Differential callus type formation by auxins and cytokinin in in vitro cultures of pepper (Capsicum annuum L.)

ABSTRACT

Two types of callus were produced by pepper explants cultured in various media containing auxins, the cytokinin 6-benzylaminopurine (BAP) and the auxin transport inhibitor 2,3,5-triiodobenzoic acid (TIBA). Callus produced on media containing auxins alone was friable, grey-green or green-orange in colour and more compact, whereas when BAP was added to culture media with a low concentration of auxin or when the medium contained TIBA alone, the callus produced was white and very hard. This type of callus was also produced in cultures of older tissues and of young tissues cultured on hormonefree medium. Results are discussed in relation to the role of cytokinins in wounding, phenylpropanoid metabolism and lignin biosynthesis.     

Micropropagation of Calluna vulgaris cv. ‘H.E. Beale’

Axillary shoot producing cultures were obtained from microcuttings and shoot tips of Calluna vulgaris cv. H.E. Beale. For cultures derived from microcuttings the highest multiplication rate of 38 shoots (5 mm or longer) was obtained on a reduced salt medium with the addition of 0.5 mgl^-1 2-isopentenyladenine (2iP) during an 8 week subculture. For shoot tip derived cultures 0.2 mgl^-1 6-benzyladenine (BA) was the best cytokinin and led to a multiplication rate of 26 for a 6 week subculture. The addition of 1 g/l casein hydrolysate to a multiplication medium enhanced shoot proliferation in presence of 0.5 mgl^-1 BA.Despite various auxin treatments shoots formed no roots in vitro but rooted readily if transferred to a peat substrate ex vitro. A high rooting percentage (80%) was also obtained with shoots taken from the end of a multiplication phase and rooted directly. An additional subculture on low auxin containing media before transfer to peat substrate is recommended because the shoot condition can be improved in this way. A high number of rooted plantlets was produced, so the methods described will allow mass propagation.     

Endogenous hormone levels during direct somatic embryogenesis in Medicago falcata

Endogenous indole-3-acetic acid, abscisic acid and cytokinins (zeatin, zeatin riboside, N-isopentenyladenine and N-isopentenyladenosine) were evaluated in initial explants (leaves) of in vitro propagated plants of alfalfa ( Medicago falcata L.) lines varying in embryogenic capacity and during the somatic embryogenesis process. Fast embryo-genic induction was correlated with high IAA and low ABA levels in the initial explants. No significant differences were observed in the cytokinin contents. Our results suggest that a certain hormone balance is necessary to allow the expression of the embryogenic potential. The consistent stages of the direct somatic embryogenesis are also characterized by changes in hormonal levels.     

Seed Treatment with Auxins Modulates Growth and Ion Partitioning in Salt-stressed Wheat Plants

Experiments were performed to determine whether seed priming with different concentrations (100, 150, and 200 mg/L) ofauxins (indoleacetic acid (IAA), indolebutyric acid (IBA), or their precursor tryptophane (Trp)) could alter salinity inducedperturbances in salicylic acid and ion concentrations and, hence, growth in wheat (Triticum aestivum L.) cultivars, namelyM. H.-97 (salt intolerant) and Inqlab-91 (salt tolerant). Primed and non-primed seeds were sown in Petri dishes in a growthroom, as well as in a field treated with 15 dS/m NaCl salinity. All priming agents, except IBA, increased the final germinationpercentage in both cultivars. The seedlings of either cultivar raised from Trp-treated seeds had greater dry biomass whenunder salt stress. In field experiments, Trp priming was much more effective in mediating the increase in grain yield,irrespective of the cultivar, under salt stress. The alleviatory effect of Trp was found to be associated with reduced uptakeof Na~ in the roots and subsequent translocation to the shoots, as well as increased partitioning of Ca~(2 )in the roots ofsalt-stressed wheat plants. Plants of both cultivars raised from Trp-and IAA-treated seeds accumulated free salicylic acidin their leaves when under salt stress. Overall, the Trp priming-induced improvement in germination and the subsequentgrowth of wheat plants could be related to ion homeostasis when under salt stress. The possible involvement of salicylicacid in the Trp priming-induced better growth under conditions of salt stress is discussed.     

Phytohormone contents in Corylus avellana and their relationship to age and other developmental processes

Endogenous levels of indole-3-acetic acid, abscisic acid, and cytokinins (Z-type: dihydrozeatin, dihydrozeatin riboside, zeatin, and zeatin riboside; iP-type: N ⁶-isopentenyl adenine and N ⁶-isopentenyl adenosine), were determined in leaves of hazelnut (Corylus avellana L.) (adult material from spring, autumn and forced outgrowth, and juvenile material). Our results showed high levels of indole-3-acetic acid, abscisic acid and total cytokinins in spring samples and low levels of the same hormones in autumn and forced outgrowth materials. The ratios of iP-type/Z-type cytokinins were low in autumn and spring leaves, while they were high in the juvenile and forced outgrowth samples. Both juvenile and forced-outgrowth hazel tissues also showed a high morphogenetic potential, suggesting that the ratio of iP-type/Z-type cytokinins may be a good index of in vitro potential of hazelnut materials.     

Inhibition by 2,4-D of somatic embryogenesis in carrot as explored by its reversal by difluoromethylornithine

The development of somatic embryos is, in many plants, inhibited by 2,4-dichlorophenoxyacetic acid (2,4-D) and other auxins. The finding that difluoromethylornithine (DFMO) can counteract this inhibition has been used to test some of the hypotheses for the mechanism of inhibition.
Inhibition of somatic embryogenesis in carrot ( Daucus carota L.) by exogenous ethylene (from ethephon), antioxidants (ascorbic acid and glutathione), ethanol/acetaldehyde and abscisic acid was not counteracted by DFMO, indicating that the inhibitory effect of 2,4-D is not manifest through the formation of these compounds. Embryogenesis was abolished by micromolar concentrations of the polar auxin transport inhibitors 2, 3, 5-triiodobenzoic acid (TIBA), N-1-naphthylphthalamic acid (NPA) and 9-hydroxyfluorene-9-carboxylic acid (HFCA). This inhibition was counteracted to a considerable extent by DFMO. Inhibition by relatively high concentrations of the antiauxin 2-( p -chlorophenoxy)-isobutyric acid (CPIB), which does not affect polar auxin transport, was in contrast not counteracted by DFMO. These findings indicate that exogenous auxins may inhibit embryogenesis by interfering with the ability of postglobular embryos to set up internal auxin gradients necessary for polarized growth.     

Seed Treatment with Auxins Modulates Growth and Ion Partitioning in Salt-stressed Wheat Plants

Experiments were performed to determine whether seed priming with different concentrations （100, 150, and 200 mg/L） of auxins （indoleacetic acid （IAA）, indolebutyric acid （IBA）, or their precursor tryptophane （Trp）） could alter salinity induced perturbances in salicylic acid and ion concentrations and, hence, growth in wheat （Triticum aestivum L.） cultivars, namely M.H.-97 （salt intolerant） and tnqtab-91 （salt tolerant）. Primed and non-primed seeds were sown in Petri dishes in a growth room, as well as in a field treated with 15 dS/m NaCl salinity. All priming agents, except IBA, increased the final germination percentage in both cultivars. The seedlings of either cultivar raised from Trp-treated seeds had greater dry biomass when under salt stress. In field experiments, Trp priming was much more effective in mediating the increase in grain yield, irrespective of the cultivar, under salt stress. The alleviatory effect of Trp was found to be associated with reduced uptake of Na^＋ in the roots and subsequent translocation to the shoots, as well as increased partitioning of Ca^＋ in the roots of salt-stressed wheat plants. Plants of both cultivars raised from Trp-and IAA-treated seeds accumulated free salicylic acid in their leaves when under salt stress. Overall, the Trp priming-induced improvement in germination and the subsequent growth of wheat plants could be related to ion homeostasis when under salt stress. The possible involvement of salicylic acid in the Trp priming-induced better growth under Conditions of salt stress is discussed.     

Effect of growth regulators and ethylmethane sulphonate on growth,and chlorophyll,sugar and proline contents in <Emphasis Type="Italic">Dracaena sanderiana</Emphasis> cultured <Emphasis Type="Italic">in vitro</Emphasis>

A high efficient four step protocol (callus initiation, regeneration, shoot elongation and rooting) for in vitro propagation of Dracaena sanderiana Sander ex Mast was developed. Callusing was achieved from nodal stem segment explants treated with various concentrations of ethylmethane sulphonate (EMS) on MS medium supplemented with 2,4-dichlorophenoxyacetic acid (2,4-D; 1.5 g m⁻³). A significant increase in callus induction percentage and biomass production was noticed from lower EMS treated lines (ET₁ and ET₂) comparatively to control and other (ET₃, ET₄ and ET₅) lines. Calli of ET₁ line showed high regeneration potential on MS medium with N ⁶-benzylaminopurine (BAP; 1.75 g m⁻³). Length of microshoots, which was reduced by EMS, restored by addition of gibberellic acid (GA₃; 0.4 g m⁻³). A marked increase in rooting with increasing EMS concentration was noticed on MS medium fortified with 3-indolebutyric acid (IBA; 1.5 g m⁻³).