Luis A. Mroginski

Foliar sprays with ABA promote growth of Ilex paraguariensis by alleviating diurnal water stress

Stomatal closure, relative water content (RWC) and vegetative growth were monitored in Ilex paraguariensis plants grown under well-watered conditions with a photosynthetic photon flux density (PPFD) varying from 100% to 1.5%, and sprayed weekly with either distilled water (control) or 1.89 mM abscisic acid (ABA). ABA treatments caused stomatal closure, ranging from 62% to 73%. These treatments also increased RWC in the early evening from 82% to 92% and 88% to 94% in mature and immature leaves, respectively. Such alleviation of the water stress was correlated with increases in leaf area, leaf dry weight (DW), shoot length and shoot DW. On day 35 from the beginning of the experiment, the increases in DW of both leaves and shoots were 1.5-fold at the 1.5% PPFD and 3-fold (for leaves) and 4.5-fold (for shoots) under 100% PPFD. In water-sprayed control plants grown under 1.5% PPFD shoot length also increased significantly, although these shoots contained more ABA (assessed by capillary gas chromatography–mass spectrometry) than those of plants grown under 100% PPFD. These results show that ABA sprayed on to leaves promotes growth in I. paraguariensis plants by alleviating diurnal water stress.

Thidiazuron Promotes In Vitro Plant Regeneration of Arachis correntina (Leguminosae) via Organogenesis

Arachis correntina (Burkart) Krapov. & W.C. Gregory is a herbaceous perennial leguminous plant growing in the Northeast of the Province Corrientes, Argentina. It is important as forage. The development of new A. correntina cultivars with improved traits could be facilitated through the application of biotechnological strategies. The purpose of this study was to investigate the plant regeneration potential of mature leaves of A. correntina in tissue culture. Buds were induced from both petiole and laminae on 0.7% agar-solidified medium containing 3% sucrose, salts and vitamins from Murashige and Skoog (MS) supplemented with 0.5–25 μM thidiazuron (TDZ). Shoot induction was achieved by transference of calli with buds to MS supplemented with 5 μM TDZ. Fifty-four percent of the regenerated shoot rooted on MS + 5 μM naphthaleneacetic acid. Histological studies revealed that shoots regenerated via organogenesis.

Somatic embryogenesis and plant regeneration from immature zygotic embryos of Melia azedarach (Meliaceae)

SummaryA procedure for the regeneration of ‘paradise tree’ (Melia azedarach, Meliaceae) plants from immature zygotic embryos via somatic embryogenesis was developed. Somatic embryos were induced from explants cultured on Murashige and Skoog medium supplemented with 0.45, 4.54, or 13.62 μM thidiazuron. Histological examination revealed that somatic embryos were induced directly from the explants. Further development of somatic embryos was accomplished with Murashige and Skoog medium at quarter-strength with 3% sucrose. A large number of plants were regenerated from somatic embryos and successfully established in soil in a greenhouse. These plants are morphologically similar to those of seed-derived plants. This system may be beneficial for mass propagation as well as for genetic manipulation of the ‘paradise tree’.

PLANT REGENERATION, ORIGIN, AND DEVELOPMENT OF SHOOT BUDS FROM ROOT SEGMENTS OF MELIA AZEDARACH L. (MELIACEAE) SEEDLINGS

SummaryIn vitro regeneration of plants from root culture of Melia azedarach seedlings was obtained. The origin and mode of development of the regenerated shoot buds were studied by means of histological analysis and scanning electron microscopy (SEM). Maximum shoot bud regeneration was achieved when root segments were cultured on Murashige and Skoog (MS) medium at quarter strength with 3% sucrose and 0.44 μM benzyladenine (BA) and kept under light (116 μmol m−2 s−1). Shoot bud elongation was achieved on MS with 0.44 μM BA, 0.46 μM kinetin (KIN), and 3.26 μM adenine sulphate (AD). Regenerated shoots were rooted on MS with 12.26 μM indole-3-butyric acid (IBA) for 4 d and subsequently in MS lacking plant growth regulators for 26 d. Plants were established in a potting substrate. Histological analysis of roots from intact seedlings (without treatment) demonstrated that during the early life of the roots, M. azedarach lacks preformed buds. In contrast, when the roots were excised and cultured in vitro, the histology and SEM observations revealed that buds originated from meristematic groups of cells, which had been formed from the pericycle and several layers beneath. These meristematic groups of cells grew towards the periphery of the cortex by crushing the outer layer of cortical cells. Further develoment led to the differentiation of leaf primordia and a shoot apical meristem.

In Vitro Culture of Rudimentary Embryos of Ilex paraguariensis: Responses to Exogenous Cytokinins

Abstract. The effects of benzyladenine (BAP), kinetin (KIN), zeatin (ZEA), isopentenyladenine (2iP), and thidiazuron (TDZ) were studied on in vitro growth of rudimentary embryos of Ilex paraguariensis St. Hil. Heart stage zygotic embryos were removed from seeds of immature, light green fruits and cultured aseptically on quarter-strength Murashige and Skoog medium containing 3% sucrose, 0.65% agar, and supplemented with or without three concentrations of BAP, KIN, ZEA, 2iP, or TDZ. Cultures were incubated in darkness at 27 ± 2°C. Media containing 4.4 × 10−6m BAP, 4.6 × 10−6m KIN, or 4.9 × 10−6m 2iP were totally ineffective in inducing embryo growth after culture for 28 days. However, lower concentrations of these compounds (4.4 × 10−8m BAP, 4.6 × 10−8m KIN, 4.5 × 10−8m ZEA, or 4.9 × 10−8m 2iP) promoted embryo growth. TDZ at 9.9 × 10−9m, 9.9 × 10−8m, or 9.9 × 10−7m induced embryo growth at similar rates. The maximum percentage of embryos converted to seedlings was achieved when the medium was supplemented with 4.5 × 10−7m ZEA.

In vitro Plant Regeneration of Melia azedarach L.: Shoot Organogenesis from Leaf Explants

In vitro regeneration of Melia azedarach L. was studied. Shoots were regenerated from calli initiated from leaflets of in vitro growing plants. The best medium for establishment of cultures was Murashige and Skoog (MS) medium with 4.44 μM benzylaminopurine (BAP) + 0.46 μM kinetin (KIN) + 16.29 μM adenine sulphate (ADE). Regenerated shoots were multiplied in MS + 0.44 μM BAP + 0.37 μM KIN + 3.26 μM ADE. Maximal rooting of 89 % was achieved by culture of regenerated shoots in MS + 12.26 μM indole-3-butyric acid for 3 d and subsequently in MS lacking growth regulators for 27 d. Rooted shoots were acclimatized and successfully transferred to soil.

IN VITRO PLANT REGENERATION OF ILEX PARAGUARIENSIS (AQUIFOLIACEAE)

SummaryNodal segments as well as shoot tips and apical meristems of 2-yr-old “maté” plants (Ilex paraguariensis St. Hil.) were cultured in vitro to establish micropropagation systems. Maximum shoot regeneration was achieved when nodal segments were cultured with 1/4 Murashige and Skoog (MS) medium with 3% sucrose. We induced roots to differentiate by transferring the regenerated shoots onto the same medium, solidified with 2.5 g Phytagel per 1 and supplemented with indole-3-butyric acid (7.4 µM) and finally transferring shoots to 1/4 MS medium with 3% sucrose and lacking growth regulators. Plants were successfully established in soil.

Plant regeneration in Arachis pintoi (Leguminosae) through leaf culture

Abstract Plant regeneration in Arachis pintoi was obtained via two developmental pathways: organogenesis and somatic embryogenesis. Organogenic callus cultures were initiated from pieces of leaf on MS medium supplemented with NAA or 2,4-D in combination with BA, KIN or 2iP. The most suitable combination for plant regeneration through organogenesis was an initial medium composed of 10 mg/l NAA+1 mg/l BA followed by transfer of the callus to a shoot induction medium (MS+1 mg/l BA). Rooting of regenerated shoots was readily achieved by culture on MS+0.01 mg/l NAA. Embryogenic callus cultures were initiated from pieces of leaf on MS medium supplemented with PICL in combination with KIN, ZEA, BA or 2iP, and the most suitable combinations were 20 mg/l PICL+1 mg/l BA or 2iP. When pieces of embryogenic callus were subcultured on MS+1 mg/l BA, somatic embryos were differentiated and developed further into well-developed plants in MS+1 g/l AC followed by MS medium devoid of plant growth regulators.

Plant regeneration in weeping lovegrass, (Eragrostis curvula) through inflorescence culture

Plant regeneration from four genotypes of weeping lovegrass (Eragrostis curvula (Schrad.) Nees), is reported via three developmental pathways: embryogenesis, organogenesis and direct regeneration. Organogenic and embryogenic callus cultures were initiated from young inflorescence segments on Murashige and Skoogs medium supplemented with 2,4-d and BA at different concentrations. The most suitable concentrations of 2,4-d for callus growth and development were 9 and 18 μM combined with a BA concentration of 0.044 μM. Genotypical differences were observed in the morphogenetic capacity. Direct regeneration was observed under similar culture conditions (culture medium, temperature and photoperiod) but with high light intensity (66 μmol m-2 s-1). Young plants were successfully transplanted to pots and grown to maturity in the greenhouse.

Colchicine, trifluralin, and oryzalin promoted development of somatic embryos in Ilex paraguariensis (Aquifoliaceae)

A protocol for somatic embryogenesis and plant regeneration of Ilexparaguariensis St. Hil. from embryos cultures was developed. Heart stage zygotic embryos were removed from seeds of immature, light green fruit and treated with antimicrotubule agents (0.1; 0.2, and 0.5% colchicine for 24 and 48 h; 1; 10, and 20 μM of either trifluralin, ααα- trifluoro- 2,6-dinitro-N,N- dipropyl-p-toluidine, or oryzalin, 3,5-dinitro-N4, N-dipropylsulphate during 48 h). The embryos were cultured aseptically on quarter-strength Murashige and Skoog medium containing 3% sucrose, 0.65% agar (1/4MS), and 0.46 μM zeatin. Cultures were incubated in darkness at 27 ± 2 °C. All thetreatments provoked a diminution of the number of germinated embryos and in some of the treated embryos somatic embryogenesis was induced. Somatic embryo maturation and conversion into whole plants could be achieved by culturing the embryos on 1/4MS lacking hormones and incubated at 27 ± 2 °C, 14 h photoperiod (116 μmol m-2s-1). Mostof the plants regenerated from somatic embryos appeared morphologically normaland grew under greenhouse conditions. Only 2 plants out of 152 studied contained the tetraploid number of the chromosomes (2n = 4x = 80), meanwhile the rest of the plants had the normal diploid number of chromosomes (2n =2x = 40). Somatic embryos with abnormal morphology were also observed.