In vitro plant regeneration from callus derived from root explants of Lathyrus sativus

Protocols have been developed for the in vitro production of plants from callus derived from root explants of Lathyrus sativus cv. P-24. Callus and shoot regeneration were achieved only in MS medium supplemented with 10.7 M naphthaleneacetic acid and an increased concentration of kinetin (0.9 M for 14 days to 1.4 M for 18 days) during callusing. The shoots obtained rooted in 1/2 MS supplemented with 0.5 M indolebutyric acid. During the year plants have been regenerated several times. The requirement for growth regulators is very specific and narrow.Abbreviations IAA indoleacetic acid - IBA indolebutyric acid - NAA naphthaleneacetic acid - BA benzyladenine     

Effect of plant growth regulators on somatic embryogenesis in leaf cultures of Coffea canephora

The effects of plant growth regulators on somatic embryogenesis were studied in leaf cultures of Coffea canephora. The maximum number of somatic embryos were obtained on media that contained only cytokinin as a plant growth regulator. All of the auxins tested (NAA, IBA, IAA and 2, 4-D) inhibited the formation of embryos. The optimal concentration of each cytokinin (2-iP, BA and kinetin) for somatic embryogenesis was 5 M. Under optimal conditions, each explant formed more than 100 embryoids with little callus and few adventitious roots. Embryoids were formed only at the cut edges of the leaf discs. Cytokinins were absorbed only at the cut edges of leaf discs that were in contact with the medium, and were not transported to other parts of the explant.Abbreviations 2-iP iso-pentenyladenine - BA benzyladenine - NAA 1-naphthaleneacetic acid - IBA indole-3-butyric acid - IAA indole-3-acetic acid - 2,4-D 2,4-dichlorophenoxyacetic acid     

In vitro plant regeneration in lateral bud explants of Cryptanthus bromelioides Var. Tricolor M. B. Foster

Lateral buds of Cryptanthus bromelioides var. tricolor variegated ornamental bromeliad, were cultured on Murashige and Skoog's basal medium (MS) supplemented with napthaleneacetic acid, indolebutyric acid, and kinetin, each at 2 mg/l. The basal part of the cultured buds proliferated and produced adventitious shoots. The shoots developed into plants of three different phenotypes: (a) entirely green, (b) striated (green and yellowish-white) and (c) yellowish-white. Rooting could be induced in (a) and (b) types but successful transplantation was possible only with green plants some of which underwent color changes after establishment in soil.Abbreviations BA benzyladenine; 2,4-D, 2,4-dichlorophenoxyacetic acid - IAA indoleacetic acid - IBA indolebutyric acid; 2,i-P, 6 -dimethylallylaminopurine; Kn, kinetin - NAA naphthaleneacetic acid, Z, zeatin     

Plant regeneration by somatic embryogenesis from callus cultures of sweet sorghum

Tissue culture methods were developed for the induction, maintenance, and regeneration of embryogenic callus in sweet sorghum (Sorghum bicolor) cultivars Keller, Rio, and Wray. No significant differences were observed in production of embryogenic callus in cultures established from developmentally immature or mature embryo explants cultured on LS medium with 2 mg/1 2,4-D plus 0.5 mg/1 kinetin. Prolific callus production did not occur until the third four-week culture period. Long-term maintenance of embryogenic callus was dependent upon the selective transfer of embryogenic callus, with other callus types discarded. High-frequency plant regeneration was achieved and quantified on a fresh weight basis of embryogenic callus.Abbreviations 2,4-D 2,4-dichlorophenoxyacetic acid - BA benzyladenine - IAA indoleacetic acid - IBA indolebutyric acid - LS Linsmaier and Skoog basal medium (Linsmaier and Skoog, 1965)     

Studies on a drought resistant legume: The moth bean,Vigna aconitifolia (Jacq) marechal. II. morphogenetic studies

Plantlets regenerated from shoot apices, cotyledons and callus cultures in Moth bean, Vigna aconitifolia (JACQ) Marechal, a drought resistant legume and pulse crop, were rooted and transferred to soil. Explants for these studies were derived from seedlings pre-conditioned by germination of seeds on B5BA and WMB (control).Abbreviations MS Murashige and Skoog (1962) - B5 B5 basal medium (Gamborg et al 1968) - B5BA B5 basal medium containing BA (2.25 mg/l) - WMB Modified White's medium (Mascarenhas et al 1976) - BA 6-benzyladenine - IAA indole-3-acetic acid - NAA 1-napthaleneaceticacid - 2,4-D 2,4-dichlorophenoxyacetic acid - IBA indolebutyric acid - 2iP N(^–2 isopentyl) adenine - CM coconut milk NCL Communication No. 3375     

High frequency embryoid and plantlet formation from tissue cultures of the Finger millet-Eleusine coracana (L.) Gaertn

Compact nodulated embryogenic callus differentiated from cultured seeds of Eleusine coracana (Finger Millet) on Murashige and Skoog (1962) basal medium with 2,4-dichlorophenoxyacetic acid (1.0, 3.0 mg ^l). This embryogenic callus was maintained on a medium with a lower level of 2,4 — dichlorophenoxyacetic acid. At every subculture the embryogenic callus had some preexisting embryoids in it. With this method of subculture the callus has retained its morphogenic potential for four years. Following transfer to media with different levels of auxins and cytokinins, the callus showed varied patterns of growth and morphogenesis. Embryoids could be germinated in profusion to form plantlets which could be transferred to the field. Shoot buds also differentiated from the whole surface of the embryoid or from the flattened meristemoids.Abbreviations BAP 6-benzylaminopurine - IAA indole-3-acetic acid - 2,4-D 2,4-dichlorophenoxyacetic acid - NAA napthaleneacetic acid - IBA indolebutyric acid - KN kinetin - MS Murashige and Skoog (1962) - GA₃ Gibberellic acid     

Production of the triterpenoid quassin in callus and cell suspension cultures of Picrasma quassioides Bennett

Plant cell and suspension cultures have been established from stem cuttings of Picrasma quassioides Bennett. The effect of 244 different types/concentrations of plant growth regulators on growth and quassin accumulation in callus tissue was investigated. Best growth, in terms of wet/dry weight after four weeks growth, was obtained on B5 media supplemented with 2% glucose, 10% coconut milk, 0.5 mg.l^–1 zeatin riboside and 1.5 mg.l^–1 IBA. The highest yields of quassin (0.014–0.018%) were detected on this same media supplemented with 1.0 mg.l^–1 IBA and varying concentrations of zeatin riboside. Suspension cultures were easily established on B5 media supplemented with 2% glucose, 1.0 mg.l^–1 2,4-D and 0.5 mg.l^–1 kinetin. The carbon source had a marked effect on quassin accumulation with 0.32% quassin being detected when cells were grown in 2% galactose. This is comparable to the highest reported quassin yield for the whole plant.Abbreviations IAA indole-3-acetic acid - IBA indolebutyric acid - IpA N⁶-(-isopentenyl) adenine - IpAR N-(-isopentenyl) adenine riboside - NAA naphthalene acetic acid - 2,4-D 2,4-dichlorophenoxyacetic acid - 6BA 6-benzyladenine     

Effect of cytokinin on morphological changes of suspension cultured cells of the moss, Barbula unguiculata

Suspension cultured cells of the moss, Barbula unguiculata, grow actively in both light and dark culture. Light-grown cells contain chlorophyll and exhibit an undifferentiated callus form. When cells are transferred to a dark condition, they develop into protonemata. Protonemata formation in the dark can be inhibited by the addition of 5 M benzyladenine or 6-furfurylaminopurine but is not affected by the addition of 5 M 2,4-dichlorophenoxyacetic acid or naphthalene acetic acid.Abbreviations BA 6-benzyladenine - 2,4-D 2,4-dichlorophenoxyacetic acid - kinetin 6-furfurylaminopurine - NAA naphthalene acetic acid     

In vitro organogenesis and somatic embryogenesis from leaf explants of Leucosceptrum canum sm.

Plantlets were obtained from leaf explants of a Labiatae tree — Leucosceptrum canum Sm. using plant tissue culture techniques. Two types of calli proliferated from the leaf explants when grown on different media, one of which was amenable to somatic embryogenesis. Differentiation of the embryoids started from the fourth passage of culture and continued up to the seventh passage. The number of embryoids decreased with the age of the callus. The capacity of such embryoids to form entire plantlets was studied using different nutrient mileux. Embryoids formed plantlets on Murashige and Skoog's (MS) medium fortified with benzylaminopurine plus indolebutyric acid. Organogenesis was observed in shoot-buds derived from explants of in vitro regenerated plantlets on MS basal medium supplemented with benzylaminopurine. Culture regenerated plantlets were transferred to MS medium without sucrose and growth hormones; finally transferred to pots containing sterile vermiculite where they are growing.Abbreviations MS Murashige and Skoog's medium - 2,4-D 2,4-dichlorophenoxy acetic acid - NAA naphthaleneacetic acid - IAA indoleacetic acid - IBA indolebutyric acid - Kn kinetin - BAP benzylaminopurine - CW coconut water     

Callus induction and shoot regeneration in Sempervivum tectorum

Leaf and shoot explants of Sempervivum tectorum L., taken from 14- and 30-day-old plants germinated in vitro, have been studied by using Murashige-Skoog and White basal media with cytokinins (benzyladenine, kinetin) and auxins (indoleacetic acid, naphthaleneacetic acid, indolebutyric acid) in various concentrations. Explants taken from 14-day-old plants died but 30-day-old leaves and shoots produced yellow and soft, as well as green and hard calluses on Murashige-Skoog medium with 4.4–8.8 M benzyladenine and 0.57 M indoleacetic acid. Shoot organogenesis was induced from green, hard callus in a medium with 2.2 M benzyladenine plus either 1.1 M indoleacetic acid or 2.5 M indolebutyric acid. Whole plants were grown on Murashige-Skoog medium without plant growth regulators. On the other hand, White medium was not suitable for raising Sempervivum tectorum in vitro.Abbreviations BA benzyladenine - IAA indoleacetic acid - IBA indolebutyric acid - MS Murashige-Skoog - NAA napthaleneacetic acid - W White     

Growth and differentiation of callus cultures of Pinus

Segments of hypocotyl and cotyledons of aseptically-grown seedlings of Pinus strobus L. (white pine) and P. echinata Mill (shortleaf pine) were used as explants for establishing tissue cultures. Growth and differentiation of callus were studied on a modified Murashige and Skoog's medium containing nutrients and plant growth regulators. Meristems below the surface of callus tissue of P. strobus could be induced on media supplemented with -naphthaleneacetic acid alone or in combination with certain other plant growth regulators. Occasionally, differentiation of shoot buds also occurred on callus cultures. These shoot buds could be grown in vitro but roots did not develop.Abbreviations ABA abscisic acid - BA 6-Benzyl-aminopurine - 2-ip N⁶-(²-isopentanyl)-adenine - GD Gresshoff and Doy's medium - GE Gamborg and Eveleigh's medium - MS Modified Murashige and Skoog's medium - NAA -naphthaleneacetic acid - SC Sommer and Caldas' medium - TIBA 2,3,5-Triiodobenzoic acid     

Callus formation from leaf mesophyll protoplasts of Malus Xdomestica Borkh. cv. Greensleeves

Large yields (1.85 × 10⁷/g.f.wt.) of viable protoplasts were obtained from leaves of axenic shoot cultures of Malus Xdomestica Borkh. cv. Greensleeves. Protoplasts cultured in liquid or agarose semi-solidified KM8P medium underwent cell wall regeneration and colony formation.Protoplast-derived cell colonies developed to callus on semi-solid KM8 medium. This is the first report of callus formation from mesophyll protoplasts of apple.Abbreviations BAP 6-benzylaminopurine - K kinetin - Z zeatin - GA₃ gibberellic acid - IBA 3-indole butyric acid - NAA 1-naphthalene acetic acid - IAA 3-indole acetic acid - ABA abscisic acid - f.wt. fresh weight - MS Murashige and Skoog (1962)     

Somatic embryogenesis and shoot organogenesis from interspecific hybrid embryos of Vigna glabrescens and V. radiata

Somatic embryogenesis occurred on cotyledons of morphologically abnormal embryos derived from Vigna glabrescens x V. radiata crosses and cultured on Murashige and Skoog (MS) medium without growth regulators. The frequency of 15–17 day old embryos that gave rise to somatic embryos increased from 8% to 29% by application a mixture of 100 mg/l gibberelllc acid, 25 mg/l -naphthaleneacetic acid (NAA) and 5 mg/l kinetin daily to the pedicels of the developing pods. However, only callus formed on immature hybrid embryos of the reciprocal cross. These callus tissues occasionally gave rise to shoots via organogenesis when transferred to MS medium with 2 mg/l N⁶-benzyladenine and 0.05 mg/l NAA. Treatment of pods with growth regulators did not influence the frequency of organogenic callus. Selfed embryos of the parents did not form somatic embryos in culture, nor did callus derived from the selfed embryos produce shoots. Thus, the ability to redifferentiate appears to be associated with interspecific hybridity.     

Nodule culture and regeneration of Eucalyptus grandis hybrids

Callus of three superior Eucalyptus grandis hybrids was induced from immature inflorescences, floral parts, shoot tips, zygotic embryos, and hypocotyl explants on various auxin (2,4-D or NAA) and cytokinin (kinetin) supplemented media. Hypocotyl callus initiated on 4 mg/l NAA and 1 mg/l kinetin formed massive nodular structures, and shoots and roots after four weeks on hormone free-medium. Callus from all other expiants turned brown and died upon transfer to hormone free or reduced hormone media. The nodular structures originating from hypocotyl-callus were maintained by subculture for over three years and retained the ability to form thousands of shoots. Shoots were successfully rooted (98% rooting) and plantlets developed were transferred to mist-greenhouse and then to greenhouse conditions with 95% survival. Plantlets were grown for six months in the greenhouse without sign of abnormal growth.Abbreviations NAA naphthalene acetic acid - 2,4-D 2,4-dichlorophenoxyacetic acid - IAA indoleacetic acid - MS Murashige and Skoog Medium (1962) - IBA indolebutyric acid     

Regeneration of plantlets from cell suspension culture derived callus of white poplar (Populus alba L.)

Friable calli derived from the stem tissues of Populus alba were used to establish cell suspension cultures which were characterized for in vitro growth and regeneration capacity. Suspended cells and callus recovered from these cells were maximal on a fresh weight basis using MS liquid medium containing 0.44 M BAP and 4.52 M 2,4-D. Shoot regeneration from the recovered callus was observed within 30 to 40 days of culture. The number of shoots was increased by subculturing the shoot-forming callus 2 to 3 times on MS medium supplemented with 19.7 M 2iP and 0.05 M IBA. Regenerated shoots were easily rooted on half-strength MS medium lacking growth regulators, and the plantlets were transferred to pots containing vermiculite for greenhouse growth.Abbreviations BAP 6-benzylaminopurine - 2iP 2-isopentenyladenine - NAA 1-naphthaleneacetic acid - IAA indole-3-acetic acid - IBA indole-3-butyric acid - 2,4-D 2,4-dichlorophenoxyacetic acid - PCV packed cell volume - MS medium Murashige and Skoog medium (1962)     

Rapid micropropagation for Mucuna pruriens f. pruriens L.

A rapid micropropagation system was developed for Mucuna pruriens f. pruriens using explants from 1-week-old aseptically grown seedlings. Multiple shoot regeneration occurred following an initial callus growth on Revised Tobacco (RT) medium supplemented with 2.7 M NAA and 9.8 M 2iP. Maximum number of shoot regeneration was achieved only from seedling explant 6 to 7 days old. More than 90% of the regenerated shoots could be rooted on half-strength liquid RT medium supplemented with 2.7 M NAA. Plantlets readily adopted to greenhouse conditions. This system provides a new tool for micropropagation of Mucuna pruriens f. pruriens, an important medicinal plant.Abbreviations BAP 6-benzylaminopurine - Kn kinetin - 2iP 2-isopentenyladenine - AdS adenine sulphate - NAA alpha-naphthalene acetic acid - IAA indole-3-acetic acid - IBA indole-3-butyric acid - MS Murashige and Skoog (1962) medium - RT Revised Tobacco medium (Kaul and Staba, 1968)     

In vitro regeneration through organogenesis of Meconopsis simplicifolia — An endangered ornamental species

Meconopsis simplicifolia (D.Don) Walp. could be propagated by induction of adventitious shoots from callus produced on hypocotyl, cotyledon and rosette leaf explants of 4-month-old seedlings. Callus was initiated on agar-solidified Murashige and Skoog medium supplemented with 1 M kinetin +10 M -naphthaleneacetic acid (NAA). Shoots formed when the callus was subcultured on medium supplemented with kinetin or benzyladenine (BA) in combination with NAA, indoleacetic acid, indolebutyric acid or gibberellic acid. Excised shoots were rooted on medium containing auxin with 10 M NAA producing the best rooting (55%).Abbreviations BA 6-benzyladenine - 2,4-d 2,4-dichlorophenoxy-acetic acid - FAA formalin-acetic acid-alcohol - GA₃ gibberellic acid - IAA indole-3-acetic acid - IBA -indolebutyric acid - NAA -naphthaleneacetic acid     

Studies on endosperm culture of Annona squamosa Linn

Mature endosperm tissue excised from germinated seeds (2–4 days after radicle emergence) of Annona squamosa grew and proliferated on White's basal medium supplemented with two cytokinins, an auxin and gibberellic acid. The callus obtained could be periodically subcultured. Shoot differentiation and root induction were obtained from callus on media of different compositions. Analyses of the root and young leaf tips showed triploid number of chromosomes (3n=21).Abbreviations Kin Kinetin - BAP 6-Benzylaminopurine - IAA Indole-3-acetic acid - IBA Indole-3-butyric acid - NAA naphthalene abetic acid - GA₃ Gibberellic acid Communication No. 3885     

Somatic embryogenesis and plant regeneration in Carica papaya L. tissue culture derived from root explants

The regeneration potential of shoot tip, stem, leaf, cotyledon and root explants of two papaya cultivars (Carica papaya cv. Solo and cv. Sunrise) were studed. Callus induction of these two cultivars of papaya showed that the shoot tips and stems are most suitable for forming callus, while leaves, cotyledons and roots are comparatively difficult to induce callus. Callus induction also varied with the varities. Somatic embryogenesis was obtained from 3-month-old root cultures. A medium containing half strength of MS inorganic salts, 160 mg/l adenine sulfate, 1.0 mg/1 NAA, 0.5 mg/1 kinetin and 1.0 mg/1 GA₃ was optimal for embryogenesis. The callus maintained high regenerative capacity after two years of culture on this medium. Plants derived from somatic embryos were obtained under green-house conditions.Abbreviations MS Murashige and Skoog's (1962) medium - NAA Naphthaleneacetic acid - IAA Indole-3-acetic acid - GA Gibberellic acid - PRV Papaya ring spot virus     

Plant regeneration from hypocotyl and petiole callus of Trifolium pratense L.

Red clover (Trifolium pratense L.) seedlings were screened for the ability to regenerate plantlets from hypocotyl-derived callus tissue. Media sequences described by Beach and Smith (1979) and Collins and Phillips (1982) and a variation using media from both sequences were tested. Plantlets were regenerated from three out of 642 genotypes. In all three cases, callus was initiated on B5C medium and regeneration was accomplished on SPL medium. Attempts to regenerate plants from petiole-derived callus tissue have so far been successful only with regenerants of clone F49. Petiole callus from epicotyl-derived F49 plants proved to be non-regenerative. Pollen viability varied significantly among individuals regenerated from callus cultures of clone F49. Root tip squashes from F49 regenerants revealed the normal diploid chromosome number (2n=14). The frequency of regeneration within progeny from reciprocal crosses between F49 regenerants and several non-regenerative genotypes was 29%.Abbreviations 2,4-D 2,4-dichlorophenoxyacetic acid - BAP benzylaminopurine - KN kinetin - NAA -naphthaleneacetic acid