In Vitro Multiple Shoot Regeneration in Syzygium aromaticum

Multiple shoots were induced from nodal segments of seedlingsof Syzygium aromaticum, on Murashige and Skoog's (MS) basalmedium at half strength salts and Gamborg's medium (B5), supplementedwith BAP and NAA. Six to eight shoots were obtained when 3 mgl^–1 BAP and 0.5 mg l^–1 NAA were used in the medium.Both MS medium and B5 medium showed more or less similar resultsregarding the proliferation of the explants. Syzygium aromaticum (L) Merr and Perry (clove), multiple shoots, regeneration     

Regeneration of Lasiurus scindicus from Tissue Culture

Embryogenic callus cultures were initiated from mature embryosof Lasiurus scindicus on Murashige and Skoog's medium supplementedwith 6 mg l^–1 2,4-Dichlorophenoxyacetic acid (2,4-D).These cultures were maintained on 2 mg l^–1 2,4-D. Plantletswere regenerated via somatic embryogenesis when the calli weretransferred onto hormone-free MS basal medium. Young plantswere successfully transplanted to pots and grown to maturityin a greenhouse. Grass, Lasiurus scindicus, Thar Desert, drought tolerant, somatic embryogenesis, plant regeneration     

Somatic Embryogenesis from Clonal Leaf Tissues of Cassava

Leaf lobes were isolated from palmate leaves of clonal cassava(Manihot esculenta Crantz) material growing in vitro or in glasshouseconditions and subjected to a two-stage culture procedure involvingincubation on Murashige and Skoog (MS2) basal medium supplementedwith 2–12 mg l^–1 2,4-D for 20 d (Stage I) beforetransfer to MS2 basal medium supplemented with 0.01 mg l^–12,4-D and 0.1 mg l^–1 6-benzylamino purine (BAP) (StageII medium). Embryogenetic tissues, foliose structures and somatic embryosdeveloped from leaf lobes at all Stage I 2,4-D concentrations,except on those explants isolated from shoot-tip cultures incubatedon MS2 basal medium supplemented with 0.1 mg l^–1 NAA and1.0 mg l^–1 BAP. Leaf lobes isolated directly from glasshouse plants showed optimalembryogenetic competence when subjected to a Stage I cultureperiod of 17 d, although foliose structure initiation was optimalwith shorter Stage I durations. Leaf lobes of 2–4 mm lengthand those isolated from phyllotaxic leaf numbers 4 and 5 showedthe greatest embryogenetic competence. Manihot esculenta, cassava, somatic embryogenesis, tissue culture, morphogenetic competence     

In vitro Regeneration of Mustard Plants (Brassica juncea var. RAI-5) on Cotyledon Explants from Non-irradiated, Irradiated and Mutagen-treated seed

Shoot formation in cotyledon explants of mustard (Brassica junceavar. Rai-5) was observed on Murashige and Skoog's medium supplementedwith NAA^* (1 mg l–1) and BA (1 mg l–1). Hypocotylsegments failed to differentiate shoots. Complete plants wereobtained when shoots were rooted in MS medium with NAA (1 mgl–1). EMS, a chemical mutagen, had an inhibitory effecton shoot regeneration. Gamma rays in doses above 2 kR suppressedshoot regeneration but stimulated callus growth. Brassica juncea, mustard, regeneration, tissue culture     

Callus Culture of Sainfoin (Onobrychis viciifolia) and Plant Regeneration through Somatic Embryogenesis

Callus of sainfoin (Onobrychis viciifolia Scop.) was initiatedfrom stem and root explants which were obtained from seedlingsgrowing in vitro, on Linsmaier Skoog (LS) medium supplementedwith 1 mg l^–1 2, 4-D and 1 mg l^–1 BA or only 1 mgl^–1 BA, and the Vacin and Went medium without hormones.Somatic embryos were formed on LS medium containing 1 m l^–1BA. Embryos developed into complete plants on filter paper saturatedwith hormone-free LS medium. Onobrychis viciifolia, somatic embryogenesis, callus culture, plant regeneration     

Micropropagation of Pissardi Plum

An efficient medium for multiplication of Prunus cerasifera,J. F. Ehrh. cv. ‘Atropurpurea’, Pissardi plum, consistedof Linsmaier-Skoog basal medium (LS) supplemented with 3% sucrose,10mg 1^–1 N⁴-(²-isopentenyl) adenine (2iP), and 162 mg1^–1 phloroglucinol (Phl). Phl in the medium significantlyenhanced growth (P < 0.1 %) over cultures maintained on LSmedium with 10 mg 1^–1 2iP and no Phl. Plantlets were rootedon a half-strength Murashige-Skoog (MS) medium supplementedwith 0.2 mg 1^–1 indole-3-butyric acid (IBA). Prunus cerasifera, Pissardi plum, micropropagation, phloroglucinol, N⁶-(²-isopentenyl) adenine     

Somatic Embryogenesis and Plant Regeneration from Inflorescence Culture of Java Citronella: (Cymbopogon winterianus)

Embryogenic callus was induced from immature inflorescence segmentsof Java citronella (Cymbopogon winterianus) and maintained for2 years on Murashige and Skoog's medium supplemented with 2,4-D(l mg l^–1). The callus cells retained the original chromosomenumber of 2n = 20. The somatic embryos germinated into plantletson MS basal medium or medium with IAA, NAA, BAP or KN individually(l mg l^–1). The regenerated plantlets developed a goodroot system on full strength solid MS inorganics medium withIAA (1 mg l^–1). The regenerated plants were similar tothe donor plant in morphology and had the same chromosome number,but showed some variation in the essential oil content. Java citronella, Cymbopogon winterianus, somatic embryogenesis, regeneration, inflorescence culture     

In Vitro Micropropagation of Passiflora edulis (Purple Passionfruit)

A technique for the micropropagation of Passiflora edulis fromseedling explants is described. A multiplication rate of 4.5per month can be achieved in an Murashige and Skoog medium supplementedwith 2 % sucrose and 2 mg l^–1 BAP. Using this technique,no callus is produced. Problems of chlorosis and a method foravoiding subsequent death after prolonged culture are discussed. Passionfruit, Passiflora edulis, micropropagation, tissue culture, chlorosis     

Growth Stimulation by Conditioned Medium and Spermidine in Low-Density Suspension Cultures of Rice

Suspension cultures of Oryza sativa L. var IR 20 grew in Murashigeand Skoog medium (MS) supplemented with 2,4-D and kinetin ina density-dependant manner with a critical minimum inoculation-densityof 10,000 cells ml^–1. Conditioned medium obtained fromthese cultures and added to MS+2,4-D+kinetin induced the growthof cultures at 1,000 cells ml^–1. Growth stimulation byconditioned medium was mimicked by spermidine but not by otherpolyamines viz. putrescine and spermine. This is the first reportof a polyamine substituting for conditioned medium in cultures. ² Present address: Vice-Chancellor, Pondicherry University,Pondicherry 605 014, India.     

Determinate Floral Buds of Plantain (Musa AAB) as a Site of Adventitious Shoot Formation

Floral buds of the ‘False Horn’ plantain clonesMusa (AAB) ‘Harton Verde’, ‘Harton Negra’,and ‘Currare’ terminate in a large single floralstructure. The apices of these floral buds are here designatedas determinate since they have lost the ability to produce additionalfloral initials or buds. Terminal peduncle segments can be culturedin a modified Murashige and Skoog (1962) medium supplementedwith N⁶-benzyl-aminopurine (5 mg I^–1). Under these conditions,this apparent inability to yield buds can be overcome as vegetativeshoot clusters form in the axils of the bracts. Rooted plantletsare obtainable by treating shoots with naphthaleneacetic acid(1 mg I^–1) and activated charcoal (0.025%). The adventitiousorigin of the shoots has been established. Musa cultivars, plantains, floral bud, adventitious buds, tissue culture     

Tissue 3Vicia Faba

The establishment of a callus culture of Vicia faba root tissuegrowing on a modified Bonner and Devirian medium supplementedwith 10^-6 M 2,4-dichlorophenoxyacetic acid and 5.0 g/l yeastextract is reported. Experiments have been carried out to determinethe optimum growth conditions of the callus on semisolid mediaand an investigation made of the potential of this materialas an initial source of friable tissue yielding cell suspensionson dissociation in liquid medium. The development of two morphologicallydistinct types of callus tissue was observed on semi-solid mediumand chemical analysis has demonstrated a correlation betweenthe friability and cell-wall composition of these tissues.     

Multiplication In Vitro of Limonium estevei Fdez. Casas

Plantlets of Limonium estevei Fdez. Casas, an endangered Spanishspecies, were successfully regenerated from nodal segments excisedfrom young seedlings. Initiation of multiple adventitious budswere obtained in MS modified medium plus 1 mg l^–1 IBAand 0·1 mg l^–1 BAP. Rooting was achieved by transferof the isolated shoots to fresh MS medium without plant growthregulators. Fully grown plants were established in a pottingmix and are growing well in a greenhouse. Limonium estevei, in vitro multiplication, adventitious regeneration     

Plantlets from somatic callus tissue of the East Indian Rosewood (Dalbergia latifolia Roxb.)

Callus-mediated shoot bud formation was demonstrated in Dalbergia latifolia Roxb. (East Indian Rosewood). Cultures were raised from shoot explants of six year-old plants on Murashige and Skoog (MS) medium supplemented with naphthaleneacetic acid (NAA) and benzyladenine (BA). A sequential treatment of callus with increasing BA levels and decreasing NAA ensured shoot bud induction. Rooting of shoots was achieved by a three-step culture procedure involving 1) White's(W) liquid medium containing indoleacetic acid (IAA), naphthaleneacetic acid and indolebutyric acid (IBA), 2) half-strength MS agar-solidified medium with charcoal (0.25%) and 3) half-strength MS liquid medium.Abbreviations BA Benzyladenine - IAA Indoleacetic acid - IBA Indolebutyric acid - MS Murashige and Skoog - NAA a-naphthaleneacetic acid - PVP Polyvinylpyrrolidone - W White's medium - 2,4-D 2,4-dichlorophenoxyacetic acid     

Effect of Sudden Salt Stress on Ion Fluxes in Intact Wheat Suspension Cells

Although salinity is one of the major problems limiting agriculturalproduction around the world, the underlying mechanisms of highNaCl perception and tolerance are still poorly understood. Theeffects of different bathing solutions and fusicoccin (FC),a known activator of plasma membrane ATPase, on plasma membranepotential (E_m) and net fluxes of Na⁺, K⁺and H⁺were studied inwheat suspension cells (Triticum aestivum) in response to differentNaCl treatments. E_mof cells in Murashige and Skoog (MS) mediumwas less negative than in cells exposed to a medium containing10 mM KCl + 0.1 m M CaCl₂(KSM) and to a basic salt medium (BSM),containing 1 m M KCl and 0.1 m M CaCl₂. Multiphasic Na⁺accumulationin cells was observed, peaking at 13 min after addition of 120m M NaCl to MS medium. This time scale was in good agreementwith net Na⁺flux changes measured non-invasively by moving ion-selectivemicroelectrodes (the MIFE system). When 120 m M NaCl was addedto all media studied, a quick rise of Na⁺influx was reversedwithin the first 20 min. In both 120 and 20 m M NaCl treatmentsin MS medium, net Na⁺efflux was observed, indicating that activeNa⁺transporters function in the plant cell response to saltstress. Lower external K⁺concentrations (KSM and BSM) and FCpre-treatment caused shifts in Na⁺fluxes towards net influxat 120 m M NaCl stress. Copyright 2000 Annals of Botany Company Sodium, potassium, proton, membrane potential, fusicoccin, salt stress, wheat, Triticum aestivum     

In vitro Culture of Abscissed Immature Avocado Embryos

The efficiency of an avocado hybridization programme, usingsmall potted glasshouse plants, was reduced by a high rate ofabscission of immature fruitlets bearing embryos too young forconventional germination. This was overcome in part by culturein vitro of the shed embryos on a liquid medium supplementedwith 0.5 mg l^–1 benzyladenine. Most embryos younger than6 weeks did not survive in culture, but older embryos slowlyproduced multiple shoots, with axillary shoot growth being furtherstimulated by removal of both cotyledons. Embryo response wasnot related to cultivar. Shoots removed from culture could begrafted to seedling rootstocks. Grafting was considered morereliable than dependence on growth of the main root or adventitiousroots in vitro to produce established plants. Persea americana Miller, avocado, abscissed fruitlets, in vitro embryo culture     

Early Cellular Events During Direct Somatic Embryogenesis in Cotyledon Explants of Solanum aviculare Forst.

A detailed light and electron microscope study of early cellularevents at the onset of somatic embryogenesis in cotyledon explantsof Solanum aviculare Forst., cultured on MS medium supplementedwith 1 mg l^–1 2,4-dichloro-phenoxyacetic acid (2,4-D)for periods of 0–12 d in darkness, is described. Examinationsof longitudinal sections in a plane offset from the centralveins indicated that the earliest embryogenic events in explantstook place within the first 3–4 d of culture in the parenchymacells associated with the vascular traces closest to the cutbasal ends of cotyledons. Thereafter, parenchyma associatedwith more distal vascular traces became active in an apparentlysequential manner such that, by the second week of culture,progressive stages of embryogenesis could be observed alongthe lengths of cotyledon sections. Despite the fact that epidermalcells and palisade tissues were exposed directly to the 2,4-Dmedium, initiation of embryogenic development was never observedin cells other than those directly associated with vasculartraces. None of the embryogenic events characterized at theultrastructural level were observed in cotyledons cultured onMS medium in the absence of 2,4-D with the exception that starchaccumulated in decreasing amounts from the wounded basal endto the distal end of each cotyledon. This system provides a valuable model with which to study earlybiochemical and molecular events occurring in explants duringthe onset of somatic embryogenesis because they occur in a predictablefashion at sequentially situated sites along the explant tissues. Somatic embryogenesis, Solanum aviculare, cotyledon explants, cellular changes     

Direct Organogenesis from Petiole and Thin Cell Layer Explants in Sugar Beet Cultured In Vitro

Detrez, C., Tetu, T., Sangwan, R. S. and Sangwan-Norreel, B.S., 1988. Direct organogenesis from petiole and thin cell layerexplants in sugar beet cultured in vitro.—J. exp. Bot.39: 917–926. Plant regeneration was obtained by direct bud formation frompetiole as well as from thin cell layer explants taken fromsugar beet (Beta vulgaris L.) plants grown in vitro. The budswere mainly induced in the blade-petiole transition zone ofthe explants. High frequency bud regeneration was observed inpetiole and thin layer explants of 10 different breeding linesof sugar beet tested. Organogenesis resulted when petiole explantsexcised from 8-d-old seedlings grown on half-strength Murashigeand Skoog medium (MS) containing 3.0 mg dm^–3 naphthaleneacetic acid (NAA), 3.0 mg dm^–3 6-benzylaminopurine (BAP)and 1.0 mg dm^–3 2, 3, 5, triiodobenzoic acid (TIBA) werecultured on MS with 3.0 mg dm^–3 NAA and 3.0 mg dm^–3BAP. Thin cell layer strips isolated from shoot apices culturedon MS medium supplemented with 0–9 mg dm^–3 BAP or1.0 mg dm^–3 indolebutyric acid (IBA) formed adventitiousbuds on MS medium containing 0–5 mg dm^–3 NAA + 5.0mg dm^–3 BAP. Histological studies confirmed the sub-epidermalorigin of shoots. Key words: Beta vulgaris, direct organogenesis, in vitro culture, petiole, regeneration, thin cell layer     

Somatic Embryogenesis and Its Hormonal Regulation in Tissue Cultures of Freesia refracta

Somatic embryogenesis can be induced in tissue cultures of Freesiarefracta either directly from the epidermal cells of explants,or indirectly via intervening callus. These two pathways ofsomatic embryogenesis can be controlled and regulated by varyingthe combinations and levels of exogenous hormones. When younginflorescence segments were cultured in vitro on modified N₄(MN₄) medium supplemented with 2 mg l^–1 indoleacetic acid(IAA) and 3 mg l^–1 6-benzylaminopurine (BAP), some ofthe epidermal cells began to exhibit the features of embryogeniccells. These cells produced embryoids and developed into newplants through direct somatic embryogenesis. If the same explantswere placed on Murashige and Skoog's (MS) medium containing2 mg l^–1 IAA, 05 mg l^–1 BAP and 05 mg l^–1naphthaleneacetic acid (NAA), pale-yellow translucent nodularcalluses appeared on the surface of the explants. When thiskind of callus was transferred to MN6 medium with 2 mg l^–1IAA and 3 mg l^–1 BAP, embryoids formed which further developedinto plantlets. The regenerated plants were morphologicallynormal and possessed the normal diploid chromosome number of2n = 22. A similar result has also been obtained with youngleaf explants of this plant. The early segmentations of embryogeniccells and the development of embryoids were studied using histologicaland scanning electron microscopic techniques, and the resultshave been discussed in association with the ontogeny and originof the embryoids. Freesia refracta Klatt, somatic embryogenesis, plant regeneration, exogenous hormones     

In Vitro Embryo Culture and Induction of Multiple Shoots in Lychee (Litchi chinensis Sonn.)

Immature embryos of different sizes and ages from commercialvarieties of lychee (Litchi chinensis Sonn.) were cultured ina range of different media. Embryos as small as 3 mm could becultured using in vitro techniques and subsequently grown intoplants. MS solid medium with 2% sucrose supplemented with 150ml l^–1 coconut water was most effective in stimulatingthe germination of immature lychee embryos. Embryos of lycheewere treated to induce adventitious buds from embryonic shootsas a means of achieving multiplication. The different varietiesexhibited differences in response, with Bengal embryonic shootsproducing 15 adventitious buds after pretreatment with 100 mgl^–1 BAP for 3 h. Root formation was achieved in 65% ofadventitious shoots using MS medium supplemented with 0.5 mgl^–1 NAA and activated charcoal. These plants were successfullydeflasked and grown on in the glasshouse. This technique providesof means of producing some multiple shoots from lychee embryosand has value for multiplication in a breeding program wherea method of micropropagation is unavailable. Litchi chinensis Sonn., lychee, embryo culture, multiple shoots, in vitro     

Effects of Sodium Chloride Stress on Callus Cultures of Cicer arietinum L. cv. BG-203: Growth and Ion Accumulation

Growth and ion accumulation were measured in callus culturesof Cicer arietinum L. cv. BG-203, grown on media supplementedwith 0–200 mol m^–3 NaCl. Fresh and dry weights decreasedat concentrations ranging from 100–200 mol m^–3,the reduction being greater during the third and fourth weeksof culture. Slight stimulation of growth was observed at 25and 50 mol m^–3 NaCl. There was also a decrease in tissuewater content (fresh weight: dry weight) at 100–200 molm^–3 NaCl. The concentration of Na⁺ and Cl^– in thetissue increased with increasing salinity of the medium. Mostof the accumulation of these ions occurred by the first weekwhile significant growth inhibition became apparent by onlythe third week of culture. Tissue K⁺ and Mg²⁺ decreased withincreasing salinization, the decrease being greater in K⁺ levels.Levels of Ca²⁺, however, were maintained throughout the experimentalrange. Key words: Cicer arietinum, NaCl stress, Callus cultures, Ion accumulation