Rapid propagation of tuna (Opuntia ficus-indica) and plant establishment in soil

Explants from young joints of mature plants of tuna (Opuntia ficus-indica Mill.) were cultured on Murashige and Skoog (MS) medium containing 8.8 M benzyladenine (BA) and 0.5 M naphthaleneacetic acid (NAA). Shoots produced were utilized as secondary explants. Each shoot was cut longitudinally from apex to base into two explants, and some of these explants were cut transversely into proximal and distal explants. The size and number of shoots produced was affected by size and position of the explant within its source. The shoots were rooted in vitro or ex vitro and plants were successfully established in soil from both rooting methods.Abbreviations AC activated charcoal - BA benzyladenine - IBA indolebutyric acid - MS Murashige & Skoog (1962) medium - NAA naphthaleneacetic acid     

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     

In vitro propagation of the leguminous tree Swartzia madagascariensis

Shoot induction frequency for the leguminous tree Swartzia madagascariensis Desv. was higher on MS and WP media than on B5. Explants incubated on media solidified with agar produced more shoots with a lower tendency to hyperhydricity than explants on agarose or Gelrite media. Maximum shoot induction was obtained with an agar-solidified MS medium containing 2.2 M benzyladenine (37 shoots/explant). Shoots rooted after transfer to half-strength MS medium supplemented with 26.8 M naphthaleneacetic acid.Abbreviations BA benzyladenine - IBA indolebutyric acid - NAA naphthaleneacetic acid - WP(M) woody plant (medium)     

In vitro shoot proliferation and production of sets from garlic and shallot

A procedure is described to regenerate shoots and bulbs in vitro with high frequency from shoot tips of garlic and shallot plants using benzyladenine or thidiazuron. Regenerated shoots were induced to form bulbs in Murashige and Skoog medium (1962) containing 5 g l^-1 activated charcoal and 120 g l^-1 sucrose under a long-day photoperiod. Bulbs formed in vitro were transferred to soil without acclimatization and produced viable plants. This method could be useful to produce low-cost bulbs, which are easy to handle and store until needed.Abbreviations AC activated charcoal - BA benzyladenine - IBA indolebutyric acid - MS Murashige & Skoog's (1962) medium - NAA naphthaleneacetic acid - TDZ thidiazuron     

In vitro induction of multiple shoots and plant regeneration from shoot tips of mung bean (Vigna radiata (L.) Wilczek)

Conditions for plant regeneration from excised shoot tips of Vigna radiata were studied. Complete plants were regenerated directly without an intervening callus phase from shoot tips on basal medium (MS salts+B₅vitamins). Regeneration frequency varied with genotype, explant size and growth regulator combinations in the medium. Addition of cytokinins induced a variable amount of callus at the base of the shoot tip, followed by multiple shoot formation. Benzyladenine (BA), kinetin and zeatin at 5×10^-6 M each induced multiple shoots in 100% of the explants but the highest number of regenerants per explant (9) was produced with BA. The efficacy of BA for shoot multiplication was not improved when it was supplemented with naphthaleneacetic acid (NAA) or indoleacetic acid (IAA). NAA or adenine sulphate, when applied alone, induced complete plantlets. The growth regulator requirement of explants for the induction of multiple shoots varied with explant size. The shoot tip explants maintained proliferation ability on subculture. None of the treatments was effective in inducing shoot bud differentiation from callus. Regenerated shoots were rooted on MS basal medium and MS supplemented with either IAA or indolebutyric acid. The rooted plants were transferred to the field; 60% subsequently survived and grew.Abbreviations BM basal medium [MS (Murashige & Skoog 1962) salts+B₅ (Gamborg et al. 1968) vitamins] - BA 6-benzyladenine - AdS adenine sulphate - IAA indole-3-acetic acid - NAA-1 naphthaleneacetic acid - IBA indolebutyric acid     

Plant regeneration of Mangifera indica using liquid shaker culture to reduce phenolic exudation

The exudation of phenolics from the cut ends of mango explants greatly hinders their regenerative ability in any in vitro growth medium. However, pretreatment of explants using liquid shaker culture helps in overcoming this problem. Explants kept in liquid MS medium supplemented with 1% polyvinylpyrolidone in 250 ml conical flasks on an automated shaker at 75 rpm were able to produce shoots when inoculated on gelled MS medium supplemented with different concentrations of growth regulators.Abbreviations BA benzyladenine - IAA indoleacetic acid - NAA naphthaleneacetic acid - IBA indolebutyric acid     

Micropropagation procedures for Leontochir ovallei

Leontochir ovallei Phil., an endangered Chilean species in the Alstroemericeae, was micropropagated on Murashige & Skoog medium supplemented with 4 M benzyladenine, 1 M indolebutyric acid and 146 mg l^-1 glutamine. Over 88% of the shoots rooted in vitro when treated with 10 M naphthaleneacetic acid and micropropagated plantlets were successfully transplanted into the greenhouse.Abbreviations BA benzyladenine - IBA indolebutyric acid - 2iP isopentenyladenine - NAA naphthaleneacetic acid - MS Murashige and Skoog (1962) medium     

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     

Production of clonal plantlets of Grevillea robusta in in vitro culture via axillary bud activation

A micropropagation method for Grevillea robusta A. Cunn. was developed using explants from mature trees cultured on Woody Plant Medium plus 4.4 M benzyladenine and 0.27 M naphthaleneactic acid (NAA) for shoot proliferation. Each nodal explant produced three to five shoots within 10 to 12 weeks in three successive transfers. Rooting was obtained by dipping the shoots in 0.54 mM NAA solution. Rooted shoots were established well in soil.Abbreviations BA benzyladenine - GA₃ gibberellic acid - IBA indolebutyric acid - NAA naphthaleneactic acid     

Adventitious in vitro plantlet formation from immature floral stems of Crinum macowanii

Adventitious shoots and plantlets were regenerated in vitro from floral stem explants of Crinum macowanii (Bak.) (bush lily). The length (age) of the floral stem as well as the orientation and position of the explant disc in the floral stem were the most important factors affecting shoot regeneration. The highest number of shoots were regenerated when immature floral stems of 70–100 mm were used as starting material, using the middle or basal parts of the stem, and orientating the discs with their proximal ends on the medium. Combinations of kinetin (4.65 M) and either indoleacetic acid (0.57 M) or naphthaleneacetic acid (0.54 M), or a combination of benzyladenine (4.44 M) and 2,4-dichlorophenoxyacetic acid (0.45 M) resulted in the highest numbers of shoots being regenerated. Although a slight degree of callus formation was noticed on the cut-edges of the discs, shoot formation did not occur via callus, but directly from the floral stem epidermis. Unrooted shoots were rooted on MS-medium containing 0.17 M sucrose.Abbreviations BA benzyladenine - IAA indoleacetic acid - IBA indolebutyric acid - NAA naphthaleneacetic acid - 2,4-d 2,4 dichlorophenoxyacetic acid     

Preliminary studies of micropropagation of Hopea odorata,a dipterocarp tree

Plantlet development from in vitro cultures of Hopea odorato Roxb. is described. Embryos excised from seeds and cultured on Gamborg's B5 or modified Murashige and Skoog (MS) medium with benzyladenine (BA, 2.2–22.2 M) produced axillary shoots at cotyledonary and/or stem nodes. Shoot production was greatest in germinated embryos on modified MS medium with 8.9 M BA. Excised axillary shoots formed few buds when cultured on medium with BA and limited root development occurred on Woody Plant Medium with naphthaleneacetic acid. Nodal explants from aseptically grown plantlets sprouted axillary shoots in modified MS medium with BA.Abbreviations BA 6-benzyladenine - 2,4-d 2,4-dichlorophenoxyacetic acid - GA₃ gibberellic acid - IBA indolebutyric acid - NAA naphthaleneacetic acid - PVPP polyvinylpolypyrrolidone     

High-frequency induction of adventitious shoots from hypocotyl segments ofLiquidambar styracifiua L. by thidiazuron

The effects of thidiazuron (TDZ) on adventitious bud and shoot formation from hypocotyl segments of sweetgum (Liquidambar styracifiua) were tested alone and in combination with 2,4-dichlorophenoxyacetic acid (2,4-D). The combination of 1 mg/1 TDZ with 0.01 mg/l 2,4-D resulted in the highest frequency of bud production. Lower concentrations of TDZ stimulated shoot production, generating the most shoots at 0.1 mg/1 TDZ with 0.01 mg/1 of 2,4-D. Inhibition of shoot elongation by TDZ was overcome by transferring shoot cultures to a shoot proliferation medium lacking TDZ or containing naphthaleneacetic acid and benzyladenine in addition to TDZ. Shoot production in liquid culture was significantly greater than that in solid culture. Comparisons of in vitro and ex vitro rooting of the adventitious shoots demonstrated that ex vitro rooting produced plants with faster growth rates and more extensive root systems.Abbreviations BA Benzyladenine - IBA indole-3-butyric acid - NAA naphthaleneacetic acid - PGR plant growth regulator - TDZ thidiazuron - 2,4-D 2,4-dichlorophenoxyacetic acid     

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     

Thidiazuron stimulates shoot organogenesis and somatic embryogenesis in white ash (Fraxinus americana L.)

Immature and mature nonstratified seeds of white ash (Fraxinus americana L.) were dissected transversely and 2/3 of each seed was placed onto agar-solidified Murashige and Skoog medium. Adventitious buds, shoots, and somatic embryos formed on callus, cotyledons, and hypocotyls of the resulting seedlings. Shoot organogenesis was induced on explants cultured on medium with 10 M thidiazuron but not on explants on media with benzyladenine (BA) or isopentenyladenine. Not all seed sources were equally capable of shoot organogenesis and embryogenesis. Atypical of adventitious regeneration of other woody plants, mature seed explants of white ash were more organogenic with shoots that elongated better than explants from immature seeds. Somatic embryogenesis was observed in cultures where mature seeds were first cultured for 4 weeks on a medium containing 10 M adenine 2,4-dichlorophenoxyacetic acid in combination with 0.1 and 1.0 M thidiazuron, followed by transfer to a medium containing 0.05 M 6-benzyladenine and 0.5 M naphthaleneacetic acid. Adventitious shoots and epicotyls from both seedlings and germinated somatic embryos were rooted under intermittent mist and acclimatized to the greenhouse.Abbreviations BA 6-benzyladenine - 2,4-d 2,4-dichlorophenoxyacetic acid - IBA indolebutyric acid - 2iP isopentenyladenine - NAA naphthaleneacetic acid - TDZ thidiazuron-N-phenyl-N-1,2,3-thiadiazol-5-ylurea - WPM woody plant medium     

Organogenesis in vitro of Nothofagus alpina (P. et E.) Oerst, Fagaceae

In vitro shoot and root regeneration of 2-year-old Nothofagus alpina plants was achieved from several types of expiants cultured in vitro on a modified Woody Plant Medium formulation. Multiple shoot formation was obtained from leaf expiants using 0.45–2.27 M thidiazuron and 0.0049–0.098 M indolebutyric acid. Excised axillary buds formed shoots and roots in the presence of 0.0049 M benzyladenine and 2.46 M indolebutyric acid, or in the absence of plant growth regulators. Nodal sections rooted when 2.46 M indolebutyric acid at was supplied in the medium. Subcultured shoots originating from nodal sections showed a high regeneration rate through multiple shoot and root formation.Abbreviations BA benzyladenine - GA₃ giberellic acid - IBA indolebutyric acid - NAA -naphthaleneacetic acid - TDZ thidiazuron - WPM McCown's Woody Plant Medium - Z zeatin     

In vitro culture of Sophora toromiro (Papilionaceae), an endangered species

Sophora toromiro (Phil.) Skottsb. is a woody species in imminent danger of extinetion. Since natural regeneration by seeds is poor and plant growth is very slow, asexual propagation is necessary. In vitro regeneration from 3- to 4-month-old seedlings was achieved. A range of naphthaleneacetic acid and benzyladenine concentrations induced root formation in nodal segment explants, developing plantlets, and also promoted axillary bud development. In subculture, nodal sections derived from axillary growth initiated multiple shoot formation and roots in a liquid medium leading to plantlet formation.Abbreviations NAA naphthaleneacetic acid - BA benzyladenine - IBA indolebutyric acid - GA₃ gibberellic acid     

In vitro shoot proliferation and propagation of guava (Psidium guajava L.) from germinated seedlings

Guava seeds were germinated on Murashige and Skoog (MS) medium with or without 8.8 μM benzyladenine (BA). BA increased the rate of germination and the number of lateral shoots (3.4 vs 1.2 per seedling). Stem nodes from these lateral shoots were cultured on proliferation media with 4.4 μM BA, and multiple shoots (3.5) were formed within 4 weeks of culture. Increasing the concentration of BA or the addition of naphthaleneacetic acid (NAA) did not affect shoot formation. Shoots produced from explants and lateral shoots from germinated seedlings were rooted in media containing activated charcoal (AC) or 9.8 μM indolebutyric acid (IBA). Shoots rooted with IBA had a higher rooting percentage (100% vs 75%) and a greater number of roots (5.5 vs 3.2) but the shoots were shorter (2.6 vs 3.4 cm) than when rooted in AC, and they required an additional 4 weeks of culture in media with AC to achieve shoot elongation. About 80% of the shoots with roots survived in the glasshouse and produced normal phenotypic plants.     

Micropropagation of Spilanthes acmella Murr.

Multiple shoots of Spilanthes acmella Murr. were induced from hypocotyl segments obtained from 1-week-old seedlings on Murashige and Skoog's (MS) medium containing benzyladenine (BA), isopentenyl adenine, and naphthaleneacetic acid (NAA). High frequency shoot proliferation (95 %) and maximum number of shoots per explant (10 ± 0.6) were recorded with 0.5 mg dm^–3 BA in combination with 0.1 mg dm^–3 NAA. A proliferation was achieved by repeatedly subculturing the nodal segments on shoot multiplication medium. About 95 % of the in vitro shoots developed roots after transfer to half strength MS medium containing indole-3-butyric acid (1.0 mg dm^–3). 95 % of the plantlets were successfully acclimatized and established in soil. Transplanted plantlets showed normal flowering without any morphological variation.     

Enhancement of shoot regeneration from cotyledon explants of Brassica rapa ssp oleifera through pretreatment with auxin and cytokinin and use of ethylene inhibitors

The presence of benzyladenine or naphthaleneacetic acid in seed germination medium markedly enhanced subsequent shoot regeneration from the base of the excised cotyledon explants of Brassica rapa cv. Horizon. Cotyledon explants from younger seedlings (3 or 4-day old) produced more shoots than those from older seedlings. Addition of the ethylene inhibitor aminoethoxyvinylglycine (1.0 M) to the regeneration medium improved shoot regeneration three fold.Abbreviations AVG aminoethoxyvinylglycine - BA benzyladenine - MGBG methylglyoxal-bisguanylhydrazone - MSBN ms (murashige & skoog 1962) medium supplemented with 4.4 m BA & 5.4 m NAA, 2% sucrose - NAA naphthaleneacetic acid     

Micropropagation of Actinidia kolomikta

Nodal segments of female A. kolomikta shoots were cultured on Murashige and Skoog modified medium with different growth regulator concentrations. The highest multiplication rate (9.5) was achieved on a medium with 10 M benzyladenine and 0.1 M indolebutyric acid. Over 90% of shoots rooted in vivo after pulse stimulation with indolebutyric acid.Abbreviations BA benzyladenine - IBA indole-3-butyric acid - NAA 1-naphthaleneacetic acid