Plant regeneration from shoot callus of rosewood (Dalbergia latifolia Roxb.)

In vitro propagation of trees using cell, tissue and organ culture is a fast emerging area. We report here the clonal propagation of Indian rosewood (Dalbergia latifolia Roxb.) from shoot callus cultures of 5 year old trees. Bud regeneration was obtained on MS media supplemented with BA and NAA. About 35% of the cultures showed organogenesis. Shoots measuring about 3–5 cm can be excised and rooted in White's medium supplemented with 1–2 mg/L IAA. Rooted plants were successfully established in soil.Abbreviations BA Benzyladenine - CM Coconut milk - 2,4-D 2,4 dichlorophenoxyacetic acid - IAA Indoleacetic acid - IBA Indolebutyric acid - K Kinetin - NAA Naphthaleneacetic acid - PVP-360 Polyvinyl pyrrolidone     

Direct somatic embroygenesis from immature embryos of rosewood (Dalbergia latifolia Roxb.)

Direct regeneration of somatic embryos was obtained from immature zygotic embryos of Dalbergia latifolia. Immature embryos dissected from green pods 90 d after flowering gave the highest frequency of somatic embryo formation. Preculture on high 2,4-D medium for 4 weeks induced direct somatic embryogenesis, which was expressed during the second culture phase in the presence of low 2,4-D along with a high sucrose concentration. Embryos were separated and transferred to the maturation medium containing MS + 0.5–1.0 mg/L BAP, where embryos developed into plantlets. Somatic embryos failed to convert into complete plants without BAP treatment. This method of direct regeneration of somatic embryos without a callus phase has direct application for genetic manipulation studies.Abbreviations MS Murashige and Skoog (1962) medium - 2,4-D 2,4-Dichlorophenoxyacetic acid - BAP 6-benzylaminopurine - ABA Abscisic acid - KIN Kinetin     

Efficient plant regeneration from cell suspension-derived callus of East Indian rosewood (Dalbergia latifolia Roxb.)

A procedure is outlined for the establishment of a proliferating cell suspension culture of East Indian rosewood (Dalbergia latifolia Roxb.) and efficient plant regeneration from callus derived from such cultures. Callus was induced from hypocotyl segments derived from 1-week-old axenic seedlings on Murashige and Skoog (1962) medium (MS) containing 10.8 μM naphthaleneacetic acid (NAA) and 2.2 μM benzyladenine (BA). Calli were increased by subculturing on MS supplemented with same growth regulators and 10% coconut water (CW). Friable calli were used to initiate cell suspension cultures. Optimum cell proliferation occurred in MS containing 10.8 μM NAA, 2.2 μM BA and 10% CW, using an initial inoculum cell density of 2%. Cell clumps composed of 20–25 cells harvested from suspension cultures at the exponential growth phase readily formed callus within 3 weeks following plating on the semi-solid MS as above. High-frequency shoot-bud differentiation was induced in these calli on MS containing 2.7 μM NAA and 13.3 μM BA. The regeneration frequency declined at higher BA concentrations. The organogenic potential of the cell suspensions was influenced by the age of the culture. Regenerated shoots were rooted on half-strength MS containing 5.7 μM indole-3-acetic acid, 4.9 μM indole-3-butyric acid and 5.3 μM indole-3-propionic acid. The plantlets were acclimatized and established in soil. Received: 22 August 1997 / Revision received: 21 May 1998 / Accepted: 1 June 1998     

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     

Rapid In Vitro Propagation of East Indian Rosewood (Dalbergia latifolia Roxb.) through High Frequency Shoot Proliferation from Cotyledonary Nodes

An efficient protocol is described for large scale in vitro propagation of east Indian rosewood (Dalbergia latifolia Roxb.) using cotyledonary nodes derived from axenic seedlings. Of the four different cytokinins tested, BA was most effective in inducing multiple shoot buds in the explants. High frequency shoot proliferation (93%) coupled with maximum number of shoot formation (10-12 shoots/explant) was recorded on Murashige and Skoog’s medium supplemented with 2.0 mg/l BA. The frequency of shoot proliferation declined at higher levels of cytokinins. Shoot culture was multiplied by subculturing the original cotyledonary node on a fresh shoot multiplication medium each time aHer excising the newly formed shoots. Shoots obtained from each passage were multiplied further as nodal explants and each node produced 3-4 shoots. In two months from a single cotyledonary node, about 90 shoots were obtained. Rooting was induced in 72% of the regenerated shoots on half-strength MS containing IAA, IBA and IPA each at 1.0 mg/l. Rooted plantlets were hardened off and eventually established in soil.     

In vitro clonal multiplication of mature trees of Dalbergia sissoo Roxb.

Multiple shoots were obtained from nodal explants of 30-year-old trees of Dalbergia sissoo Roxb. on a defined medium, MS (Murashige & Skoog medium) supplemented with auxin-cytokinin combinations. IAA (Indole accetic acid) alone promoted 15% rooted shoot buds. A combination of IAA+Kn (Kinetin) gave 100% rooted shoot buds. A combination of NAA (napthaleneacetic acid) + Kn and NAA+BAP (6-benzylaminopurine) also gave high percentages of rooted shoot buds. Ascorbic acid in the medium prevented the death of callus and plantlets, which followed darkening of the medium.     

In vitro micropropagation of Baliospermum montanum (Willd.) Muell-Arg--a medicinal plant

Micropropagation of B. montanum was achieved on Murashige and Skoog's (MS) medium augmented with BAP using nodal segments. Maximum number of shoots (3.4 +/- 0.25) were found in MS medium fortified with BAP (3.10 microM). In vitro raised shoots were rooted on half strength MS medium augmented with various concentrations and combination of auxins viz.. IAA, IBA and NAA. Maximum number of roots were observed on half strength MS medium fortified with IBA (9.84 microM) combined with NAA (5.37 microM).     

Rapid In Vitro Multiplication of Dalbergia sissoo Roxb

Micropropagation of Dalbergia sissoo Roxb was achieved through in vitro proliferation of axillary buds from 30 to 40 years old mature tree. Bud-break was achieved within six days when nodal explants were cultured on MS medium supplemented with kinetin (9.2 μm), indole-3-butyric acid (2.46 μM) apd 6-benzyladenine (13.2 μM). Multiple shoot formation occurred from nodal explants of in vitro raised shoots on MS medium with reduced levels of major salts and kinetin. Roots were Induced within 5 days on in vitro generated shoots on MS medium supplemented with 1-naphthalene acetic acid (0.53 μM) and indole-3-butyric acid (9.8 μM).     

Evaluation of biochemical changes during different stages of somatic embryogenesis in a vulnerable timber tree Dalbergia latifolia (Indian rosewood)

In Vitro Cellular & Developmental Biology - Plant - In Dalbergia latifolia Roxb. (Indian rosewood), a premium-quality vulnerable timber tree species belonging to Family Fabaceae, direct somatic...     

Characterization of microsatellite markers in the rosewood (Dalbergia monticola Bosser & R. Rabev.)

Dalbergia monticola is one of the major components of the oriental forest of Madagascar. This economically and ecologically important tree is threatened because of the dramatic decrease of the Madagascar forest. We have estimated the genetic diversity and structure of the species by studying nuclear microsatellites. We have developed eight pairs of primers to analyse 215 individuals distributed from the north to the south of the island. These markers will be useful for genetic and ecological studies of this species.     

Growth and Leaf Gas Exchange Characteristics in Dalbergia sissoo Roxb. and D. latifolia Roxb. Under Water Deficit

Forty two-month-old plants of Dalbergia sissoo and D. latifolia were subjected for 56 d to water deficit induced by withholding water. Drought stress caused a significant reduction in plant height, stem diameter, net photosynthetic rate (P _N), transpiration rate (E), and stomatal conductance (g _s) in both species, but the reduction was greater in D. sissoo than in D. latifolia. Water use efficiency (P _N/E) was adversely affected due to water stress only in D. latifolia, and intrinsic water use efficiency (P _N/g _s) was increased in both species. There was a slight effect of water stress on variable to maximum fluorescence (F_v/F_m) (quantum yield of photosystem 2) in both species, but the species did not differ significantly in this attribute.     

In vitro micropropagation of the macarronesian evergreen treePersea indica (L.) K. Spreng

Summary Shoot propagation ofPersea indica (L.) K. Spreng was achieved using seedling axillary buds cultured on MS (Murashige and Skoog, 1962) medium with 1 mg/l (2.8 μM) N⁶-benzyladenine (BA). Forty percent of the obtained shoots did not elongate, but showed bud proliferation, which was maximal (three axillary buds per shoot) at the end of the seventh subculture. Sixty percent of the shoots elongated, did not show bud proliferation, and formed calluses at their base. Successful rooting (84.6%) was achieved dipping the base of each elongated shoot in 3 g/l (16.11 mM) indolebutyric acid (IBA) for 1–2 s, and transferring to half strength MS medium without growth regulators. These shoots presented an acclimatization success of 100%. Results suggest that micropropagated elongated shoots ofP. indica can be adequately used in reforestation programs.     

Gibberellic acid and thidiazuron promote micropropagation of an endangered woody tree (Pterocarpus marsupium Roxb.) using in vitro seedlings

Plant Cell, Tissue and Organ Culture (PCTOC) - Pterocarpus marsupium Roxb., which belongs to the Fabaceae family, is a promising herbal drug producing forest tree widely known as the ‘Indian...     

In vitro propagation of the bamboo (Bambusa tulda Roxb.) through shoot proliferation

An efficient protocol has been developed for the in vitro propagation of Bambusa tulda through shoot proliferation. Shoots from 3-week-old aseptically grown seedlings were used to initiate cultures. Multiple shoots were obtained on liquid Murashige and Skoog (MS) medium supplemented with 6-benzylaminopurine (8×10^–6M) and kinetin (4×10^–6M). Continuous shoot proliferation at a rate of 4–5 fold every three weeks was achieved through forced axillary branching. More than 90% of the shoots could be rooted on a modified MS medium containing indoleacetic acid (1×10^–5M) and coumarin (6.8×10^–5M). Following simple hardening procedures, the in vitro raised plants were transferred to the soil with more than 80% success.Abbreviations BAP 6-benzylaminopurine - 2-ip 6-,-dimethylallylaminopurine - Kn kinetin - IAA 3-indoleacetic acid - IBA 3-indolebutyric acid - NAA 1-naphthaleneacetic acid     

In vitro micropropagation of Lawsonia inermis (Lythraceae)

A successful protocol was developed for mass propagation of Lawsonia inermis Linn., an important medicinal plant. Multiple shoots were induced in apical and axillary meristems derived from mature explants of L. inermis on Murashige and Skoog (1962) medium supplemented with 0.25 mg/l 6-benzylaminopurine (BA), 0.25 mg/l Kinetin (Kn), 0.5 mg/l ascorbic acid and 3% (w/v) sucrose. The rate of multiplication was higher when the cultures were incubated under continuous light rather than the 14 hr photoperiod. Rooting was readily achieved upon transferring the microshoots onto MS basal semi-solid medium supplemented with 0.25 mg/l indole-3-butyric acid (IBA) after ten days of culture. Micropropagated plantlets were acclimatized and successfully grown in soil.     

In vitro micropropagation of Ruscus aculeatus

We have developed three protocols for the rapid micropropagation of Ruscus aculeatus. The primary explants utilised were immature embryos, aerial buds excised from rhizomes and shoot buds regenerated from organogenic calli. In order to increase the plant regeneration from the primary explants, we used organogenic calli from cladode, stem and rhizome segments. We tested more than 20 culture media for callus induction and shoot regeneration and the best results were obtained when rhizome segments were cultured on Murashige and Skoog medium supplemented with 0.5 mg dm⁻³ 2,4-dichlorophenoxyacetic acid and 1 mg dm⁻³ kinetin.     

In vitro micropropagation of Arctostaphylos uva-ursi (L.) Sprengel.: Comparison between two methodologies

In vitro micropropagation of Arctostaphylos uva-ursi was performed to increase the number of ground cover species able to serve as substitute for members of the Rosaceae susceptible to fire blight. Explants (node segments) excised from plants growing in the greenhouse were established in vitro on a medium containing 10 M -naphthaleneacetic acid (NAA) and activated charcoal (2 g I^-1). Using in vitro grown shoots, two propagation procedures were used:- Culture of nodal fragments with 50 M NAA resulted in the growth of 6 to 7 nodes every 4 weeks, yielding 1 700 almost rootable shoots after 4 subcultures;- Development of axillary shoots obtained with media containing 25 M benzyladenine (BA) and 20 M indoleacetic acid (IAA) yielded almost 500 rootable shoots after 4 subcultures. The rate of propagation decreased after the 3^rd subculture.Percentage of in vitro rooted shoots reached 98% with diluted micronutrients and 10 M NAA but 31% of the plants died during acclimatization.Abbreviations BA benzyladenine - BM basal medium - HID high intensity discharge - IAA indoleacetic acid - IBA indolebutyric acid - NAA -naphthaleneacetic acid - PAR photosynthetic active radiation - 2iP 2-isopentenyladenine     

紫穗槐的离体快速繁殖

以子叶节为外植体,建立起了紫穗槐的快速离体再生系统.经过四周的培养,在附加8mg·L-16-BA的MS培养基上能够获得再生频率为100%,平均每个外植体5.21个芽点的高效再生植株.以再生植株的茎节为外植体所进行的继代能够在相同的培养基上连续的产生新的不定芽,但芽点数要少于起始培养.经过3周的培养,有82.53%切下的再生茎段能够在含2.0mg·L-1IAA的MS培养基上生根.在所有进行分析过的再生植株中,它们的染色体数目都没有发生变异(2n=40).经过练苗以后,再生植株成功地定植于土壤当中并展示了一致的外部形态和生长特性.