Ljubinka Ćulafić

Relationship between the concentration of macroelements, their uptake and multiplication of cherry rootstock Gisela 5 in vitro

Sweet cherry rootstock Gisela 5 was micropropagated on Murashige and Skoog (MS) medium, on MS medium containing double-strength macro salts (MS 2×), 1/2 strength (MS 1/2) and 1/4 strength (MS 1/4) with 4.4 μM BA, 0.5 μM NAA, and 0.3 μM GA3. All the analyses and parameters were monitored on day 0, day 20 and day 40 after subculturing. As fresh and dry weight of the explants increased during subculturing, the fresh and dry weight of the media decreased. The pH of the media declined during subculturing following by slow increase on media MS 1/2 and MS 1/4. Gisela 5 showed the best growth and development on MS 2× and MS media with the highest N and P uptake. Growth and multiplication depend on the uptake of these elements from the medium.

Micropropagation of Pinus heldreichii

Micropropagation by organogenesis from mature embryos of Pinus heldreichii Christ. was achieved. The frequency of adventitious bud induction was higher on embryos grown on Gresshoff and Doy medium than on Von Arnold and Eriksson, or Murashige and Skoog medium. The greatest number of buds and developed shoots was obtained after induction with benzyladenine at 2.22 or 4.40 μM for four weeks. Shorter induction time was less effective for bud induction, but subsequent shoot elongation was accelerated. Shoots elongated on half-strength, growth regulator-free medium supplemented with activated charcoal. After pulse treatment with 1 mM indole-3-butyric acid twenty shoots were rooted, while agar-solidified medium supplemented with α-naphthaleneacetic acid (0.27 or 1.08 μM), or indole-3-butyric acid (0.25 or 0.98 μM) induced callus formation only.

Induction of somatic embryogenesis and embryo development in Rumex acetosella L.

Axillary buds of the dioecious plant Rumex acetosella L. were isolated and cultured in vitro. The callus tissue which developed at the basal parts of the explants displayed a high capacity for shoot formation. This morphogenetic pattern was predominant on Murashige and Skoog (MS) medium supplemented with 2% sucrose, 2.2 mgl-1 benzylaminopurine and 0.17 mgl-1 indole-3-acetic acid. Somatic embryogenesis was induced when the osmolality of the medium was increased by adding 6% sucrose instead of 2%, or hexitols in addition to 2% sucrose. Most of the embryogenic calli were formed on the basal parts of leaf laminae and bracts. Development and maturation was strongly promoted by transferring the tissue to a solid or liquid medium lacking benzylaminopurine and indole-3-acetic acid and supplemented with 10 mgl-1 gibberellic acid. The embryos germinated and developed into normal rosette plants when transferred to vermiculite moistened with hormone-free, half-strength MS salt solution. The histology of successive embryogenic stages is presented.

Flowering ofin vitro grown spinach shoots in the presence of the Herbicide Sandoz 9789

Stem tips ofSpinacia oleracea were isolated and grown in sterile culture, with Sandoz 9789 added to the medium. Although the herbicide provoked a complete loss of all chloroplast pigments, developed shoots were able to flower, in response to long days; in short days flowering was inducible by adding gibberellins. Sandoz 9789 decreased the elongation of stems by up to 30%; the percentage of flowering plants in culture was also decreased. However, among those plants that flowered, the majority were male, so that Sandoz 9789 produced a shift of the sex ratio toward maleness.

In vitro organogenesis in two dioecious species, Rumex acetosella L. and R. acetosa L. (Polygonaceae)

Callus cultures were established from dioecious plant species Rumex acetosella and R. acetosa, using cotyledons, hypocotyls and stem tips of aseptically germinated seedlings as primary explants. Cultures were also established from male and female R. acetosella adult plants, starting from vegetative lateral buds. Cell division was induced using a high 2,4-D concentration, while bud induction and multiplication were stimulated on a medium with high BAP/IAA ratio. Cotyledon fragments of both species produced only rhizogenic calli. Hypocotyl-derived calli of R. acetosella produced buds, while those of R. acetosa showed no bud forming response under these conditions. Bud multiplication occurred in stem tip cultures of both species and in lateral bud cultures of R. acetosella. Calli derived from male plants produced more buds than those from female. Shoots were easily rooted using IBA, and plantlets were effectively transferred to soil. Flowering was not induced in culture. The sex of regenerated male and female plants was not altered by the culture conditions.

The behaviour of the shoot apical meristem inChenopodium rubrum under conditions non-inductive for flowering

The results of different photoperiodic treatments preventing flowering and representing the control vegetative treatments in the studies of floral induction and differentiation were studied inChenopodium rubrum seedlings. A fully vegetative growth pattern of the meristem was maintained only in continuous light or after a photoperiodic treatment which consisted in a 15 min light break of the 8 h dark periods which themselves are a threshold for flowering inChenopodium. Light breaks applied to 10 h and longer dark periods did not prevent the changes resembling the early events of transition to flowering. Disappearance of zonal pattern, stimulation of apical growth, precocious initiation of leaf primordia and weakening of apical dominance have been observed. Flower formation did not follow.

Effect of photoperiod during growth of Chenopodium rubrum mother plants on properties of offspring

Using in vitro culture, we determined the effect of photoperiod during growth of Chenopodium rubrum mother plants on vegetative and reproductive development of offspring. Photoperiod during flowering induction of mother plants (the first 6 d after the germination) has the key influence on seed germination and offspring growth, while offspring flowering and seed maturation is determined by photoperiod their mothers experienced during, and shortly after, flowering induction. The mechanism can be through changes in seed protein pattern which we found dependent on photoperiod experienced by mother plants.

Hairy roots formation in recalcitrant-to-transform plant Chenopodium rubrum

Susceptibility of C. rubrum to Agrobacterium-mediated transformation was demonstrated by inoculating the petioles of in vitro grown plants with A. rhizogenes strain A4M70GUS. Hairy roots were produced in 8 % of explants. They were isolated and maintained on plant growth regulator-free solid or liquid half-strength Murashige and Skoog medium for two years. Hairy root fresh mass increased 30 — 90 folds when grown in liquid medium, which was superior to solid medium, where most of the hairy roots produced calli. When these calli were grown on medium supplemented with 0.5 mg dm-3 thidiazuron, embryo-like structures were obtained. Transgenic status of long-term callus and hairy root cultures was confirmed by histochemical GUS assay, by PCR specific to the uidA, rolA&B and ags genes and by Southern hybridization.

Endogenous cytokinins in the vegetative and reproductive phases of development in the dioecious plant Rumex acetosella L .

The content of endogenous cytokinins has been analysed in leaf and inflorescence extracts of male and female R. acetosella plants, using gas chromatography. Plant parts were extracted at four stages of development: leaves of juvenile plants, leaves of adult plants at the time of flower initiation and in full bloom, and upper internodes of the inflorescence stalks. Cytokinins with characteristics similar to isopentenyl adenine and adenosin, zeatin, zeatin riboside, and a bound form of zeatin, were all found in the extracts. The total amount of cytokinins was higher in female than in male plants during all these stages.

Indole auxins in spinach plants grown in long and short days

The endogenous auxin-like substances were analyzed in the shoot extracts of young spinach seedlings, exposed to photoperiodic induction. At least eight indole auxins were found. One of them was identified as tryptophan, the other one is most probably IAA. The plants grown in long days had a higher level of ether soluble auxins than the controls in short days. Separate extractions of plants after each of the eight inductive days showed that the auxin content was not constant, but subjected to irregular oscillations. However, parallel oscillations were also found in control plants grown in short days. Staminate plants were found to contain more endogenous auxins than the pistillate ones. It is concluded that the quantitative changes in auxins during the photoperiodic induction are probably not related to flowering, but to some other growth process, common to all plants in that phase of growth. The higher level of auxins in staminate plants may be the cause of their faster elongation before the onset of flowering.AbstractV extraktech nadzemních částí klíčních rostlin špenátu vystavených fotoperiodické indukci byly stanoveny endogenní auxiny. Bylo zjištěno nejméně osm indolových auxinů. Jeden z nich byl identifikován jako tryptofan, další je s velkou pravděpodobností IOK. Rostliny rostoucí při dlouhém dni měly vyšší hladinu auxinů rozpustných v eteru než kontrolní rostliny rostoucí při krátkém dni. Jednotlivé extrakce rostlin vždy po osmi indukčních dnech ukázaly, že obsah auxinů není konstantní, ale že podléhá nepravidelným oscilacím. Paralelní oscilace však byly zjištěny i u kontrolních rostlin pěstovaných při krátkém dni. Samčí rostliny měly vyšší obsah endogenních auxinů než rostliny samičí. Ze zjištěných výsledků vyplývá, že kvantitativní změny v obsahu auxinů během fotoperiodické indukce pravděpodobně nejsou v žádném vztahu ke kvetení, ale k jiným růstovým procesům, společným všem rostlinám v této vývojové fázi. Vyšší hladina auxinů v samčích rostlinách pravděpodobně způsobuje jejich rychlejší prodlužování před nástupem kvetení.