Effect of different plant growth regulators on callus and adventitious shoots induction, polysaccharides accumulation and antioxidant activity of Rhodiola dumulosa / 中草药·英文版

OBJECTIVE@#As a medicinal plant, the resource of Rhodiola dumulosa is deficient along with the large collection. For the protection and utilization of R. dumulosa, the influence of plant growth regulators (PGRs) on callus induction and adventitious shoots differentiation, polysaccharide production and the antioxidant activity were tested.@*METHODS@#Internodes of R. dumulosa were used as explants and cultured on MS medium plus different plant growth regulators (PGRs). The anti-oxidative activities of polysaccharides were evaluated using radical scavenging assays.@*RESULTS@#By response surface plot, 0.85 mg/L N6-benzyladenine (BA), 0.34 mg/L naphthaleneacetic acid (NAA) and 0.33 mg/L 2,4-dicholorophenoxyacetic acid (2,4-D) were the optimal factors for callus induction (90.03%) from internodes explants on MS medium. The fresh weight of green callus increased 47.26 fold, when callus was inoculated on MS + thidiazuron (TDZ) 0.5 mg/L + NAA 2.0 mg/L. Adventitious buds regenerated from callus on the media of MS were fortified with BA 1.0 mg/L plus NAA 0.5 mg/L, and the induction rate was 40.00%. MS plus indole-3-butyric acid (IBA) 1.0 mg/L produced the highest rooting rate with 10 to 15 roots in a length of 2-3 cm per shoot. The content of total polysaccharides in callus developed on MS + TDZ 0.5 mg/L + NAA 2.0 mg/L and MS + BA 1.0 mg/L + NAA 0.5 mg/L was as high as 1.72%-2.15%. At the dose of 0.5 mg/mL polysaccharides extracted from different callus induced on MS + NAA 2.0 mg/L + TDZ 0.5 mg/L or MS + BA 1.0 mg/L + NAA 0.5 mg/L or MS + BA 0.5 mg/L + 2,4-D 0.5 mg/L, the ABTS radical eliminating percentages were 82.78%, 80.18% and 68.59%, respectively, much higher than that of wild plant.@*CONCLUSION@#A rapid micropropagation system for R. dumulosa has been developed. The combination of TDZ and NAA or BA and NAA can increase the yield of the total polysaccharides. The polysaccharides isolated from callus and whole wild plants had stronger free radicals scavenging activities, indicating that polysaccharides from R. dumulosa are the potential pharmaceutical supplements.

Evaluation of antioxidant activity and phytochemicals of selected methanol rattan shoot extracts from Morong, Bataan

Background@#Rattan is a member of the Arecaceae family grown in the tropical or subtropical climates. These plants are used as raw materials for furniture-making. In some cases, its shoots are also used as food and may possess bioactive potentials. In this study, rattan root methanolic extracts were screened for phytochemicals and evaluated for antioxidant activities.@*Methodology@#Six (6) methanolic extracts of selected rattan shoots were subjected to in vitro antioxidant assays, which include DPPH, FRAP, metal chelation, superoxide, and nitric oxide. Qualitative and quantitative phytochemical analyses were also employed. @*Results@#Shoots of Calamus sp. 02 (Bangaw-Bangaw; 85.59±0.430%), C. usitatus (Talola; 82.45±2.215%), and C. spinifolius (Kurakling; 75.54±1.599%) showed the most effective DPP radical inhibition at 66.67μg/mL. Concentration-dependent reducing power (>30% scavenging at 6.64μg/mL) with no-to-low metal chelating activity was also observed in these plant food. C. sp 02 and C. usitatus demonstrated an effective scavenging activity against superoxide anions at 227.3μg/mL. Nitric oxide scavenging activity was observed in all shoots with C. merrilli (Palasan) exhibiting highest at 78.13μg/mL. Qualitative phytochemical analyses showed that rattan shoots all contained reducing sugars, phenolics, terpenoids, and quinone compounds. Quantitative phytochemical analyses revealed that C. sp. 02 (66.024±4.183mg GAE/g) and C. merrilli (1.767±0.056mg QE/g) contained the highest amounts of phenolic and flavonoids, respectively. These phytochemicals present may explain their behavior as antioxidants. @*Conclusion@#The study revealed that different rattan shoots showed different capacities to scavenge particular oxidants. Of these, C. sp. 02, C. spinifolius, and C. merrilli may be considered promising sources of natural antioxidants.

Cytokinin induces the development of gabirobeira root cuttings / Citocininas induzem o desenvolvimento de estacas de raiz de gabirobeira

ABSTRACT: The gabirobeira is a species native to the Brazilian Cerrado with potential for use in cropping systems. This study evaluated the effect of the cytokinin 6-benzylaminopurine (BAP) on root cuttings of gabirobeira (Campomanesia adamantium). The plant material was obtained from gabirobeira progenies of one and two-years-old. The cuttings were segmented in 5 cm length and 1.90 to 3.22 mm diameter, immersed in the following BAP concentrations: 0.0; 1.0; 2.0 and 4.0 mg L-1 for 15 seconds and planted in trays containing the substrate Bioplant®. A complete randomized experimental design was adopted in a factorial scheme 2x4, (cuttings age x BAP concentrations) with fifteen replicates per treatment. After 140 days the number of cuttings with shoots, number of shoots, number of leaves, and diameter of the main root were evaluated. The better development of the cuttings was observed on progenies of two-years-old. The lowest cytokinin concentrations promoted the better emission and number of shoots of the progenies from both ages.

RESUMO: A gabirobeira é uma espécie nativa do Cerrado brasileiro com potencial para uso em sistemas de cultivo. O objetivo deste estudo foi avaliar o efeito da 6-benzilaminopurina (BAP) em estacas radiculares de progênies de Gabirobeira (Campomanesia adamantium). As estacas radiculares foram obtidas de progênies de Gabirobeira de um e dois anos de idade. Estes foram segmentados em 5 cm de comprimento e apresentavam entre 1,90 a 3,22 mm de diâmetro, imersos nas concentrações: 0.0; 1.0; 2.0 e 4.0 mg L-1 de BAP por 15 segundos e plantados em bandejas contendo Bioplant®. O delineamento experimental foi o inteiramente casualizado, em esquema fatorial 2x4 (idades das estacas x concentrações de BAP), com quinze repetições por tratamento. Após 140 dias, foram avaliados o número de estacas com brotações, número de brotações, número de folhas e diâmetro da raiz principal. Estacas de raízes de progênies de dois anos de idade apresentaram melhor desenvolvimento. Menores concentrações de citocinina trouxeram melhores resultados de emissão e número de brotações das progênies de ambas as idades.

Main nutrients and functional ingredients in shoots of Polygonatum cyrtonema / 中国中药杂志

In order to reveal the main nutrients and functional ingredients in the shoots of Polygonatum cyrtonema, the polysaccharides, proteins, amino acids, and total phenols were determined. The tested samples cultured in Ma'nijiaonong, Hengtang village, Tianmushan town, Lin'an, Zhejiang, which were collected from three provenances(Pan'an and Longquan in Zhejiang and Qingyang in Anhui). The results showed that the polysaccharide content of the shoots varied from 2.34% to 12.73%, roughly one-third of rhizomes. The protein content varied from 107.75 to 192.49 mg·g~(-1), nearly 5.50 times more than rhizomes. Moreover, the average of total amino acid content was 193.13-248.74 mg·g~(-1), approximately 4.16 times of rhizomes. And the essential amino acids account for 35.57%-39.44% of the total amino acids content, which was close to the standard of the ideal protein proposed by FAO/WHO(the essential amino acid/total amino acid is about 40%). In addition, the taste amino acids(TaAA) changed from 160.12 to 208.29 mg·g~(-1), revealing the material basis of "shoots were extremely delicious" in Chinese ancient herbal medicine. Additionally, the total phenols varied from 51.21-58.76 mg·g~(-1), about 2.96 times of rhizomes. The DPPH free radical scavenging rate of tested shoots was over 95%, which obviously superior to rhizomes. Therefore, the shoots of P. cyrtonema is a very high-quality vegetable and functional food with good development potential. Furthermore, the main nutrients and functional substances in P. cyrtonema shoots are closely related to the provenances and harvesting seasons. It is important to improve the quality and yield of the shoots by strengthening the variety of breeding and cultivation techniques.

Clonal Propagation of Strawberry (Fragaria x ananassa Duch.) through In vitro Runner Tip Culture through Incorporation of Growth Hormones

Runner tips explants of strawberry give rise to multiple shoots when cultured on MS medium supplemented with different concentrations and combinations of BAP with KIN or NAA or GA3.The highest response of shoot multiplication was obtained on MS containing 2.5 mgl-1 BAP + 0.5 mgl-1 Kin + 0.5 mgl-1 GA3. The maximum frequency of rooting (83%) and highest number of roots (3.49) was produced in medium containing 1.0 mgl-1 IBA. The well grown rooted plantlets were acclimatized and successfully established in autoclaved vermiculate soil and as well as natural condition. Using our established protocol, it is also possible to provide large numbers of micropropagated plantlets of this cultivars to produce high quality strawberry fruit for commercial cultivation practices.

De novo in vitro shoot morphogenesis from shoot tip-induced callus cultures of Gymnema sylvestre (Retz.) R. Br. ex Sm

BACKGROUND: Gymnema sylvestre is a medicinal woody perennial vine known for its sweetening properties and antidiabetic therapeutic uses in the modern and traditional medicines. Its over-exploitation for the therapeutic uses and to meet the demand of pharmaceutical industry in raw materials supply for the production of anti-diabetic drugs has led to considerable decline in its natural population. RESULTS: An efficient system of shoot bud sprouting from nodal segment explants and indirect plant regeneration from apical meristem-induced callus cultures of G. sylvestre have been developed on Murashige and Skoog (MS) medium amended with concentrations of cytokinins. Of the three growth regulators tested, N6-benzylaminopurine (BAP) was the most efficient and 2.0 mg L-1 gave the best shoot formation efficiency. This was followed by thidiazuron (TDZ) and kinetin (Kin) but, most of the TDZ-induced micro shoots showed stunted growth. Multiple shoot formation was observed on medium amended with BAP or TDZ at higher concentrations. The produced micro shoots were rooted on half strength MS medium amended with auxins and rooted plantlets acclimatized with 87% survival of the regenerates. CONCLUSIONS: The developed regeneration system can be exploited for genetic transformation studies, particularly when aimed at producing its high yielding cell lines for the anti-diabetic phytochemicals. It also offers opportunities for exploring the expression of totipotency in the anti-diabetic perennial vine.

Development of a direct transformation method by GFP screening and in vitro whole plant regeneration of Capsicum frutescens L

Background: Capsicum is a genus of an important spice crop that belongs to the chili lineage. However, many Capsicum species (family Solanaceae) are known to be recalcitrant to genetic transformation and in vitro regeneration, thus hampering the effort in using Capsicum species for detailed biological investigation. In this study, we have developed an optimized protocol for the direct transformation of Capsicum frutescens L. cv. Hot Lava using a biolistic particle delivery system. In addition, a procedure for in vitro whole plant regeneration from the hypocotyl explants of C. frutescens was established. Results: In this study on the biolistic system, explant target distance, bombardment helium (He) pressure, and the size of the microcarrier were the key parameters to be investigated. The optimized parameters based on the screening of GFP expression were determined to have a target distance of 6 cm, helium pressure of 1350 psi, and gold particle (microcarrier) size of 1.6 µm. The greatest number of shoots was obtained from hypocotyls as explants using Murashige and Skoog medium supplemented with 5.0-mg/L 6-benzylaminopurine and 0.1-mg/L 1-naphthaleneacetic acid. On an average, five shoots per explant were formed, and of them, one shoot managed to form the root and developed into a whole plant. Conclusions: We obtained an optimized protocol for the biolistic transformation of chili and in vitro regeneration of chili plantlets. The establishment of the protocols will provide a platform for molecular breeding and biological studies of chili plants.

Simultaneous determination of eight flavonol glycosides in Toona sinensis shoots by UPLC / 中成药

AIM To establish a UPLC method for the simultaneous content determination of glycosides,rutinoside,myricitrin,hyperoside,isoquercitrin,guaijaverin,astragalin,quercitrin and afzelin in the shoots of Toona sinensis Roemer.METHODS The analysis of 60％ methanol extract of T.sinensis shoots was performed on a 35 ℃ thermostatic Waters XBridge Shield RP18 column (2.1 mm × 150 mm,1.7 μm),with the mobile phase comprising of acetonitrile-water flowing at 0.35 mL/min in a gradient elution manner,and the detection wavelength was set at 350 nm.RESULTS Eight flavonol glycosides showed good linear relationships within their own ranges (r≥ 0.9992),whose average recoveries were 98.3％-103.5％ with the RSDs of 0.46％-3.36％.CONCLUSION This accurate,stable and reproducible method can be used for the quality control of T.sinensis shoots.

Simultaneous determination of eight flavonol glycosides in Toona sinensis shoots by UPLC / 中成药

AIM To establish a UPLC method for the simultaneous content determination of glycosides,rutinoside,myricitrin,hyperoside,isoquercitrin,guaijaverin,astragalin,quercitrin and afzelin in the shoots of Toona sinensis Roemer.METHODS The analysis of 60％ methanol extract of T.sinensis shoots was performed on a 35 ℃ thermostatic Waters XBridge Shield RP18 column (2.1 mm × 150 mm,1.7 μm),with the mobile phase comprising of acetonitrile-water flowing at 0.35 mL/min in a gradient elution manner,and the detection wavelength was set at 350 nm.RESULTS Eight flavonol glycosides showed good linear relationships within their own ranges (r≥ 0.9992),whose average recoveries were 98.3％-103.5％ with the RSDs of 0.46％-3.36％.CONCLUSION This accurate,stable and reproducible method can be used for the quality control of T.sinensis shoots.

Comparison on progeny regeneration capacity among different mating patterns and optimization of cultivation condition in Swertia mileensis / 中草药

Objective: To establish and optimize the technique of tissue culture and rapid propagation, and compare the progeny regeneration capacity among different mating patterns in Swertia mileensis, meanwhile, to explore the reproductive assurance of the fragment populations. Methods: Sterilized seeds from different mating patterns were cultured on MS media for germination, which included self-breed, inbreeding, outbreeding, and nature pollinating, and counted the germination rate 60 d later. The study used the different source explants maintained on MS medium supplemented with different types and concentration of plant growth regulator combinations to explore its capacity of callus induction and adventitious shoots differentiation with orthogonal design. Results: There is no significant difference in the germination rates among all kinds of seed as touched above. The optimal medium for callus induction is MS + ZT 0.5 mg/L + 2,4-D 0.05 mg/L, the callus induction rates were 100%, 96.67%, 96.55%, and 96.29%; The optimal medium for adventitious shoots differentiation was MS + BA 2.0 mg/L + KT 0.1 mg/L + NAA 1.0 mg/L, and the differentiation rates all reached up to 100%; The optimal medium for rooting was 1/2 MS + NAA 1.0 mg/L, the rooting rate both were as high as 100% and there was no significant difference in the growth of plantlets. Conclusion: This study optimizes the cultivation conditions and provides an effective solution for protecting the wild resources and sprouts multiplication of S. mileensis. Meanwhile, from the plant physiological basis, it could be preliminarily predicted that the advantage of inbreeding would make up the fitness cost associated with it and the progeny regeneration capacity shows no significant difference between inbreeding and outbreeding.

A simple cost effective method for mass propagation of Chrysanthemum morifolium and antibacterial activity assessment of in vitro raised plantlets.

An efficient low cost procedure for in vitro propagation of Chrysanthemum morifolium has been developed with subsequent assessment of antibacterial property of in vitro raised plantlets. Optimal axillary shoot multiplication was achieved on MS medium supplemented with low concentration of BAP. Psyillium husk and market sugar were standardized as suitable alternatives to the conventionally used agar and sucrose, cutting down the production cost of tissue culture raised plantlets to over six times. Optimal in vitro rooting was obtained on half strength MS medium containing IBA. Regenerated plantlets with well developed shoots and roots were acclimatized successfully and transferred to field conditions where they flowered. The leaves of ex vitro growing tissue culture raised plantlets were later assessed for activity against bacterial pathogens. The present protocol ensures minimal cost input in large scale production of a commercially important ornamental plant and opens up scope of scientific interventions directed at its allied therapeutic usage. Abbreviations: MS: Murashige and Skoog (1962); HgCl2 : Mercuric chloride; PGR: Plant growth regulator, TCR: tissue culture raised; BAP: 6, Benzylaminopurine; NAA: α-Naphthalene Acetic Acid; IBA: Indole-3 butyric acid; IAA: Indole-3 acetic acid; min: minutes; ***:significant at 99.9%.

Direct multiple shoots proliferation of black night shade (Solanum nigrum l.) from shoot tip explants induced by thidiazuron .

The present investigation was undertaken in Black Night Shade (Solanum nigrum L.) which is an important medicinal plant. Direct multiple shoots proliferation was achieved from shoot tip. The shoot tips were cultured on MS medium fortified with Thidiazuron (TDZ) (1.0-7.0 mg/L) for multiple shoot induction. Multiple shoots proliferation was best observed at 3.0 mg/L TDZ from the shoot tip explants within three weeks of culture. Shoot number per explant ranged between 2 and 10. Individual shoots were aseptically excised and sub cultured in the same media for shoot elongation. The elongated shoots were transferred to Indole Acetic Acid/Indole Butyric Acid (IAA/IBA) (0.5mg/L–2.0mg/L) for root induction. Rooting was observed within two weeks of culture. Rooted plantlets were successfully hardened under culture conditions and subsequently established in the field conditions. The recorded survival rate of the plants was 86%. Plants looked healthy with no visually detectable phenotypic variations.

Efficient induction system of adventitious shoots and plant regeneration in Solanum pseudo-capsicum / 中草药

Objective: To investigate the conditions for improving the efficiency of plant regeneration system for Solanum pseudo-capsicum via the induction of adventitious shoots. Methods: Sterilized seeds of S. pseudo-capsicum were cultured on MS, OM, and WPM basal media for germination and seedling growth. Effectiveness of different combination of tissue culture media and growth regulators on the callus induction, adventitious shoots production, and plant regeneration were compared, using the explants excised from the seedlings of 45 d. Results: WPM is the best medium for the growth of S. pseudo-capsicum. Leaf is the best explant for callus induction with the frequency up to 100% cultured on WPM medium containing 0.5 mg/L 6-BA and 0.01 mg/L NAA after 7 d and the callus divided into green adventitious shoots whose frequency reached 100% after 10 d; The callus whose frequency was 81.7% could be induced from the stem with buds cultured on the same medium as mentioned above, the adventitious shoots generated 20 d later. However, the stem tip induced callus without capacity of differentiation. The optimal medium for adventitious shoots proliferation is WPM + 0.5 mg/L 6-BA + 0.5 mg/L NAA + 1.0 mg/L KT, the proliferation coefficient is far more 6.0 after 30 d. The optimal medium for plantlet rooting is 1/2 WPM + 0.01 mg/L NAA, the strong plant regeneration occurred 30 d later with the rooting rate up to 100%. Over 90% plantlets survived after transplanting into sand with constant temperature and humidity condition for 25 d. Conclusion: The current study provides an effective solution for maintaining the improved seeds trait and sprouts multiplication of S. pseudo-capsicum, which is helpful for the research of cultivation and genetic transformation.

Chemical constituents from shoots of Phyllostachys edulis (I) / 中草药

Objective: To investigate the chemical constituents from the shoots of Phyllostachys edulis. Methods: Compounds were isolated by using a combination of various chromatographic techniques including silica gel, ODS, Sephadex LH-20, reversed-phase HPLC, and others. And the structures were elucidated by the nuclear magnetic resonance (NMR), mass spectrometry, and other modern spectroscopy. Results: Eighteen compounds were isolated from the shoots of P. edulis and identified as thymidine (1), uracil (2), 5-ethyluracil (3), 4-hydroxy-3-methoxybenzoic acid (4), p-hydroxy-benzaldehyde (5), adenosine (6), uridine (7), 2'-O-methyluridine (8), ethyl-p-hydroxybenzoate (9), cyclo (L-Val-L-Ala) (10), cyclo (L-Phe-L-Leu) (11), flazine (12), ethyl 4-(sulfooxy) benzoate (13), 1H-indole-3-carboxylic acid, methyl ester (14), cis-p-hydroxyl ethyl cinnamate (15), trans-p-hydroxyl ethyl cinnamate (16), 4-hydroxy-2-methoxy-benzaldehyde (17), and methyl p-hydroxy benzeneacetate (18). Conclusion: All the compounds are isolated from the shoots of P. edulis for the first time, and compounds 6-18 are firstly obtained from the plants of Phyllostachys Sieb. et Zucc.

In vitro regeneration and ploidy level analysis of Eulophia ochreata Lindl.

Various parameters including explant-type, medium compositions, use of phytohormones and additives were optimized for direct and indirect regeneration of E. ochreata, a medicinal orchid under threat. Protocorm-like-bodies (PLBs) proved to be the best explants for shoot initiation, proliferation and callus induction. Murashige and Skoog’s (MS) medium containing 2.5 mg L-1 6-benzylaminopurine (BAP), 1.0 mg L-1 kinetin (Kin) and additives (adenine sulfate, arginine, citric acid, 30 mg L-1 each and 50 mg L-1 ascorbic acid) was optimal for shoot multiplication (12.1 shoots and 7.1 PLBs per explant with synchronized growth), which also produced callus. Shoot number was further increased with three successive subcultures on same media and ~40 shoots per explant were achieved after 3 cycles of 30 days each. Additives and casein hydrolysate (CH) showed advantageous effects on indirect shoot regeneration via protocorm-derived callus. Optimum indirect regeneration was achieved on MS containing additives, 500 mg L-1 CH, 2.5 mg L-1 BAP and 1.0 mg L-1 Kin with 30 PLBs and 6 shoots per callus mass (~5 mm size). The shoots were rooted (70% frequency) on one by fourth-MS medium containing 2.0 mg L-1 indole-3-butyric acid, 200 mg L-1 activated charcoal and additives. The rooted plantlets were hardened and transferred to greenhouse with 63% survival rate. Flow-cytometry based DNA content analysis revealed that the ploidy levels were maintained in in vitro regenerated plants. This is the first report for in vitro plant regeneration in E. ochreata.

Large scale propagation of an exotic edible bamboo, Phyllostachys pubescens Mazel ex H. De Lehale (Moso Bamboo) using seeds.

For ex vitro propagation, seeds of P.pubescens were treated with different concentrations of gibberellic acid (GA3) and germination of seeds was tested both in plastic pots as well as by direct sowing in the nursery beds. Maximum seed germination was achieved when treated with 200 mgL–1 (w/v) GA3. For in vitro propagation, an exposure of nodal explants from in vitro raised seedlings to 0.2 mgL–1 1–phenyl–3–(1,2,3–thiadiazol–5–yl) urea and 1 mgL–1 kinetin supplemented medium for 30 days and thereafter to hormone free Murashige and Skoog basal medium resulted in axillary shoot proliferation. For rooting, in vitro raised shoots were exposed to MS medium containing 2 mgL–1 indole-3-butyric acid for 15 days and then shifted to hormone free medium. On an average, 2.8 shoots were obtained in 75% of the cultures within 4 weeks. Such in vitro raised plants were successfully hardened and shifted to field conditions.

In vitro propagation of Vriesea reitzii, a native epiphyte bromeliad from the Atlantic rainforest / Propagação in vitro de Vriesea reitzii, uma bromélia epífita nativa da Mata Atlântica

The induction of nodular culture (NC) and the subsequent development of microshoots of V. reitzii are considered an in vitro propagation model-system with high regenerative performance. Current research analyzed the determinant factors of the in vitro morphogenesis control of bromeliads. Seeds excised from mature capsules were grown on medium MS basic (MSB), liquid or gelled, supplemented or not with α- naphthaleneacetic acid (NAA), 6-benzilaminopurine (BAP) or thidiazuron (TDZ). The regeneration and elongation of microshoots were evaluated from NC sub-cultivated on MSB medium on liquid culture medium supplemented with different concentrations of indolyl-3-acetic acid (IAA) and gibberellic acid (GA3). Plant growth regulators (PGR) supplemented into the medium MSB inhibited the germination of the seeds and induced NC in the second week of growth. The induced NC on MSB medium with NAA (4 µM) and sub-cultivated on MSB medium with NAA (2 µM) plus N6(2-isopentenyl) adenine (2-iP) (2 µM) showed granular texture and high rate of proliferation. NC sub-culture in MSB medium with IAA (4 µM) provided a higher average number of microshoots (1,478 shoots g-1 of NC). Shoots over 3.0 cm resulted in more than 95% ex vitro survival.

A indução de culturas nodulares (CNs) e subsequente desenvolvimento de microbrotos de V. reitzii configuraram-se em um sistema de alta performance regenerativa in vitro. No presente trabalho foram estudados os fatores determinantes do controle da morfogênese in vitro das CNs. Sementes excisadas de cápsulas maduras foram cultivadas em meio básico MS (MSB) líquido ou geleificado e suplementado ou não com ácido naftalenoacético (ANA), 6-benzilaminopurina (BAP) ou thidiazuron (TDZ). A partir das CNs subcultivadas em meio MSB, foi avaliada a regeneração e o alongamento de microbrotos em meio de cultura líquido suplementado com diferentes concentrações de ácido indolil-3-acético (AIA) combinados com ácido giberélico (AG3). A suplementação de fitorreguladores ao meio MSB inibiram a germinação das sementes e promoveram a indução de CNs na segunda semana de cultivo. As CNs induzidas em meio MSB suplementado com ANA (4 µM) e subcultivadas em meio MSB suplementado com ANA (2 µM) mais N6(2-isopentenil) adenina (2-iP) (2 µM) apresentaram textura granular e alta taxa de proliferação. O cultivo destas CNs em meio MSB suplementado com AIA (4 µM) resultou no maior número médio de microbrotos (1.478 brotos g -1 de CN). Brotos maiores de 3,0 cm resultaram em mais de 95% de sobrevivência em ambiente ex vitro.

High-frequency in vitro plantlet regeneration from apical bud as a novel explant of Carum copticum L

Objective: To develop an in vitro regeneration system to increase the recovery of Carum copticum L. plantlets as a part of developing a metabolic engineering program.Methods:3-acetic acid and indole butyric acid on direct shoot regeneration and rooting of ajowan from apical bud explants were assessed. All explants were cultured on Murashige and Skoog (MS) medium supplemented with different combinations of 6-benzyl amino purine (BAP) (0, 2.2, 4.4, 8.8μ The efficacy of different concentrations and combinations of 6-benzyladenine, indole-Results: The maximum shoot regeneration frequency (97.5%) and the highest number of shoots produced from apical buds (34 shoots per explant) were obtained on MS medium fortified with BAP (4.4 μmol/L) and IAA (0.5 μmol/L). Low shoot regeneration frequency was observed in BAP free treatments. The effects of different strengths of MS medium and various concentrations of IAA and indole-3- butyric acid on rooting rate, length and average number of roots were also investigated. Application of indole-3- butyric acid (6 μmol/L) in full-strength MS medium, was more effective than IAA and resulted in highest shoot regeneration frequency with the rooting rate of 100% and highest mean number of roots per shoot (41.8). The rooted plantlets were acclimatized successfully in greenhouse conditions with a survival rate of 90%. mol/L) and indole-3-acetic acid (IAA) (0, 0.5, 1.1, 2.2 μmol/L). Conclusion: In this study, a simple and reliable regeneration and acclimatization protocol for Carum copticum has been presented. This protocol can be found very advantageous for a variety of purposes, including mass multiplication of Carum species, medicinal plant breeding studies and transgenic plant production.

High-frequency in vitro plantlet regeneration from apical bud as a novel explant of Carum copticum L

<p><b>OBJECTIVE</b>To develop an in vitro regeneration system to increase the recovery of Carum copticum L. plantlets as a part of developing a metabolic engineering program.</p><p><b>METHODS</b>The efficacy of different concentrations and combinations of 6-benzyladenine, indole-3-acetic acid and indole butyric acid on direct shoot regeneration and rooting of ajowan from apical bud explants were assessed. All explants were cultured on Murashige and Skoog (MS) medium supplemented with different combinations of 6-benzyl amino purine (BAP) (0, 2.2, 4.4, 8.8 µmol/L) and indole-3-acetic acid (IAA) (0, 0.5, 1.1, 2.2 µmol/L).</p><p><b>RESULTS</b>The maximum shoot regeneration frequency (97.5%) and the highest number of shoots produced from apical buds (34 shoots per explant) were obtained on MS medium fortified with BAP (4.4 µmol/L) and IAA (0.5 µmol/L). Low shoot regeneration frequency was observed in BAP free treatments. The effects of different strengths of MS medium and various concentrations of IAA and indole-3- butyric acid on rooting rate, length and average number of roots were also investigated. Application of indole-3- butyric acid (6 µmol/L) in full-strength MS medium, was more effective than IAA and resulted in highest shoot regeneration frequency with the rooting rate of 100% and highest mean number of roots per shoot (41.8). The rooted plantlets were acclimatized successfully in greenhouse conditions with a survival rate of 90%.</p><p><b>CONCLUSION</b>In this study, a simple and reliable regeneration and acclimatization protocol for Carum copticum has been presented. This protocol can be found very advantageous for a variety of purposes, including mass multiplication of Carum species, medicinal plant breeding studies and transgenic plant production.</p>

Regeneration of Shoots from Callus of Ceropegia pusilla Wight and ARN.

This study was undertaken to evaluate the most suitable concentration of plant growth regulators and suitable explants for callus induction and subsequent organogenesis of an endangered medicinal plant Ceropegia pusilla. The best performance of callus induction and morphogenesis was found on MS medium supplemented with 6- benzylaminopurine (BA) and naphthalene acetic acid (NAA), from node and internode the maximum account of callus initiation was recorded on MS medium supplemented with BA+NAA (2.22μM + 5.37 μM) and the maximum % of callus induced on subculture is 5.47  0.68. Rooting was best achieved on MS medium augmented with IBA (2.46μM). The maximum number of roots (4.72  0.66 cm) and root length (4.8  0.49 cm) were recorded). Plantlets regenerated in vitro with well-developed shoot and roots were successfully acclimatized in pots containing a mixture of decomposed coir waste, perlite and compost 1: 1:1 ratio and grown in a shade house with 81  3.16 percent survival rates.