In Vitro Culture and Plant Regeneration in Vicia faba subsp. Equina (var. Spring Blaze)

Explants of stem, leaves, roots, and cotyledons from etiolatedaxenically grown Vicia faba seedlings were cultured on a rangeof media. Shoot organogenesis was only obtained with nodal stemand cotyledonary node explants when cultured on MS medium with3% sucrose, 2.0 mg 1^–1 BAP and 02 mg 1^–1 NAA. Callusproliferation accompanied shoot organogenesis from nodal stemexplants. Successive subculture of nodal stem callus resultedin proliferation of regenerative callus which contained severalshoot bud initials. The capacity for shoot regeneration fromthis callus was maintained for 9 months. Histological studiesreveal de novo formation of meristematic centres in callus andtheir further development into bud primordia. High frequencyrooting of these adventitious shoots was obtained on half-strengthMS medium with 1.5% sucrose, 0.1 mg 1^–1 NAA and 0.5 mg1^–1 kinetin. Key words: Vicia faba, adventitious shoots, axillary shoots, de novomeristem formation, organogenesis, tissue culture     

Callus Initiation and Organized Development from Zamia pumila Embryo Explants

Explants derived from Zamia pumila embryos were cultured ona Murashige and Skoog basal medium supplemented with naphthaleneaceticacid (NAA), N⁴-benzylaminopurine (BAP), or combinations of thetwo at 27 °C in darkness. NAA was invariably required forcallus initiation, and its minimal effective concentration was0.1 mg l^–1. BAP was not always required, and dependingon the explant type and NAA concentration, BAP either enhanced,suppressed, or had little effect on the frequency of callusinitiation. High frequency of callus initiation occurred with1.0 mg l^–1 NAA combined with 0.01 or 1.0 mg l^–1BAP. When the concentration of NAA was high relative to thatof BAP, friable callus was produced. As the relative BAP concentrationwas increased, a more compact callus formed. Compact-nodularcallus developed at equal concentrations of NAA and BAP overa wide range of absolute concentrations. Friable callus formedroots only. Compact-nodular callus formed roots, shoots andembryo-like structures. Root and shoot formation predominatedand were of nearly equal frequency. Formation of embryo-likestructures was infrequent. Zamia pumila, callus differentiation, callus formation, embryo culture, naphthaleneacetic acid, N⁴-benzylaminopurine     

Morphogenesis in Cultured Floral Parts of African Violet

Callus tissue was induced from floral parts of African violetcultured on MS medium containing NAA (2 mg I^–1) and BAP(0.2 mg I^–1). When maintained on this medium in the presenceof light, the callus produced many shoots and roots. Large numbersof adventitious shoot buds were formed apparently in the absenceof callusing when ovary, sepal, and petal tissue was culturedon MS medium supplemented with BAP (1 mg I^–1) and NAA(1 mg I^–1). In contrast, culturing the same floral partson MS medium augmented with kinetin (1 mg I^–1) and NAA(0.5 mg I^–1) and NAA (0.5 mg 1-¹) led to the profuse developmentof roots. Organs seemed to be initiated from the epidermis ofcultured floral parts and did not appear to be related to particularcells or loci. Transfer of shoots to MS medium deviod of growthsubstances resulted in the formation of plantlets, which ata height of 3 cm could be transferred to soil and grown to maturitywithout variation in morophology or cytology.     

In vitro Propagation of Narcissus

Adventitious shoots were induced on leaf, scale and stem explantstaken from the basal plate region of flowering-size bulbs onmedia containing 2–16 mg l^–1 6-benzylaminopurineand 0·25–4·0 mg 1^–1 1-naphthal-eneacetic acid (NAA). Only the levels of NAA had a significanteffect on the numbers of shoots produced. When trimmed and splitin half, 6 mm or more diameter in vitro shoots regenerated furtheradventitious shoots which in turn grew in size suitable forsplitting within 16 weeks. The vigour of the first generation of shoots was proportionalto the hormone levels used for their initiation. All shootseventually declined in vigour, senesced and formed dormant bulbils.Split senescent shoots regenerated only a few secondary shootswhich quickly became senescent. A total of 500–2000 bulbilscould be obtained from each initial bulb within 18 months. Bulbilsrequired 10 weeks at low temperature before planting to breakdormancy. Histological observations showed that in twin scales and splitshoots, adventitious shoots were regenerated from at least twosuperficial layers of menstematic cells near to the basal plate.This multicellular mode of origin suggests that plants multipliedfrom in vitro adventitious shoots could be as genetically uniformas those from natural vegetative increase. Narcissus, tissue culture, propagation, adventitious shoots, histology     

Production of Plantlets of Shorea roxburghii G. Don. from Embryonic Axes Cultured In Vitro

Development of axillary shoots was induced in embryonic axesof the dipterocarp Shorea roxburghii G. Don. cultured on a modifiedMS medium containing 6-benzyl-aminopurine (BAP) at an optimumconcentration of 5 mg I^–1. Excised axillary shoots wereused in multiplication and rooting experiments. Vigorous rootdevelopment occurred in shoots supported on filter paper bridgesin liquid medium containing naphthaleneacetic acid (NAA) andindolebutyric acid (IBA) (0.1 mg I^–1 each). Shorea roxburghii, Dipterocarpaceae, tissue culture, plantlet formation     

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     

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     

Morphogenic Potential of Cultured Explants of Crocus chrysanthus Herbert. cv. E. P. Bowles

Explants of leaves, basal plates, petals, anthers and ovariesof young growing corms of Crocus chrysanthus var. E. P. Bowleswere cultured on MS basal media with 20 different combinationsof either kinetin and NAA or BAP and 2, 4-D in the dark. Nomajor change was observed except on ovary explants. The ovaryexplants produced callus at 5.0 mg 1^–1 and 10 mg^–1BAP and subsequently stigma-like structures formed on the surfaceof the callus. Transfer to light resulted in the stigma-likestructures developing a yellow pigmentation whereupon they cameto resemble the naturally-grown stigmas. Corm formation andshoot regeneration was obtained from the callus when the ovaryexplants were cultured on media containing 5.0 and 10 mg I^–1BAP with 0.5 mg 1^–1 2, 4-D. Increasing the level of 2,4-D markedly reduced the number of shoots produced per explant. Key words: Crocus chrysanthus, callus, ovary explants     

Effects of Growth Regulators and Glutamine on In Vitro Development of Zygotic Embryos of Taro (Colocasia esculenta var. antiquorum)

Zygotic embryos of taro, Colocasia esculenta var. antiquorumcultured on Linsmaier-Skoog (LS) medium without the additionof hormones develop into mature plants only in the presenceof endosperm tissue. Growth is usually evident within the firstweek of culture when embryos swell and become green. Embryosexcised from endosperm and cultured on LS containing 0-01 mg1^–1 naphthaleneacetic acid (NAA), and 0–01 mg 1^–16-dimethylaminopurine (6-DMAP) grow at a rate comparable withcontrols for the first week of culture. During the second week,growth rates are higher than controls primarily because embryosform elongated hypocotyl regions which often produce swollentissues and/or callus. In the presence of 200 mg 1^–1 glutamineand a range of concentrations of 6-dimethylaminopurine, benzyladenine,or NAA, elongation of the hypocotyl axis is inhibited, and acompact callus may develop. Embryos grown on LS containing 200mg 1^–1 glutamine and 2.0 mg 1^–1 2, 4, 5-trichlorophenoxyaceticacid form friable callus which was used to generate short-livedsuspension cultures. Growth Regulators, Glutamine, tygotic embryos, Colocasia esculenta, endosperm     

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     

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     

Studies on the Formation of Roots and Shoots in Wheat Callus Cultures

Callus tissues were initiated from root, embryo and inflorescenceexplants of wheat. These callus cultures were used to studythe formation of roots and shoots in the absence and presenceof selected plant hormones. On a basal medium alone, only newly-initiatedembryo callus formed both roots and shoots while root callusonly formed roots. Inflorescence callus showed no signs of differentiation.The regenerative capacity of root and embryo callus tissueson medium lacking hormones decreased with increasing periodsof culture. Calluses which failed to differentiate in the absenceof hormones were selected for studies on hormone-mediated differentiation.NAA (1 mg 1^–1) was effective in inducing roots from allcalluses irrespective of their origin or age. In contrast, shootformation was elicited by incubating newly-formed callus onbasal medium supplemented with kinetin (5 mg 1^–1) andNAA (1 mg 1^–1) but rapidly decreased with longer periodsof culture. The differences observed in differentiation of thecallus in the absence and presence of hormones is discussed.     

Differentiation of Shoot Buds in vitro in Tissue 1Sisymbrium irio L.

Shoot bud formation was induced in the stem callus of Sisymbriumirio L., a Cruciferous plant. The callus was established onMurashige and Skoog medium with IAA (1?0 mg l^–1) and kinetin(0?5 mg l^–1). The effect of three purines (kinetin, 6-benzylaminopurine,and 6-methylaminopurine) incorporated singly along with IAAin MS medium was investigated. It was found that kinetin orMAP (3–5 mg l^–1) along with IAA (0?5 mg l^–1)were the most effective in inducing shoot bud formation. Adeninesulphate (10 mg l^–1) with kinetin (1?0 mg l^–1) alsoinduced bud differentiation. The morphogenetic potential of the callus to differentiate shootbuds was seemingly lost in 2 year old callus cultures. However,on successively subculturing on a regeneration medium shootbuds differentiated and the number of buds formed improved onfurther subculture. Two types of meristematic outgrowths were recognized: (i) arisingfrom superficial cells and (ii) arising from deep-seated cellsin the vicinity of tracheidal elements. However, both typesformed meristematic nodules on the surface of which shoot budsdifferentiated. Some embryoids were also recognized arisingsuperficially.     

Haploid Plantlet Formation from Female Gametophytes of Ephedra foliata Boiss. in vitro

Female gametophytes (at the archegonial stage) excised fromyoung ovules of Ephedra foliata Boiss, were cultured on a basalmedium (Murnshige and Skoog's combinations of major and minorsalts, Iron source, vitamins, myo-inositol along with 2 percent sucrose and 10 per cent coconut milk) under aseptic conditions.Growth and morphogenetic responses of the explants to auxinswere compared at different concentrations and a study of theirinteractions with cytokinins has also been made. At 2 mg 1^–1,2, 4-D induced profuse callusing which subsequently producedroots. NAA at 4 mg 1^–1 was optimal for callus growth androoting. Combinations of 2,4-D and kinetin were more effectivein inducing roots and shoot buds than those of 2,4-D and benzylamino-purine (BAP). Addition of BAP (0.05 mg ^1–1) to themedium containing optimal concentrations of NAA resulted information of a large number of roots. Kinetin induced only rootingin the presence of 4 mg 1^–1 NAA. A high concentrationof BAP (8 mg 1^–1), stimulated shoot bud formation. Forthe further development of shoot buds, neither auxin nor cytokininwas needed. Cytological observations revealed the presence ofhaploid number of chromosomes, i.e. seven. Ephedra foliata, tissue culture, callus, regeneration, 2,4-dichlorophenoxyacetic acid, naphthalene acetic acid, kinetin, benzyl amino-purine     

Effect of External Supply of Growth Substances on Axillary Proliferation and Development in Dioscorea bulbifera

Bulbil development in cultured nodes of D. bulbifera proceededin the absence of growth substances from the medium. When IAAwas incorporated into the medium at the concentrations of 5mg l^–1 and 10 mg l^–1 the cultured nodes producedlarger bulbils than in its absences. When the concentrationof IAA was increased to 15 mg l^–1, however, the culturednodes produced a callus instead of a properly organized bulbil.The dry weight of bulbils increased when kinetin was added tothe medium at the concentrations of 0.05, 0.5, and 2.5 mg l^–1.The greatest increase was with 0.5 mg l^–1 kinetin. Onincreasing the concentration of kinetin in the medium to 5.0mg l^–1 the tissue produced had smaller dry weight thanthose produced in the absence of growth substances. Additionof different combinations of IAA and kinetin to the basal mediumresulted in the production of normal bulbils, roots, and shootsin some instances (suitable combinations) and in the productionof callus and abnormal shoots in others (non suitable combinations).     

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     

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     

Regeneration of Shoots on Root Explants of Flax

Regeneration of shoots from adventitious root explants of flaxwas achieved for two of five cultivars tested. Root explantsof the cultivar Bombay regenerated buds on MS medium with varioussupplements, the best combination being 002 mg I^–1 NAA,1 mg I^– 6-BA, 20 mg I^– adenine and 500 mg I^–cefotaxime. Adenine, in the presence of 6-BA, and cefotaxime,in the presence of both 6-BA and adenine, were found to stimulatebud initiation, but the most important factor influencing budregeneration from flax roots was genotype     

Shoot and Plantlet Formation in Organ and Callus Cultures of Albizzia lebbek Benth

Plantlets were produced in vitro from root and hypocotyl explantstaken from seedlings of the tree legume, Albizzia lebbek. Theseexplants formed shoots when cultured with 5.0 mg l^–1 kinetinand 1.0 mg l^–1 IAA in MS medium. Shoots were also inducedin large numbers from callus treated with benzylaminopurine.About 20 per cent of the shoots rooted and were grown into plants. Albizzia lebbek Benth, tree legume, hypocotyl, root, in vitro cultures, shoot-plantlet induction     

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