Callus formation from protoplasts of Artemisia sphaerocephala Krasch and some factors influencing protoplast division

Round wormwood (Artemisia sphaerocephala Krasch) seeds were germinated on Murashige & Skoog (1962) medium without plant growth regulators. The hypocotyls of seedlings were sliced and cultured on M1 medium with 2,4-dichlorophenoxyacetic acid (9.05 M) to induce callus. The induced calluses were subcultured on the same medium. Ten day old calluses were used to isolate protoplasts in an enzyme solution with 0.65 M mannitol. Protoplast yield strongly depended upon the state of callus cultures. Certain amount of hemicellulase could improve protoplast isolation. Purified protoplasts were cultured in modified Kao & Michayluk (1975) medium with 0.60 M mannitol as osmoticum, suggesting that protoplasts of A. sphaerocephala need a high initial osmolarity. Protoplasts generally divided evenly and the percentage of first division could reach 10%. Kinetin exhibited a positive effect on initial cell division. Furthermore, we studied the effect of protoplast density and vitamin C on sustained growth of protoplasts. After forty days, 1 mm calluses in diameter formed.Abbreviations CH casein hydrolysate - 2,4-D 2,4-dichlorophenoxyacetic acid - KM8P Kao & Michayluk (1975) protoplast medium - MS Murashige & Skoog (1962) medium - MES-2 (N-morpholino)ethanesulfonic acid     

Neogene diversification in the temperate lichen-forming fungal genus Parmelia (Parmeliaceae,Ascomycota)

The lichen-forming genus Parmelia Acharius occurs worldwide but its centre of distribution is in the northern hemisphere and it is widespread in boreal-temperate Eurasia and North America. Recent molecular work on Parmelia has identified phylogenetic relationships within two major groups of the genus: P. saxatilis s. lat. and P. sulcata s. lat. However, little is known about the diversification and historical biogeography of these groups. Here we have used a dataset of two genetic markers and 64 samples to estimate phylogenetic relationships within Parmelia. The dated phylogeny provides evidence for major diversification during the Neogene and Pleistocene. These diversification events are probably correlated with climatic changes during these periods. Evidence of gene flow within species between populations from North America and Europe has been found in three species: P. sulcata Taylor, P. saxatilis (L.) Acharius and P. barrenoae Divakar, M.C. Molina & A. Crespo. Cryptic species recently segregated on the basis of molecular differences (P. encryptata A. Crespo, Divakar & M.C. Molina vs. P. sulcata and P. saxatilis vs. P. mayi Divakar, A. Crespo & M.C. Molina) do not share a common ancestor. Moreover, the P. saxatilis complex is remarkably diverse. Two morphotypes of P. saxatilis s. lat. were shown to represent independent monophyletic lineages. Consequently, two species (P. sulymae Goward, Divakar, & M.C. Molina & A. Crespo and P. imbricaria Goward, Divakar, M.C. Molina & A. Crespo) are newly described here.     

基于分子证据确认秦岭藤属与驼峰藤属(夹竹桃科)的系统位置

秦岭藤属(Biondia Schltr.)是中国特有属,约13种,驼峰藤属(Merrillanthus Chun&Tsiang)是仅在我国与柬埔寨分布的单种属,目前两个属均被归并到白前属(Vincetoxicum Wolf),由于取样和系统发育分析尚存一些欠缺,因此其系统位置和归属尚需进一步确认。该文使用2个核糖体基因序列片段(ITS、ETS)、5个叶绿体基因序列片段(psbA-trnH、trnG、trnL、trnL-F、trnT-L),以及二者的合并数据,重建娃儿藤亚族(Tylophorinae)(包含属的模式秦岭藤[B.chinensis Schltr.=V.shaanxiense(Schltr.) Meve&Liede]与驼峰藤[M.hainanensis Chun&Tsiang=V.hainanense(Chun&Tsiang) Meve, H. H. Kong&Liede])的系统发育树。结果表明：秦岭藤属与驼峰藤属均包含在白前属中,秦岭藤与青龙藤[B.henryi(Warb.) Tsiang&Li=V.henryi(Warb....     

A New Nonparametric Approach to the Comparison of K Independent Samples of Response Curves II: A K Sample Generalization of the FRIEDMAN Test

A K sample generalization of the FRIEDMAN test (1937) is introduced which can be used as a nonparametric procedure for testing the homogeneity of the profiles of K independent samples of response curves measured at T identical points of time. While a similar procedure in LEHMACHER & WALL (1978), section 3, is based on T combined tests, each of them at level a/T, here a finite and asymptotic test is presented which is based on a single test statistic. The application of the new multivariate test is illustrated by the same numerical example as in LEHMACHER & WALL (1978). The properties of this test are discussed and compared with the combined test mentioned above.     

Factors affecting in vitro shoot formation from vegetative shoot apices of apple and relationship between organogenic response and cytokinin localisation

ABSTRACT

The effects of macro- and micro-elements, benzyladenine (BA) concentration, and the period of auxin application on adventitious shoot formation from callus originating from vegetative shoot apices were tested on apple (Malus domestica Borkh) rootstock Jork 9. The putative relationship between organogenic response and cytokinin localisation was also studied by an immunolocalisation technique for in situ determination of free cytokinins. The use of MS (Murashige & Skoog, 1962) salts in the medium instead of those of LP (Quoirin & Lepoivre, 1977) had a strong positive effect both on shoot formation rate and on the number of shoots produced. The highest organogenic response from callus was induced using 17.8 μM BA in the presence of 2.7 μM NAA and by maintaining the explants for 20 days in darkness, then transferring them to fresh auxin-free medium and to the light. The in situ localisation studies, performed using antibodies with a marked specificity against zeatin and isopentenyladenine, revealed changes in the localisation of free zeatin in the tissues during the shoot-forming process, in particular during the active cell division phase leading to callus formation, and in the initial phase of bud formation. Changes in zeatin distribution in the tissues of the vegetative shoot apex during shoot formation may indicate a role for this cytokinin free base in cell differentiation and organogenesis.     

New lichenicolous, muscicolous, corticolous and lignicolous taxa of Burgoa s. l. and Marchandiomyces s. l. (anamorphic Basidiomycota), a new genus for Omphalinafoliacea, and a catalogue and a key to the non-lichenized, bulbilliferous basidiomycetes

A catalogue and a key to the non-lichenized, bulbilliferous basidiomycetes are given. The new genus Burgella is described for the lichenicolous B. flavoparmeliae, phylogenetically close to Sistotrema oblongisporum and Multiclavula. The genera Pneumatospora and Tricellulortus are placed in synonymy of Minimedusa, the new combination M. obcoronata is proposed, and the new facultative lichenicolous M. pubescens is described. The new facultative lichenicolous Burgoa angulosa is phylogenetically close to the generic type B. verzuoliana, whilst the new B. moriformis and B. splendens are provisionally included in the genus Burgoa. A Burgoa-like species in the Ceratobasidiaceae is left unnamed. Two new species of Marchandiomyces, M. buckii and M. nothofagicola, are described. As Marchandiomyces aurantiacus is phylogenetically more close to Erythricium than to Marchandiomyces, it is proposed to exclude it from that genus and to use the holomorphic generic name Marchandiobasidium for both anamorph and teleomorph of this species. The new genus Marchandiomphalina is introduced for the lichenized Omphalina foliacea, a taxon phylogenetically close to Marchandiobasidium. Taxonomic novelties Burgella Diederich & Lawrey; B. flavoparmeliae Diederich & Lawrey; Burgoa angulosa Diederich, Lawrey & Etayo; B. moriformis Diederich, Ertz & Coppins; B. splendens Diederich & Coppins; Marchandiomphalina Diederich, Lawrey & Binder; Marchandiomphalina foliacea (P. M. J?rg.) Diederich, Lawrey & Binder; Marchandiomyces buckii Diederich & Lawrey; M. nothofagicola Diederich & Lawrey; Minimedusa pubescens Diederich, Lawrey & Heylen; M. obcoronata (B. Sutton, Kuthub. & Muid) Diederich & Lawrey     

Cellular succession in the epithelium lining the pharyngeal cavity of planarians

Regardin cell renewal of planarian epithelial tissue, little is known at present except for the epidermis (Tyler, 1984). According to Skaer (1965), the definitive epidermal cells in planarian embryos originate in the parenchyma where they develop as precursor cells which then migrate into the epidermis. This process of development is also reported in regenerating planarians (Spiegelman & Dudley, 1973; Morita & Best, 1974; Hori, 1978). Recently the validity of Skaer's conclusion has clearly been confirmed by Ishii & Sakurai (1998) using SEM and TEM.The present study deals with the origin and renewal of cells of another epithelium — that lining the pharyngeal cavity of a freshwater planarian Dugesia japonica. This epithelium is known to consist of cells with secretory granduls of fingerprint-like structure (Ishii, 1966; Bowen & Ryder, 1973; Gamo & Garcia-Corrales, 1988; Asai, 1991). The characteristic feature of these specific granules, as well as epitheliosomes (Tyler, 1984) of the epidermis, is very useful as a cell marker for distinguishing cell types and following cell differentiation. Gamo & Garcia-Corrales (1988) claimed the cell populations of the pharyngeal epithelium of Dugesia gonocephala is morphologically and functionally heterogeneous. The sequence of morphological alteration offers the oppurtunity to distinguish at least three functional stages. These represent different adaptive forms of a single cell type, and indicate transformation from one to the next induced by cell renewal. It suggests a beta cell (noeblast) origin of the epithelial cells.In the present study SEM observations revealed theat there are morphologically three distinct cell types which are considered to be juvenile, mature, and senile cells respectively, judging from the differences in cells and the many transitional forms among them (Fig. 1). When observed by TEM, the cytoplasmic patterns of these cell types are correspondingly different from one another, similarly the surface specializations of the plasma membranes of each cell type which facilitate their identification by TEM also vary. Apparently alterations of cell morphology are involved in ageing in a single cell type. Morphological features of each cell type are summarized. Juveniles (R) rarely occur, are small cells provided with prominent apical ruffles and considered to be phagocytic because the cells often contain newly formed phagosomes. Mature cells (F) predominate, are large, polygonal, flat cells, presumably with an intense secretory activity. The cells contain many profiles of granular ER and a small number of specific (secretory) granules. Senile cells (M) are intermittently distributed, irregularly contoured cells with heavy surface microvilli. The specific granules are dense in the apical cytoplasm, and profiles of granular ER decease in number.The precursor epithial cells that are seen in the parenchyma exhibit a considerable degree of differentiation before their translocation into the lining. The primordial cells observed are neoblast-like, with several cohromatoid bodies and many free ribosomes (Morita et al., 1969). These results obtain thus support the evidence and prediction of Gamo & Garcia-Corrales (1988). The observed mode of cell replacement appears to be the same as found in the epidermis (Lentz, 1967).     

Isolation of protoplasts from different Eucalyptus species and preliminary studies on regeneration

Protoplasts were isolated from different Eucalyptus clones and hybrids using mesophyll tissue, calli and cell suspension cultures. The protoplast yields differed greatly according to the starting material and adaptations of the basic procedure had to be designed in specific cases. Eucalyptus protoplasts are representative of recalcitrant woody plant systems since their proliferation is limited in culture. The best results were obtained with protoplasts from cell suspension cultures. A screening of factors increasing proliferation was performed. When some of these factors were combined the cell division frequency was enhanced and microcalli were obtained.Abbreviations B.A.P. Benzylaminopurine - 2-4D 2-4 dichlorophenoxy-acetic acid - F.D.A. Fluorescein diacetate - F.W. Fresh weight - M.E.S. Morpholino ethane sulfonic acid - M.S. Murashige & Skoog (1962) - N.A.A. Naphtalene acetic acid - TRIS Tri (hydroxymethyl amino methane) - V.KM. Medium-Kao and Michayluk medium modified by Vasil (Vasil & Vasil 1980)     

Towards the prediction of non-peptidic epitopes

In-silico methods for the prediction of epitopes can support and improve workflows for vaccine design, antibody production, and disease therapy. So far, the scope of B cell and T cell epitope prediction has been directed exclusively towards peptidic antigens. Nevertheless, various non-peptidic molecular classes can be recognized by immune cells. These compounds have not been systematically studied yet, and prediction approaches are lacking. The ability to predict the epitope activity of non-peptidic compounds could have vast implications; for example, for immunogenic risk assessment of the vast number of drugs and other xenobiotics. Here we present the first general attempt to predict the epitope activity of non-peptidic compounds using the Immune Epitope Database (IEDB) as a source for positive samples. The molecules stored in the Chemical Entities of Biological Interest (ChEBI) database were chosen as background samples. The molecules were clustered into eight homogeneous molecular groups, and classifiers were built for each cluster with the aim of separating the epitopes from the background. Different molecular feature encoding schemes and machine learning models were compared against each other. For those models where a high performance could be achieved based on simple decision rules, the molecular features were then further investigated. Additionally, the findings were used to build a web server that allows for the immunogenic investigation of non-peptidic molecules (http://tools-staging.iedb.org/np_epitope_predictor). The prediction quality was tested with samples from independent evaluation datasets, and the implemented method received noteworthy Receiver Operating Characteristic-Area Under Curve (ROC-AUC) values, ranging from 0.69–0.96 depending on the molecule cluster.     

The use of TTC reduction assay for assessment of Gentiana spp. cell suspension viability after cryopreservation

This study was aimed at improving the 2,3,5-triphenyl-tetrazoliumchloride (TTC) reduction test for initial assessment of cell survival after cryopreservation. Experiments were carried out on three embryogenic cell suspensions of different ages: 9-year-old Gentiana tibetica (King ex Hook. F.), 2-year-old G. kurroo (Royle), and 1-year-old G. cruciata (L.). The suspensions were maintained in MS medium supplemented with 1.0 mg 1⁻¹ 3,6-dichloro-o-anisic acid, 0.1 mg 1⁻¹ naphthaleneacetic acid, 2.0 mg l⁻¹ 6-benzylaminopurine, 80.0 mg 1⁻¹ adenine sulphate and 0.09 M sucrose. Four weeks before freezing, part of the tissue was subcultured to the same medium with sucrose concentrations elevated from 0.09 M (3%sMS) to 0.175 M (6%sMS) or 0.26 M (9%sMS). In freezing treatments without cryoprotection, tissue was plunged directly into liquid nitrogen (LN) or cooled gradually. In freezing treatments with cryoprotection, the cells were pretreated with 1 M sucrose, or with 0.4 M sorbitol + 0.25 M proline or + 0.08 M DMSO, or with vitrification solution (PVS2). Encapsulation was another variant. TTC reduction activity was spectrophotometrically assessed immediately, 1, 3, 5, 24 and 48 h after thawing. Cells without cryoprotection were lethally damaged, but TTC reduction activity in those cells ranged from 6.5% (tissue from 3%sMS) to 73 % (tissue from 9%sMS) directly after thawing. Formazan production was reduced to zero after 24 h. The TTC test showed 50% formazan content immediately after thawing of DMSO-protected G. tibetica tissue, but only 22.47% after 24 h and 2.9% after 48 h. Ultrastructural analysis of those cells showed lethal damage in many of them. For the PVS2 treatment, the formazan content was similar in samples analyzed directly after thawing and 24 h later. Cells treated with PVS2 did not show structural disturbances. Encapsulated cell aggregates of G. cruciata treated with concentrations of sucrose increasing up to 1 M produced 2.6 times more formazan. When applied at least 48 h after thawing, the TTC test can reflect cell viability and can be used to compare the effectiveness of cryoprotectant performance and freezing protocols, but it must be carefully evaluated, with appropriate controls.     

A cytogenetic study of four species of cockatoos and Amazon parrots

The karyotypes of three species of cockatoos, Cacatua moluccensis, Cacatua goffini, and Cacatua sanguinea, and one Amazon parrot, Amazona aestiva, are described for the first time. These karyotypes are compared to the twenty-three species of psittacines published previously. Cacatua moluccensis appears to be a link in the evolutionary tree between Probosciger aterrimus and the other cockatoo species studied. This study reaffirms that the cockatoos are probably most closely related to the loris (Loriculus sp.) and Amazon parrots based on karyotype (M. W. M. Van Dongen & L. E. M. De Boer, (Genetica 65: 109–117) 1984).     

Propagation and xanthone content of <Emphasis Type="Italic">Gentianella austriaca</Emphasis> shoot cultures

Shoot cultures of Gentianella austriaca (A. & J. Kerner) Dostal established from seedling epicotyls were maintained on MS medium supplemented with 2.22 μM BA and 0.54 μM NAA. A characteristic feature of these cultures was precocious flowering, which appeared in all rapidly elongating shoots. Flower development arrested shoot elongation and multiplication of shoot cultures. Continuous shoot propagation was possible only by use of small axillary or adventitious buds as explants for subculturing. Flowering could not be suppressed by GA₃ addition or by cultivation in short-day conditions. The highest rooting percentage (47.3% with 7.83 roots per explant) was achieved on media with 4.92 μM IBA. Shoot cultures contained the same types of secondary metabolites as plants from nature. Xanthones were the major constituents, with DMB (demethylbellidifolin), DGL (demethylbellidifolin-8-O-glucoside) and BGL (bellidifolin-8-O-glucoside) present at roughly two times lower concentrations than in samples from nature. Secondary metabolite production was strongly affected by the presence of BA in the medium.     

苦苣苔科凹柱苣苔属在越南的发现及水晶凹柱苣苔的补充描述

中越边境喀斯特地区有着异乎寻常的植物多样性,已经成为全球生物多样性研究和保护的热点区域,近年来在该区发现了多个苦苣苔科植物的新种或新属。凹柱苣苔属是2010年才被建立的新属,该属的花与叶相比,花较大,叶相对较小,叶片边缘稍外卷,花序仅单花,柱头盘状或凹坑状,蒴果长椭球形而有别于苦苣苔科的其他属。凹柱苣苔属为喀斯特专性植物,目前仅包括分布于黔西南的凹柱苣苔和分布于滇东南的水晶凹柱苣苔两个物种。作者在开展中越边境喀斯特地区植物多样性调查时,在越南北部的河江省发现了水晶凹柱苣苔,基于该新发现居群的植物标本,对其进行了补充描述,确认其花期为11月—12月,并对该物种濒危状况进行了重新评估,同时提供了野外生态照片以资辨认。目前,越南已记录苦苣苔科植物32属150余种,凹柱苣苔属在越南北部的发现,不仅丰富了该国苦苣苔科植物的多样性,再一次充分证明了中越边境喀斯特地区是一个完整的植物区系地理单元,而且为稀有的水晶凹柱苣苔提供了更加丰富的野外本底资料,对其将来的深入研究和保护具有重要的生物地理学和保护生物学意义。     

Taxonomic notes on African Orthotrichaceae I. New synonymy in Macromitrium

New synonymy and distribution records for the genus Macromitrium Brid. in sub-Saharan Africa are presented. Seven names are placed in synonymy, reducing the number of accepted taxa for Macromitrium in sub-Saharan Africa to 33. Macromitrium anomodictyon Cardot was found to be conspecific with M. serpens (Bruch ex Hook. & Grev.) Brid.; M. mauritianum Schwägr. and M. sanctae-mariae Renauld & Cardot are placed in synonymy with M. subtortum (Hook. & Grev.) Schwägr. and M. belangeri Müll.Hal., M. seriatum Paris & Broth., M. bequaertii Thér. & Naveau and M. trollii Dixon in Herz. are all placed in synonymy with M. sulcatum (Hook.) Brid. subsp. sulcatum. The precise lectotypification is implemented for six species within the genus. Macromitrium chloromitrium (Besch.) Wilbraham and M. orthostichum Nees ex Schwägr. are reported new to Tanzania and M. chloromitrium new to continental Africa.     

In vitro cold storage of cork oak shoot cultures

Friable callus cultures were initiated from cotyledons and hypocotyls of Opuntia ficus-indica. Explants from cotyledons produced significantly more callus than those from hypocotyls. Optimum callus growth was observed on Murashige & Skoog medium supplemented with 0.9 μM 6-furfurylaminopurine, 2.3 μM 2,4-dichlorophenoxyacetic acid, 1.0 μM 4-amino 3,5,6-trichloropicolinic acid, 400 mg l^-1 casein hydrolysate and 3% sucrose. The same medium without agar was used for establishing cell suspensions. This revised version was published online in June 2006 with corrections to the Cover Date.     

Pre- and post-synaptic actions of the cerebral ventral giant cell on buccal muscles ofPhiline

Summary The cerebral ventral giant cell ofPhiline exterts a selective presynaptic inhibitory modulatory action on the terminals of a buccal excitatory motoneuron in two buccal muscles. Other excitatory inputs to the muscles are not affected. The ventral giant cell also makes direct synaptic contacts on the fibres of the same muscles. In the retractor muscle M4, 5 the junction potentials are usually depolarising when measured in sea water, but in the fibres of M6 they may have either polarity. The mean membrane potential of the fibres of M4, 5 and M6 was –74.7±0.65 mV and –64±0.95 mV respectively. Depolarization of the muscles fibres by around 15 mV by immersion in 20 mM K saline abolished the junction potential in M4, 5 and converted the depolarizing potential in M6 to a hyperpolarizing response.It is concluded that the VGC junction potential results from an increase in membrane conductance to an ion with a reversal potential between –60 and –70 mV.     

Revision of the genus Diclidophora Krøyer, 1838 (Monogenea: Diclidophoridae), with the proposal of Macrouridophora n. g.

The present study re-examines the detailed morphology of the type-species, Diclidophora merlangi (Kuhn, in Nordmann, 1832) Krøyer, 1838, and other Diclidophora species parasitic on gadid fishes: D. denticulata (Olsson, 1876) Price, 1943, D. esmarkii (Th. Scott, 1901) Sproston, 1946, D. luscae (van Beneden & Hesse, 1863) Price, 1943, D. minor (Olsson, 1868) Sproston, 1946, D. palmata (Leuckart, 1830) Diesing, 1850, D. phycidis (Parona & Perugia, 1889) Sproston, 1946, D. pollachii (van Beneden & Hesse, 1863) Price, 1943 and the recently described D. micromesisti Suriano & Martorelli, 1984. An amended generic diagnosis of Diclidophora Krøyer, 1838 (synonym Diclidophora Diesing, 1850) is provided, which includes the presence of a prostatic vesicle in the terminal male genitalia and the distal fusion of the median and peripheral sclerites, a₁ and c₁ in the clamp anterior jaw. Macrouridophora n. g. is herein proposed for species previously considered in Diclidophora, which are parasitic on macrourid and morid fishes. The clamp morphology in Macrouridophora n. g. has distinct lamellate extension attachments to peripheral sclerites c₁ and the distal portion of d₁, with no distal fusion between a₁ and c₁ in the anterior jaw. Macrouridophora macruri (Brinkmann, 1942) n. comb. is chosen as the type-species. Nine other species are herein transferred to Macrouridophora n. g.: M. coelorhynchi (Robinson, 1961) n. comb., M. lotella (Machida, 1972) n. comb., M. nezumiae (Munroe, Campbell & Zwerner, 1981) n. comb. and M. tubiformis (Rohde & Williams, 1987) n. comb. are redescribed, based on the re-examination of type or voucher specimens. Macrouridophora attenuata (Mamaev & Zubtschenko, 1979) n. comb., M. caudata (Mamaev & Zubtschenko, 1984) n. comb., M. papilio (Mamaev & Avdeev, 1981) n. comb., M. paracoelorhynchi (Mamaev & Paruchin, 1979) n. comb. and M. physiculi (Mamaev & Avdeev, 1981) n. comb. have adequately described haptoral clamps in the literature. The clamp morphology in Macrouridophora sp. from Lepidorhynchus denticulatus in Australia is also considered. Diclidophora whitsonii Suriano & Martorelli, 1984 is herein transferred to the genus Macruricotyle Mamaev & Ljadov, 1975, as M. whitsonii (Suriano & Martorelli, 1984) n. comb. D. embiotocae Hanson, 1979 is herein considered a species incertae sedis. D. caudospina Khan & Karyakarte, 1983 and D. paddiforma Deo & Karyakarte, 1979 are herein considered species inquirendae. D. aglandulosa Deo, 1977, D. glandulosa Das, 1972, D. minuta Das, 1972 and D. spindale Deo, 1977 are formally dismissed as nomina nuda. The systematic position of Diclidophora Krøyer, 1838 and Macrouridophora n. g. in the subfamily Diclidophorinae Cerfontaine, 1895 (sensu Mamaev, 1976) is discussed.     

Micropropagation and encapsulation of medicinally important Chonemorpha grandiflora

Summary This study describes a protocol for in vitro propagation of Chonemorpha grandiflora (Roth) S.M. & M.R. Almeida (Apocynaceae) through axillary bud proliferation and rooting. For shoot induction, the combination of 13.3 &#x03BC;M N ⁶-benzyladenine (BA) and 2.46 &#x03BC;M indole-3-butyric acid (IBA) in Murashige and Skoog (MS) medium was better than other growth regulators. Liquid medium was superior over solid medium for root formation and growth. IBA was more effective than other auxins for root induction. Addition of silver nitrate in the presence of IBA significantly increased the number, length, and branching of roots. Shoot tips encapsulated in medium containing 0.49 &#x03BC;M IBA and 11.7 &#x03BC;M silver nitrate exhibited 95% conversion to plantlets. This protocol enables the production of 250 plantlets from a single nodal explant within 7 mo. Plantlets showed 90% survival after acclimatization in soil and were morphologically similar to the source plant under field conditions.