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A distributed and decentralised ledger system shared by a variety of users in a peer-to-peer network is defined as a blockchain. Through its decentralised approach, it makes pharmaceutical products more accessible and secure for vendors, while they can maintain and access their inventories themselves. Between a company and its suppliers, a supply chain is used to produce and distribute a specific product or service, whose efficiency is increased with its integration in the supply chain. For efficient and transparent tracking of products, blockchain will be the right choice. In view of blockchain being incorporated into the supply chain system, it solves many logistical issues that supply chains face, such as appropriate data access, ensuring data quality, and many more. This can increase the traceability of the material supply chain and improve the maintenance of the list when there are several products to be supplied. Therefore, the concept of supply chain was developed in two platforms namely, Hyperledger Fabric and Hyperledger Sawtooth. Finally, Hyperledger Sawtooth was found to be well suited for creating decentralised apps or platforms with respect to resource utilization It enables developers to separate their application domain from the core system so that business rules for apps can exist without requiring knowledge of the primary platform's underlying architecture. The proposed system shows that Hyperledger Sawtooth framework consumes lower CPU than Fabric which helps in increasing the number of transactions.
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A 4-day-old female neonate with raised cord blood thyroid-stimulating hormone (127 µIU/mL) underwent Tc thyroid scan to rule out thyroid dysgenesis. The images revealed midline focus of lingual thyroid as the only functioning thyroid tissue. In addition, bilateral focal and symmetrical breast uptake was seen without clinically palpable breast nodule on either side. Transplacental transfer of maternal hormones leading to stimulation of neonatal breasts explains this unusual scan finding. One should be aware of this rare pattern of focal breast uptake in Tc-pertechnetate scan in neonates with congenital hypothyroidism to avoid scan misinterpretation.
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Mama/metabolismo , Hipotiroidismo Congénito/metabolismo , Pertecnetato de Sodio Tc 99m/metabolismo , Transporte Biológico , Mama/diagnóstico por imagen , Hipotiroidismo Congénito/diagnóstico por imagen , Femenino , Humanos , Recién Nacido , CintigrafíaRESUMEN
The nucleation control and separation of mono and ortho polymorphs of the important pharmaceutical solid paracetamol were carried out through a crystallization process in gel media for the first time. Crystallization of mono and ortho polymorphic forms of paracetamol was achieved by optimizing the experimental parameters such as the specific gravity, pH, height of the gel column and solute concentration at ambient temperature. The optimized experimental conditions favor the generation of necessary supersaturation responsible for the nucleation of preferred polymorph at different levels in the gel column and also endure the stability of the grown orthorhombic polymorphs at ambient conditions. Accordingly the needle like metastable orthorhombic polymorph nucleates at the top portion of the gel column whereas the prismatic stable monoclinic polymorph nucleates mostly at the bottom level. Morphology of the nucleated polymorphs was analyzed and their crystalline structures were confirmed by PXRD. FTIR analysis revealed the shifting of absorption peaks of few functional groups corresponding to both the polymorphs due to the difference in their structural nature. DSC analysis revealed that the grown ortho polymorph form II transforms to mono form I at 89.47°C while the grown mono form I retains its phase until melting.
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Acetaminofén/química , Rastreo Diferencial de Calorimetría , Cristalización , Geles , Difracción de Polvo , Espectroscopía Infrarroja por Transformada de FourierRESUMEN
Analysis of any mammalian plasma proteome is a challenge, particularly by mass spectrometry, due to the presence of albumin and other abundant proteins which can mask the detection of low abundant proteins. As detection of human plasma proteins is valuable in diagnostics, exploring various workflows with minimal fractionation prior to mass spectral analysis, is required in order to study population diversity involving analysis in a large cohort of samples. Here, we used 'reference plasma sample', a pool of plasma from 10 healthy individuals from Indian population in the age group of 25-60 yrs including 5 males and 5 females. The 14 abundant proteins were immunodepleted from plasma and then evaluated by three different workflows for proteome analysis using a nanoflow reverse phase liquid chromatography system coupled to a LTQ Orbitrap Velos mass spectrometer. The analysis of reference plasma sample a) without prefractionation, b) after prefractionation at peptide level by strong cation exchange chromatography and c) after prefractionation at protein level by sodium dodecyl sulfate polyacrylamide gel electrophoresis, led to the identification of 194, 251 and 342 proteins respectively. Together, a comprehensive dataset of 517 unique proteins was achieved from all the three workflows, including 271 proteins with high confidence identified by ≥ 2 unique peptides in any of the workflows or identified by single peptide in any of the two workflows. A total of 70 proteins were common in all the three workflows. Some of the proteins were unique to our study and could be specific to Indian population. The high-confidence dataset obtained from our study may be useful for studying the population diversity, in discovery and validation process for biomarker identification.
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Proteínas Sanguíneas/metabolismo , Voluntarios Sanos , Proteoma/metabolismo , Adulto , Biología Computacional , Secuencia de Consenso , Femenino , Humanos , India , Masculino , Persona de Mediana Edad , Peso Molecular , Péptidos/metabolismoRESUMEN
OBJECTIVE: To screen Isolated Growth Hormone Deficiency (IGHD) patients with congenital Familial Isolated (FIGHD) and Nonfamilial Isolated Growth Hormone Deficiency (NFIGHD) for GH1gene deletions (6.7 kb,7.6 kb,7 kb) and Growth hormone releasing hormone receptor GHRHR(E72X) gene mutation and study genotype/phenotype correlation in this multicentre study. METHODS: Clinical, auxologic (Ht.SDS ≤ -2.5), hormonal and MRI evaluation of hypothalamic/pituitary (HP) axis, IGF1, IGFBP3 estimation and GH stimulation test confirmed IGHD in 107 patients. Of these 107 patients, 97 consented for molecular genetic studies. Height, weight and Bone Age (BA) were obtained. PCR based restriction digestion method was used for molecular genetic analysis of patients and families. Ethics committee approval was obtained. RESULTS: Based on the genotype, these 97 patients (M60,F37;1.62:1) age 3 mo to 17 y belonging to 80 families (consanguinity, 15/80), were categorized into Group I with GH1 gene deletion, n = 17 (17.5 %) from 14 families, Group II with GHRHR (E72X) mutation n = 34 (35 %) from 24 families, Group III, n = 46 (47 %) from 42 families having neither of these deletions/mutations (but with sibling involvement). In Group I, homozygous 6.7 kb and 7.6 kb deletions involved 76 % and 18 %. 6.7 kb deletion with characteristic IGHD phenotype predominated in nonconsanguineous community from Rajasthan having lowest mean FBS (55.6 mg/dl, p < 0.001) and peak GH (0.03 ng/dl, p < 0.01). In Group II phenotype was IB. Twenty one of the 23 FIGHD had homozygous GHRHR(E72X) mutation and four with IGHD had heterozygous GHRHR(E72X) mutation. IGF1 and IGFBP3 were low. MRI showed hypoplastic anterior pituitary (APH) in all. Group III is not discussed in detail. CONCLUSIONS: Genetic background is more likely in congenital Growth Hormone Deficiency (GHD). GH1gene deletions and GHRHR(E72X) mutation with characteristic phenotypes are encountered in North Western region of India. Regional studies are essential.
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Enanismo Hipofisario/genética , Adolescente , Niño , Preescolar , Consanguinidad , Electroforesis en Gel de Agar , Femenino , Eliminación de Gen , Genotipo , Hormona del Crecimiento/genética , Hormona Liberadora de Hormona del Crecimiento/genética , Humanos , Lactante , Masculino , FenotipoRESUMEN
Glioblastomas (GBMs) are the most common and lethal primary tumors of the central nervous system with high level of recurrence despite aggressive therapy. Tumor-associated proteins/peptides may appear in the plasma of these patients as a result of disruption of the blood-brain barrier in them, raising the scope for development of plasma-based tests for diagnosis and monitoring the disease. With this objective, we analyzed the levels of proteins present in the plasma from GBM patients using an iTRAQ based LC-MS/MS approach. Analysis with pooled plasma specimens from the patient and healthy control samples revealed high confidence identification of 296 proteins, of which 61 exhibited a fold-change ≥1.5 in the patient group. Forty-eight of them contained signal sequence. A majority have been reported in the differentially expressed transcript or protein profile of GBM tissues; 6 have been previously studied as plasma biomarkers for GBM and 16 for other types of cancers. Altered levels of three representative proteins-ferritin light chain (FTL), S100A9, and carnosinase 1 (CNDP1)-were verified by ELISA in a test set of ten individual plasma specimens. FTL is an inflammation marker also implicated in cancer, S100A9 is an important member of the Ca(2+) signaling cascade reported to be altered in GBM tissue, and CNDP1 has been reported for its role in the regulation of the levels of carnosine, implicated as a potential drug for GBM. These and other proteins in the dataset may form useful starting points for further clinical investigations for the development of plasma-based biomarker panels for GBM.
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Apoferritinas/genética , Biomarcadores de Tumor/genética , Neoplasias Encefálicas/genética , Calgranulina B/genética , Dipeptidasas/genética , Regulación Neoplásica de la Expresión Génica , Glioblastoma/genética , Proteínas de Neoplasias/genética , Apoferritinas/sangre , Biomarcadores de Tumor/sangre , Neoplasias Encefálicas/sangre , Neoplasias Encefálicas/diagnóstico , Calcio/metabolismo , Calgranulina B/sangre , Carnosina/metabolismo , Cromatografía Liquida , Dipeptidasas/sangre , Ensayo de Inmunoadsorción Enzimática , Femenino , Glioblastoma/sangre , Glioblastoma/diagnóstico , Humanos , Masculino , Persona de Mediana Edad , Proteínas de Neoplasias/sangre , Proteómica/métodos , Transducción de Señal , Espectrometría de Masas en TándemRESUMEN
Composite blend microbeads of sodium alginate (NaAlg) with sodium carboxymethyl cellulose (NaCMC) containing magnesium aluminum silicate (MAS) particles and enteric coated with chitosan have been prepared to achieve controlled release (CR) of amoxicillin in stomach environment. The composite beads have been characterized by X-ray diffraction (XRD) to study drug distribution, DSC for understanding thermal stability and Fourier transform infrared (FTIR) spectroscopy to investigate chemical interactions as well as to assess the structure of the drug-loaded formulations. Surface morphology of the beads was investigated by scanning electron microscopy (SEM). The size distribution of beads loaded with drug as studied by particle size analyzer was in the range of 745-889 µm. The beads exhibited quite widely varying encapsulation efficiencies from 52 to 92%. Equilibrium swelling of the beads measured in water and in vitro release of amoxicillin in pH 1.2 medium suggests that drug release depends on polymer blend composition, concentration of MAS and extent of enteric coating.
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Alginatos/química , Amoxicilina/administración & dosificación , Quitosano/química , Microesferas , Amoxicilina/química , Rastreo Diferencial de Calorimetría , Química Farmacéutica , Preparaciones de Acción Retardada/química , Ácido Glucurónico/química , Ácidos Hexurónicos/química , Tamaño de la Partícula , Espectroscopía Infrarroja por Transformada de Fourier , Difracción de Rayos XRESUMEN
Membrane proteins play key roles in the development and progression of cancer. We have studied differentially expressed membrane proteins in glioblastoma multiforme (GBM), the most common and aggressive type of primary brain tumor, by high resolution LC-MS/MS mass spectrometry and quantitation by iTRAQ. A total of 1834 membrane proteins were identified with high confidence, of which 356 proteins were found to be altered by 2-fold change or more (198 up- and 158 down-regulated); 56% of them are known membrane proteins associated with major cellular processes. Mass spectrometry results were confirmed for representative proteins on individual specimens by immunohistochemistry. On mapping of the differentially expressed proteins to cellular pathways and functional networks, we notably observed many calcium-binding proteins to be altered, implicating deregulation of calcium signaling and homeostasis in GBM, a pathway also found to be enriched in the report (Dong, H., Luo, L., Hong, S., Siu, H., Xiao, Y., Jin, L., Chen, R., and Xiong, M. (2010) Integrated analysis of mutations, miRNA and mRNA expression in glioblastoma. BMC Syst. Biol. 4, 163) based on The Cancer Genome Atlas analysis of GBMs. Annotations of the 356 proteins identified by us with The Cancer Genome Atlas transcriptome data set indicated overlap with 295 corresponding transcripts, which included 49 potential miRNA targets; many transcripts correlated with proteins in their expression status. Nearly 50% of the differentially expressed proteins could be classified as transmembrane domain or signal sequence-containing proteins (159 of 356) with potential of appearance in cerebrospinal fluid or plasma. Interestingly, 75 of them have been already reported in normal cerebrospinal fluid or plasma along with other proteins. This first, in-depth analysis of the differentially expressed membrane proteome of GBM confirms genes/proteins that have been implicated in earlier studies, as well as reveals novel candidates that are being reported for the first time in GBM or any other cancer that could be investigated further for clinical applications.
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Neoplasias Encefálicas/metabolismo , Señalización del Calcio , Glioblastoma/metabolismo , Proteínas de la Membrana/metabolismo , Proteoma/metabolismo , Secuencia de Aminoácidos , Estudios de Casos y Controles , Cromatografía Liquida , Femenino , Regulación Neoplásica de la Expresión Génica , Humanos , Masculino , Proteínas de la Membrana/química , Proteínas de la Membrana/genética , Persona de Mediana Edad , Datos de Secuencia Molecular , Fragmentos de Péptidos/química , Proteoma/química , Proteoma/genética , Espectrometría de Masas en Tándem , Análisis de Matrices TisularesRESUMEN
Glioblastoma multiforme (GBM) is the most aggressive among human gliomas with poor prognosis. Study of tumor cell secretome - proteins secreted by cancer cell lines, is a powerful approach to discover potential diagnostic or prognostic biomarkers. Here we report, for the first time, proteins secreted by three GBM cell lines, HNGC2, LN229 and U87MG. Analysis of the conditioned media of these cell lines by LC-MS/MS using ESI-IT mass spectrometer (LTQ) resulted in the confident identification of 102, 119 and 64 proteins, respectively. Integration of the results from all the three cell lines lead to a dataset of 148 non-redundant proteins. Subcellular classification using Genome Ontology indicated that 42% of the proteins identified belonged to extracellular or membrane proteins, viz. Vinculin, Tenascin XB, SERPIN F1 and TIMP-1. 52 proteins matched with the secretomes of 11 major cancer types reported earlier whereas remaining 96 are unique to our study. 25 protein identifications from the dataset represent proteins related to GBM or other cancer tissues as per Human Protein Atlas; at least 22 are detectable in plasma, 11 of them being reported even in cerebrospinal fluid. Our study thus provides a valuable resource of GBM cell secretome with potential for further investigation as GBM biomarkers.
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Glioblastoma/metabolismo , Proteínas de Neoplasias/análisis , Línea Celular Tumoral , Perfilación de la Expresión Génica , Glioblastoma/genética , Humanos , Proteínas de Neoplasias/genética , Espectrometría de Masa por Ionización de ElectrosprayRESUMEN
Glutaraldehyde crosslinked interpenetrating polymer network (IPN) blend microspheres of chitosan (CS) and hydroxyethyl cellulose (HEC) were prepared in the form of microspheres (66-82 microm dia) and investigated for the controlled release (CR) of isoniazid (INH), an antituberculosis drug. Microspheres were characterized by X-ray diffraction (XRD) to study the uniform distribution of drug in the matrices, differential scanning calorimetry (DSC) and thermogravimetry (TGA) to study thermal stabilities of IPN blends. Fourier transform infrared (FTIR) spectroscopy was used to understand chemical interactions and to assess the IPN structure. Scanning electron microscopy (SEM) was employed to investigate the morphology of microspheres. Drug-loaded microspheres were produced in spherical shapes with encapsulation efficiency ranging from 50 to 66%. Equilibrium swelling measured in pH 7.4 buffer solution as well as in vitro release of drug in pH 1.2 and 7.4 buffer solutions indicated the dependence of drug release on crosslinking as well as blend composition of the IPN matrix.
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Celulosa/análogos & derivados , Quitosano/química , Portadores de Fármacos/química , Isoniazida/química , Isoniazida/metabolismo , Microesferas , Tampones (Química) , Rastreo Diferencial de Calorimetría , Celulosa/química , Preparaciones de Acción Retardada/química , Preparaciones de Acción Retardada/metabolismo , Difusión , Concentración de Iones de Hidrógeno , Microscopía Electrónica de Rastreo , Tamaño de la Partícula , Espectroscopía Infrarroja por Transformada de Fourier , Termogravimetría , Difracción de Rayos XRESUMEN
INTRODUCTION: Wasting is the cardinal feature of tuberculosis, but not much documentary evidence supporting food supplements exists. This study was done to assess the effects of food supplements on body weight, physical function, quality of life and treatment outcomes in patients with tuberculosis and wasting. METHODS: The study was conducted in 30 Anganwadi centres of 16 villages in the catchment area of Pinnamaneni Siddhartha Institute of Medical Sciences and Research Foundation and the Gannavaram Directly Observed Treatment Short Course chemotherapy centre from August 2005 to December 2005. A total of 100 patients participated in the study. Patients who were started on anti-tubercular therapy within the previous two weeks were randomly assigned to either the control or the food supplement group. At the end of three months, their body weight was measured and physical function and quality of life were assessed. Treatment outcomes were assessed at the one-year follow-up for both groups. RESULTS: Patients who received supplements had a significant increase in body weight (8.6 percent versus 2.6 percent, p-value less than 0.001) and maximum grip strength (p-value less than 0.001), a higher sputum conversion rate (p-value is 0.039), a higher treatment completion rate (p-value is 0.031) and improvements in the quality of life scores. CONCLUSION: Intake of food supplements resulted in a definitive increase in body weight and physical function in our study sample. Improvements can be observed in all areas, including psychologically, physiologically, socially and in the treatment outcomes.
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Suplementos Dietéticos , Tuberculosis Pulmonar/dietoterapia , Síndrome Debilitante/dietoterapia , Adulto , Antituberculosos/uso terapéutico , Peso Corporal , Femenino , Humanos , Masculino , Calidad de Vida , Resultado del Tratamiento , Tuberculosis Pulmonar/tratamiento farmacológico , Síndrome Debilitante/tratamiento farmacológico , Aumento de PesoRESUMEN
Synthesis of nanocomposite thin films of CrN/Cu deposited on (100) Si and D-9 alloy substrates by pulsed magnetron sputtering as a function of copper content in the range 15.1-35.8 at.% is investigated. XRD analysis of the films deposited at 773 K with nitrogen flow rate of 10 sccm indicated that the films are nanorystalline and bi-phasic (fcc-CrN and fcc-Cu). Scanning electron microscopy showed a structureless morphology for CrN, while agglomerates were obtained for CrN/Cu nanocomposite thin films. Atomic force microscopy also confirmed the agglomeration of particles with increasing Cu content. The amount of copper content in the nanocomposite films had also shown a significant reduction in the crystallite size of CrN. The nano hardness measurements showed a peak hardness of 17 GPa for the films with copper content of 15.1 at.%. The hardness values were found to decrease significantly with Cu content > 31.1 at.%.
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Transgenic hairy roots were induced from Withania somnifera Dunal, by infecting leaf explants with Agrobacterium rhizogenes. Polymerase chain reaction for rol A gene and Southern blot confirmed the integration of T-DNA in the genome. Cultures were grown in Murashige and Skoog solid as well as in liquid medium. The antioxidant activity was assayed in roots grown in solid media and liquid media. Hairy roots grown in liquid media found to possess highly significant activity in 1,1-diphenyl-2-pecryl-hydrazyl radical, beta-carotene linoleic acid model system. The activity was 57.34%, 75.64%, and 93.41% in case DPPH model and 55.3%, 76.3%, and 90.5% in case of b-CLAMS in 25, 50, and 100 mg L(-1) concentration, respectively. In case of hydroxyl radical trapping and brain lipid peroxidation assay, the activity was more significant in hairy roots grown on solid medium in comparison with commercial formulation prepared using normal roots and standard withanaloids. Root extract grown in solid medium has shown 93.2% hydroxyl radical trapping activity at 100 mg L(-1) concentration, and 500 mg L(-1) has shown 83.6% in case of brain lipid peroxidation assay. High-performance liquid chromatography analysis demonstrated the presence of withanaloids in the hairy root extracts. The results of the study clearly indicate that there is enhancement of secondary metabolites in hairy roots, which is indicated through significant enhancement of the antioxidant activity, since these are the major constituents responsible for the activity. This is the first report on the presence of antioxidant principles in genetically modified roots of W. somnifera. These results of the present study may aid in utilization of the W. somnifera hairy roots for its rejuvenating principles.
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Extractos Vegetales/uso terapéutico , Plantas Modificadas Genéticamente , Rejuvenecimiento/fisiología , Withania , Animales , Antioxidantes/metabolismo , Encéfalo/metabolismo , Cromatografía Liquida , Peroxidación de Lípido , Extractos Vegetales/metabolismo , Transformación GenéticaRESUMEN
Hairy roots of Rauvolfia micrantha were induced from hypocotyl explants of 2-3 weeks old aseptic seedlings using Agrobacterium rhizogenes ATCC 15834. Hairy roots grown in half-strength Murashige & Skoog (MS) medium with 0.2 mg indole 3-butyric acid l-1 and 0.1 mg alpha-naphthaleneacetic acid l-1 produced more ajmaline (0.01 mg g-1 dry wt) and ajmalicine (0.006 mg g-1 dry wt) than roots grown in auxin-free medium. Ajmaline (0.003 mg g-1 dry wt) and ajmalicine (0.0007 mg g-1 dry wt) were also produced in normal root cultures. This is the first report of production of ajmaline and ajmalicine in hairy root cultures of Rauvolfia micrantha.
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Ajmalina/biosíntesis , Raíces de Plantas/metabolismo , Rauwolfia/metabolismo , Alcaloides de Triptamina Secologanina , Yohimbina/análogos & derivados , Yohimbina/metabolismo , Diferenciación Celular , Técnicas de Cultivo/métodos , Ácidos Indolacéticos/farmacología , Raíces de Plantas/citología , Raíces de Plantas/crecimiento & desarrollo , Raíces de Plantas/microbiología , Plantas Medicinales/citología , Plantas Medicinales/crecimiento & desarrollo , Plantas Medicinales/metabolismo , Plantas Medicinales/microbiología , Rauwolfia/citología , Rauwolfia/crecimiento & desarrollo , Rauwolfia/microbiología , Rhizobium/fisiologíaRESUMEN
Glucocorticosteroid therapy can occasionally lead to a serious and debilitating complication called osteone-crosis or avascular necrosis. While a prompt diagnosis of this complication can help in preventing significant morbid-ity in the affected patients, dermatologists in general, are often not conscious of this facet of steroid-related problems. The precise mechanisms responsible for this disease are not fully understood nor its prevalence in derma-tological practice. An attempt has been made in this article to review the current understanding of its pathogenesis, the risk factors associated with its occurrence and the methods for its early diagnosis and treatment. An attempt is also made to formulate a set of guidelines useful to a dermatologist for the prevention of this disabling complication.
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Rapid micropropagation of Trichopus zeylanicus Gaertn. subsp. travancoricus Burkil ex Narayanan, a rare ethnomedicinal herb endemic to the Western Ghats of southern India, was achieved by culturing shoot tips (0.3-0.5 cm) of 2-month-old axenic seedlings on Woody Plant Medium. Among the cytokinins tested, only BAP induced callus-free multiple shoot bud formation, with a maximum of 8.5±0.4 buds per explant being obtained with 2.0 mg.l(-1) BAP after 8 weeks of culture. Shoot tips containing proliferated buds were divided and subcultured on medium containing 0.2 mg.l(-1) BAP to produce 12.0±1.0 shoots per explant in 6 weeks. Excision of buds after culture initiation, with subculture of the debudded basal tissue in 2 successive passages yielded 20.0±1.0 and 13.5±0.5 buds per explant respectively. Each bud cultured in turn for 4 weeks on WPM with 1.0 mg.l(-1) BAP formed 3.8±0.4 secondary buds which were repeatedly recultured to increase bud production. Altogether this method enabled an estimated harvest of 7848 buds from a single shoot tip in 28 months. Shoots (3-5 cm) developed from bud cultures were rooted in half-strength WPM medium with 0.5 mg.l(-1) each of NAA and IBA, and 90-100% of the rooted plants were established in the field after hardening. Micropropagated plants were grown to maturity free of defects in growth, morphological, flowering and seed set characteristics.
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Clonal propagation of Adhatoda beddomei C.B. Clarke (Acanthaceae), a rare medicinal shrub, was achieved through callus-free axillary meristem proliferation from stem node explants of field-grown plants cultured in SH medium. Shoot multiplication was a function of cytokinin activity but sustained growth of the shoots was dependent on the synergistic effect with the auxin, IAA. An optimum number of 5-10 shoots per explant were obtained in 6 weeks using 3.0 mg.l(-1) BAP, 0.5 mg.l(-1) 2-ip and 1.0 mg.l(-1) IAA, Upon subculture, vertical halves of the precultured node with the differentiated shoots yielded a larger aggregate number of shoots (23-27) than the uncut precultured node left intact (15-17). Shoot multiplication was rapid and consistent over prolonged periods when the hormonal concentrations were reduced to 1.0 mg.l(-1) BAP and 0.2 mg.l(-1) IAA during subculture, and reculture of the nodal explants derived from shoot cultures. Rooting of 3-5 cm shoots thus obtained was greatly accelerated in stationary liquid medium containing 0.2 mg.l(-1) IBA or IAA. Hardening of the rooted plantlets in the humidity chamber was essential for high frequency (95%) survival. Micropropagated plants established in the field flowered after fifteen months and were free from apparent defects in cytological, growth and flowering characteristics.