxidative stress.…”

Section: Catalase Expression Inmentioning

confidence: 99%

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Αντιοξειδωτικα Ενζυμικα Συστηματα Σε Πρωτοπλαστες Απο Μεσοφυλλο Vitis Vinifera L. Και Nicotiana Tabacum L.

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Microtubules in maize leaf protoplasts in relation to donor tissue and in vitro culture

Cited by 9 publications

References 12 publications

Protoplast isolation and culture from carob (Ceratonia siliqua) hypocotyls: ability of regenerated protoplasts to produce mannose‐containing polysaccharides

Protoplast isolation and culture from carob (Ceratonia siliqua) hypocotyls: ability of regenerated protoplasts to produce mannose‐containing polysaccharides

DNA replication in maize leaf protoplasts

Αντιοξειδωτικα Ενζυμικα Συστηματα Σε Πρωτοπλαστες Απο Μεσοφυλλο Vitis Vinifera L. Και Nicotiana Tabacum L.

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