Light Requirement for Shoot Regeneration in Horseradish Hairy Roots 1

Science.gov (United States)

Saitou, Tsutomu; Kamada, Hiroshi; Harada, Hiroshi

1992-01-01

Hairy roots of horseradish (Armoracia rusticana) were induced by inoculation with Agrobacterium rhizogenes harboring Ri plasmid and cultured on phytohormone-free Murashige and Skoog medium after eliminating the bacteria. Hairy roots grew vigorously and sometimes formed yellowish calli under dark conditions. On the other hand, growth of hairy roots stopped after several weeks of culture with light, then shoots were regenerated. Frequency of shoot formation from hairy roots increased as the culture period in light lengthened and the light intensity increased. The shoot regeneration was induced by treatment with white or red light, but not with far-red light. Shoot regeneration by red light was inhibited by following treatment with far-red light. Red and far-red light reversibly affected shoot regeneration. Excised roots of nontransformed plants grew quite slowly on phytohormone-free Murashige and Skoog medium and occasionally formed shoots under white light conditions. PMID:16669041

Optimization of Protocols for In Vitro Regeneration of Sugarcane (Saccharum officinarum

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Shakra Jamil

2017-01-01

Full Text Available Sugarcane contributes 60–70% of annual sugar production in the world. Somaclonal variation has potential to enhance genetic variation present within a species. Present study was done to optimize an in vitro propagation protocol for sugarcane. The experiments included four varieties, 9 callus induction media, 27 regeneration media, and 9 root induction media under two-factor factorial CRD. Data were recorded on callus induction, embryogenic callus formation, shoot elongation (cm, root induction, and plant regeneration. Statistically significant differences existed between genotypes and treatments for callus induction (%, embryogenic callus formation (%, shoot elongation (cm, root induction, and plant regeneration (%. All parameters showed dependency on genotypes, culture media, and their interaction. Highest callus induction (95% embryogenic callus formation (95% was observed in callus induction media 5. Highest plantlet regeneration (98.9% capacity was observed in regeneration media 11 whereas maximum shoot elongation (12.13 cm and root induction (8.32 were observed in rooting media 4. G1 showed best response for all traits and vice versa for G4. Hence it was concluded that G1, callus induction media 5, regeneration media 11, and rooting media 4 are the best conditions for in vitro propagation of sugarcane.

In vitro plant regeneration of Albizia lebbeck (L. from seed explants

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S. Perveen

2013-07-01

Full Text Available Objectives: An efficient and reproducible regeneration protocol for rapid multiplication of Albizia lebbeck (L. was developed by using intact seed explants.Methods: Murashige and Skoog's (MS medium supplemented with different hormones (BA, Kn, GA3 and TDZ was used for the induction of multiple shoots from the seed explants. Ex-vitro rooting was performed by using pulse treatment method in auxins (IBA and NAA and the complete plantlets were transferred to the field.Results: High frequency direct shoot induction was found in aseptic seed cultures of A. lebbeck on Murashige and Skoog medium supplemented with 5.0 µM TDZ (Thiadiazuron. Seeds were germinated after 7 days of culture and induced maximum 8 shoots from the region adjacent to the apex of the primary shoot of the seedling upto 25 days of incubation. Proliferating shoot cultures with increased shoot length was established by sub-culture of excised sprouting epicotyls on MS medium supplied with reduced concentrations of TDZ. Maximum shoot regeneration frequency (76 % with  highest number of shoots (21 and shoot length (5.1 cm per sprouting epicotyl was observed in the MS medium supplemented with 0.5 µM TDZ after 8 weeks of culture. Different concentrations of Indole-3-butyric acid (IBA and α-naphthalene acetic acid (NAA were tested to determine the optimal conditions for ex-vitro rooting of the microshoots. The best treatment for maximum ex-vitro root induction frequency (81 % was accomplished with IBA (250 µM pulse treatment given to the basal end of the microshoots for 30 min followed by their transfer in plastic cups containing soilrite and eventually established in normal garden soil + soilrite (1:1 with 78 % survival rate. In addition, histological study was undertaken to gain a better understanding of the regenerated shoots from the epicotyl region.Conclusion: The findings will be fruitful in getting a time saving and cost effective protocol for the in vitro propagation of Albizia

In vitro plant regeneration from leaf explants of Solanum pimpinellifolium L.

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Shirley Valderrama-Alfaro

2011-01-01

Full Text Available Tomato is a species of agricultural importance. Besides, it is a source of vitamins, minerals and antioxidant compounds. Therefore it is essential to obtain varieties resistant to diseases. Solanum pimpinellifolium L. ‘Tomatillo wild’, one of the wild relatives of tomato, is considered multipurpose reservoir of genes. This characteristic must be exploited. In vitro plantlets from leaves of ‘wild tomatillo’ were regenerated with this objective. A system for in vitro germination of seeds obtained from ex vitro sources was created. The use of seed allowed obtaining adequate seedlings in a short time to start regeneration. Thein vitro response of explants was evaluated in four treatments. The Murashige and Skoog (MS basal culture medium supplemented with a-naphthaleneacetic acid (NAA and 6-benzyl aminopurine (BAP was used in different combinations. The higher percentage (30% of shoot induction was achieved with 0.1 mg l1 NAA / 1 mg l1 BAP. The presence of callus and roots was observed after seven days of culture in the combination treatment with 1 mg l1 NAA / 0.1 mg l1 BAP. Callus showed 1 or 2 adventitious shoots per explant after 30 days of culture and 3 to 6 shoots after seventy days. Furthermore, the presence of fully formed adventitious plantlets (shoot and root was observed after the fifth week of culture. Therefore, the best combination for in vitro regeneration is NAA 0.1 mg l1 / 1 mg l1 BAP. Keywords: auxin, cytokinins, indirect organogenesis

Micropropagation of chokeberry by in vitro axillary shoot proliferation.

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Litwińczuk, Wojciech

2013-01-01

The black chokeberry-aronia (Aronia melanocarpa Elliot) is a shrub native to North America although nowadays well known in Eastern Europe. The fruits are regarded as the richest source of antioxidant phytonutrients among fruit crops and vegetables. Chokeberries can be easily propagated by seeds but this method is not recommended. Micropropagation is far more efficient than other conventional cloning methods like layering or softwood cuttings. Aronia clones are propagated in vitro through four- or three-stage method based on subculturing of shoot explants. The double diluted MS or full strength MS medium with elevated 50% Ca(2+) and Mg(2+) content are used in the initiation and proliferation chokeberry in vitro cultures, respectively. They are supplemented with 0.5-1.0 mg LBA, and 0.05 mg LIBA. The double-phase medium is recommended in the last passage before shoot rooting. The regenerated shoots could be rooted both in vitro on double diluted MS with 0.05 mg L(-1) IBA or in vivo in peat and perlite substrate and subsequently grown in the greenhouse.

Regeneration of Pelargonium in vitro

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Agnieszka Wojtania

2013-12-01

Full Text Available Pelargonium sp. has been a subject of numerous studies to deterimine the effec tiveness of in vitro techniques to produce a large number of pathogen-free plants. Regeneration of pelargonium plants from the different initial explants as well via organogenesis as via somatic embryogenesis has been obtained. The most effective adventitious shoot formation has been achieved from shoot tips and axillary buds using cytokinin or cytokinin/auxin combinations. Leaf explants, whose general have lower organogenic potency, regenerate better in the presence of thidiazuron. This growth regulator stimulate the somatic embryos production from hypocotyl and cotyledone explants too. The main problem in tissue culture propagation of Pelargonium has been the high tendency to formation of vigorously growing callus with low organogenic potency and rapid senescence of cultures. Moreover, the significant differen ces in requirements to the medium composition (minerals, organic compounds and growth regulators between Pelargonium cultivars has been observed. This makes difficult to develop an universaI method of Pelargonium micropropagation.

Developmental anatomy of blueberry (Vaccinium corymbosum L. ‘Aurora’) shoot regeneration

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The culture of Vaccinium corymbosum L. ’Aurora’ leaves on regeneration medium results in the regeneration of adventitious shoots. We present anatomical evidence that these new shoot apices are directly regenerated from the cultured blades. Mounds of densely staining cells, which formed from epidermi...

Auxin-cytokinin synergism in vitro for producing genetically stable plants of Ruta graveolens using shoot tip meristems

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Mohammad Faisal

2018-02-01

Full Text Available An efficient micropropagation protocol was developed for Ruta graveolens Linn. using shoot tip meristems derived from a 4-month-old field grown plant. In vitro shoot regeneration and proliferation was accomplished on Murashige and Skoogs (MS semi-solid medium in addition to different doses of cytokinins viz.6- benzyl adenine (BA, Kinetin (Kn or 2-isopetynyl adenine (2iP, singly or in combination with auxins viz. indole-3-acetic acid (IAA, indole-3-butyric acid (IBA or α-naphthalene acetic acid (NAA. Highest regeneration frequency (27.6% was obtained on (MS medium composed of BA (10 µM with maximum number (9.4 of shoots and 4.3 cm shoot length after 4 weeks of incubation. Among various combinations tried best regeneration frequency (71% of multiple shoot formation with highest number (12.6 of shoots per shoot tip explants were achieved in MS medium augmented with a combination BA (10.0 µM and NAA (2.5 µM after 4 weeks of incubation. The optimum frequency (97% of rhizogenesis was achieved on half-strength MS medium having 0.5 µM IBA after 4 weeks of incubation. Tissue culture raised plantlets with 5–7 fully opened leaves with healthy root system were successfully acclimatized off in Soilrite™ with 80% survival rate followed by transportation to normal soil under natural light. Genetic stability among in vitro raised progeny was evaluated by ISSR and RAPD markers. The entire banding pattern revealed from in vitro regenerated plants was monomorphic to the donor. The present protocol provides an alternative option for commercial propagation and fruitful setting up of genetically uniform progeny for sustainable utilization and germplasm preservation.

A rapid and efficient in vitro regeneration system for lettuce (Lactuca sativa L.).

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Armas, Isabel; Pogrebnyak, Natalia; Raskin, Ilya

2017-01-01

Successful biotechnological improvement of crop plants requires a reliable and efficient in vitro regeneration system. Lettuce ( Lactuca sativa L.), one the most important vegetable crops worldwide, is strongly genotype-dependent in terms of regeneration capacity, limiting the potential for biotechnological improvement of cultivars which show recalcitrance under currently available protocols. The effect of different nutrient sources, plant hormone combinations and activated charcoal supplementation on shoot induction efficiency was evaluated on the cultivar 'RSL NFR', which had previously shown poor regeneration efficiency. Multiple shoot organogenesis from cotyledon explants was recorded at the highest frequency and speed on Murashige and Skoog regeneration medium supplemented with 200 mg/l of activated charcoal, 3% sucrose, 10 mg/l benzylaminopurine and 0.5 mg/l naphthaleneacetic acid, which induced shoots through direct regeneration in 90.8 ± 7.9% of explants. High shoot induction efficiency was also observed, albeit not quantified, when using this medium on some other cultivars. This activated charcoal-containing regeneration medium might offer a rapid and efficient option for direct shoot induction in some lettuce genotypes that do not respond well to common lettuce regeneration protocols. This is also the first report of the effect of activated charcoal in lettuce tissue culture.

Metabolomic homeostasis shifts after callus formation and shoot regeneration in tomato

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Kumari, Alka; Ray, Kamalika; Sadhna, Sadhna; Pandey, Arun Kumar; Sreelakshmi, Yellamaraju; Sharma, Rameshwar

2017-01-01

Plants can regenerate from a variety of tissues on culturing in appropriate media. However, the metabolic shifts involved in callus formation and shoot regeneration are largely unknown. The metabolic profiles of callus generated from tomato (Solanum lycopersicum) cotyledons and that of shoot regenerated from callus were compared with the pct1-2 mutant that exhibits enhanced polar auxin transport and the shr mutant that exhibits elevated nitric oxide levels. The transformation from cotyledon to callus involved a major shift in metabolite profiles with denser metabolic networks in the callus. In contrast, the transformation from callus to shoot involved minor changes in the networks. The metabolic networks in pct1-2 and shr mutants were distinct from wild type and were rewired with shifts in endogenous hormones and metabolite interactions. The callus formation was accompanied by a reduction in the levels of metabolites involved in cell wall lignification and cellular immunity. On the contrary, the levels of monoamines were upregulated in the callus and regenerated shoot. The callus formation and shoot regeneration were accompanied by an increase in salicylic acid in wild type and mutants. The transformation to the callus and also to the shoot downregulated LST8 and upregulated TOR transcript levels indicating a putative linkage between metabolic shift and TOR signalling pathway. The network analysis indicates that shift in metabolite profiles during callus formation and shoot regeneration is governed by a complex interaction between metabolites and endogenous hormones. PMID:28481937

Regeneration of okra ( Abelmoschus esculentus L.) via apical shoot ...

African Journals Online (AJOL)

Abelmoschus esculentus L. Monech) via apical shoot culture system. The study of apical shoot culture system was found effective for regeneration of apical shoots. The okra (A. esculentus L. Monech) N-550 line evolved at R&D, Nirmal Seeds Pvt. Ltd., ...

In vitro propagation of Cymbidium goeringii Reichenbach fil. through direct adventitious shoot regeneration.

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Park, Han Yong; Kang, Kyung Won; Kim, Doo Hwan; Sivanesan, Iyyakkannu

2018-03-01

The influence of 2,4-dichlorophenoxyacetic acid (2,4-D), benzyladenine (BA), and thidiazuron (TDZ) on direct rhizome induction and shoot formation from rhizome explants of Cymbidium goeringii was explored. Rhizome segments obtained from in vitro seed cultures of C. goeringii were placed on Murashige and Skoog (MS) medium incorporated with 5, 10, 20, or 40 µM 2,4-D and 1, 2, 4, or 8 µM BA or TDZ alone or in combination with 20 µM 2,4-D. The explants developed only rhizomes on MS medium with or without 2,4-D. The highest percent of rhizome formation (100%) was obtained on MS medium incorporated with 20 μM of 2,4-D. The morphology and number of rhizomes varied with the level of 2,4-D in the medium. Direct adventitious shoot formation was achieved on medium incorporated with BA or TDZ. The adventitious shoots produced per explant significantly increased with the supplementation of 2,4-D to cytokinin-containing medium. The highest mean of 21.8 ± 1.8 shoot buds per rhizome segment was obtained in medium fortified with 20 μM 2,4-D and 2 μM TDZ. The greatest percent of root induction (100%) and the mean of 5.3 ± 1.1 roots per shoot were achieved on ½ MS medium incorporated with 2 μM of α-naphthaleneacetic acid. About 97% of the in vitro-produced plantlets acclimatized in the greenhouse. An efficient in vitro propagation protocol was thus developed for C. goeringii using rhizome explants.

Effects of thidiazuron and paclobutrazol on regeneration potential of tulip flower stalk explants in vitro and subsequent shoot multiplication

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Małgorzata Podwyszyńska

2011-01-01

Full Text Available The effects of TDZ and paclobutrazol on the primary regeneration on tulip flower stalk explants of six cultivars and subsequent shoot multiplication were examined. Explants, flower stalk slices, were excised from cooled and subsequently forced bulbs. The explants were incubated for two months in darkness on medium containing NAA and cytokinins, 2iP and BAP, as control, or TDZ (0.5-4 mg l-1 and paclobutrazol (0.05-0.4 mg l-1. Then, the regenerating explants were subcultured on medium with TDZ and NAA applied at low concentrations. Different regeneration capabilities were found depending on cultivar and growth regulators. The percentage of explants forming leaf-like structures ranged, on the control medium, from 80% in 'Blue Parrot' and 'Prominence' to below 30% in 'Apeldoorn' and 'Mirjoran'. TDZ, applied at optimum for each cultivar concentration, greatly increased the regeneration potential up to 70-100%. Paclobutrazol, added to the TDZ-containing medium, significantly enhanced the response of explants, resulting in high numbers of leaf-like structures formed per explant (13.7-22.8. The structures developed gradually into characteristic forms: the growing up cotyledonary leaf, the probable root primordium formed at its base, the growing downwards stolon and the shoot meristem developed finely on its tip. It is suggested that such primary regeneration may have a nature of somatic embryogenesis. Then, the adventitious shoots developed and formed clusters, which were divided into 2-3 smaller ones every two months. The growth regulators, used at initial stage, markedly influenced subsequent shoot multiplication. Thus, the most intensive shoot formation was noted with TDZ at concentrations of 0.5-2 mg l-1 and paclobutrazol of 0.05-0.1 mg l-1.

A new approach for in vitro regeneration of tomato plants devoid of exogenous plant growth hormones.

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Plana, Dagmara; Fuentes, Alejandro; Alvarez, Marta; Lara, Regla M; Alvarez, Félix; Pujol, Merardo

2006-10-01

Many available methodologies for in vitro regeneration of commercial tomato varieties promote not only the production of normal shoots but also individual leaves, shoots without apical meristems and vitrified structures. All these abnormal formations influence and diminish the regeneration efficiency. At the basis of this phenomenon lies callus development. We optimized an alternative procedure by which the regeneration occurs without abnormal shoot formation. The portion including the proximal part of hypocotyls and the radicle was cultured on medium consisting of Murashige and Skoog salts, 4 mg/L thiamine, 100 mg/L mio-inositol and 3% sucrose. After two-three weeks, 60% explants showed adventitious shoot formation. No changes in the morphological characteristics of regenerated plants and fruits were observed as compared with parents. Karyotypic analysis of regenerated plants showed no variations in chromosome number. The optimized procedure offers the advantage of tomato plant regeneration avoiding callus formation, which enables normal plant recovery with an efficiency ranging from 1.45 +/- 0.05 to 2.57 +/- 0.06 shoots per explant in Campbell-28, Amalia, Lignon, and Floradel cultivars.

In vitro plant regeneration from Narbon Vetch (Vicia narbonensis L ...

African Journals Online (AJOL)

STORAGESEVER

2008-07-18

Jul 18, 2008 ... regeneration from cotyledonary node explants obtained from, in vitro raised seeds of 4 - 5 and 14 - 15 days old .... shoots in Vicia faba using different concentrations of. BAP-TDZ. ... feed legume crop for dry areas in west Asia.

In-vitro morphogenesis of corn (Zea mays L.) : I. Differentiation of multiple shoot clumps and somatic embryos from shoot tips.

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Zhong, H; Srinivasan, C; Sticklen, M B

1992-07-01

In-vitro methods have been developed to regenerate clumps of multiple shoots and somatic embryos at high frequency from shoot tips of aseptically-grown seedlings as well as from shoot apices of precociously-germinated immature zygotic embryos of corn (Zea mays L.). About 500 shoots were produced from a shoot tip after eight weeks of culture (primary culture and one subculture of four weeks) in darkness on Murashige and Skoog basal medium (MS) supplemented with 500 mg/L casein hydrolysate (CH) and 9 μM N(6)-benzyladenine (BA). In this medium, shoots formed in shoot tips as tightly packed "multiple shoot clumps" (MSC), which were composed of some axillary shoots and many adventitious shoots. When the shoot tips were cultured on MS medium containing 500 mg/L CH, 9 μM BA and 2.25 μM 2,4-dichlorophenoxyacetic acid (2,4-D), most of the shoots in the clumps were adventitious in origin. Similar shoot tips cultured on MS medium containing 500 mg/L CH, 4.5 μM BA and 2.25 μM 2,4-D regenerated many somatic embryos within eight weeks of culture. Somatic embryos were produced either directly from the shoot apical meristems or from calli derived from the shoots apices. Both the MSC and the embryos produced normal shoots on MS medium containing 2.25 μM BA and 1.8 μM indole-3-butyric acid (IBA). These shoots were rooted on MS medium containing 3.6 μM IBA, and fertile corn plants were grown in the greenhouse. The sweet-corn genotype, Honey N Pearl, was used for the experiments described above, but shoot-tip cultures from all of 19 other corn genotypes tested also formed MSC on MS medium containing 500 mg/L CH and 9 μM BA.

In vtro adventitious shoot regeneration from cotyledon explant of brassica oleracea subsp. Italica and brassica oleracea subsp. capitata using tdz and naa

International Nuclear Information System (INIS)

Salim, S.; Rashid, A.

2014-01-01

Broccoli(Brassica oleracea subsp. italica) cv. Green Dragon King and cabbage (Brassica oleracea subsp. capitata) cv. Gianty are important vegetable crops grown in Cameron Highlands, Malaysia. The cotyledons of both cultivars were used as explant source for in vitro shoot regeneration. The objective of this research was to examine the influence of the growth regulators thidiazuron (TDZ) and naphthaleneacetic acid (NAA) on adventitious shoot formation in these cultivars. This system of adventitious shoot regeneration from cotyledon explants could be useful as a tool for genetic transformation of the subspecies. Cotyledon explants of both cultivars excised from 5-day-old in vitro germinated seedlings were placed on shoot induction medium containing basal salts of Murashige and Skoog (MS) and various concentrations of TDZ and NAA. The highest percentage of cotyledon explant of broccoli cv. Green Dragon King producing shoot (76.66%) and the highest mean number of shoots produced per explant (0.9) were obtained on 0.1 mg/l TDZ with 0.1 mg/l NAA. Meanwhile, the highest percentage of cotyledon explant of cabbage cv. Gianty producing shoots (86.67%) and highest number of shoots produced per explant (1.1) were recorded on 0.5 mg/l TDZ with 0.1 mg/l NAA. Therefore, 0.1 mg/l TDZ with 0.1 mg/l NAA and 0.5 mg/l TDZ with 0.1mg/l NAA are the recommended combinations for adventitious shoot regeneration from cotyledonary explants of broccoli cv. Dragon King and cabbage cv. Gianty respectively. (author)

Induction of shoot regeneration in cotyledon explants of the oilseed crop Sesamum indicum L.

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Masochon Zimik

2017-12-01

Full Text Available Sesamum indicum is an ancient oilseed crop known for its high quality edible oil and its medicinally important lignans. The crop is said to be recalcitrant to plant tissue culture thus limiting the use of modern biotechnology for its genetic improvement. We present here a protocol describing plant regeneration through adventitious shoot formation from cotyledons dissected from sesame seeds soaked for four hours in water. Subculturing of the cotyledons after two weeks of culture on to a fresh Murashige and Skoog medium leads to differentiation of adventitious shoots from the proximal cut end of the explant. Culture of cotyledons on a medium containing 9% sucrose for a couple of weeks prior to transfer to MS medium supplemented with 3% sucrose induced a higher frequency of shoot regeneration. The highest frequency of 25% adventitious shoot regeneration was observed for S. indicum variety UMA. This variety also turned out to be the best among the ten genotypes tested for shoot regeneration through tissue culture. While addition of IAA marginally improved regeneration, silver nitrate was found essential for enhancing the frequency of shoot regeneration. The regenerated shoots formed roots on full strength MS medium supplemented with 1 mg/l IBA and the rooted plants were established in soil.

Morphological Evaluation of Shoots Regenerated from Hygromycin-Resistant Rice Callus (cv IACuba-28

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Maylin Pérez Bernal

2007-01-01

Full Text Available An evaluation system based on the morphological characteristics of regenerated hygromycin-resistant rice callus shoots was established for correlating such characteristics with shoot viability on hygromycin. Embryogenic rice calli were transformed by Agrobacterium tumefaciens (EHA105/ pCAMBIA1300, containing the hygromycin-phosphotransferase gene as selection marker. After two weeks on selection medium, hygromycin-resistant calli were transferred to regeneration medium. Regenerated shoots were extracted every 5 days (over a 30-day period and classified into three classes according to their morphological structure: class I: vigorous shoot having typical bipolar structure; class II: shoot having small root compared to apical length, or shoot without roots; class III: shoots having an abnormal appearance, such as malformed leaves or albinism. Individualised shoots were transferred to MS medium containing hygromycin for evaluating their resistance to antibiotics. A relationship was observed between regenerated shoots’ morphological characteristics and the percentage of shoots’ viability on hygromycin. Class I prevailed at early shoot extraction and was the most resistant to hygromycin. Drastic class I reduction was found with later shoot extraction, whilst classes II and III became increased. Likewise, shoot viability became radically reduced on MS medium containing hygromycin. This result might be applied for improving efficiency regarding obtaining transgenic rice plants, taking into account the best time for obtaining high percentages of hygromycin-resistant shoots having the best morphological characteristics.

In Vitro Regeneration of Foxtail Millet (Setaria italica (L. Beauv. cv. Buru Hotong

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Iriawati

2017-09-01

Full Text Available In vitro regeneration of foxtail millet (Setaria italica (L. Beauv. was done using basal shoot explants of 10-day old seedlings. Explants were cultured in MS basal medium supplemented with 2,4-dichlorophenoxyacetic acid (2,4-D, kinetin, 6-benzylaminopurine (BAP and 1.5 ppm NiSO4. Shoot multiplication and root induction were done in Murashige and Skoog (MS basal media. Plantlets were then acclimatized in rice husk charcoal, cocopeat, or mixed media. Results showed that MS basal medium containing 0.5 ppm kinetin, 2 ppm BAP, and 0.1 ppm 2,4-D was the optimal medium for shoot induction, in which 60% of explants developed direct shoot organogenesis. In addition, callus was optimally formed in MS medium supplemented by 1 ppm kinetin, 1 ppm BAP, and 0.5 ppm 2,4-D. Regenerated shoots spontaneously developed roots after being transferred into MS basal media without growth regulator. During acclimatization, the highest survival rate of plantlets (47% was obtained in rice husk charcoal. The developed method could be useful towards improvement of this species using in vitro tissue culture techniques.

IN VITRO REGENERATION OF LOURO-PARDO (Cordia trichotoma (Vellozo Arrabida ex Steudel

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Nilton César Mantovani

2010-08-01

Full Text Available This study was carried out aiming at evaluating the in vitro regeneration of louro-pardo (Cordia trichotoma (Vellozo Arrabida ex Steudel. Nodal segments were desinfested and innoculated in two basal media: WPM (Wood Plant Médium, 1981 and MS (Murashige & Skoog, 1962 supplemented with BAP (6-Benzylaminopurine, TDZ (Thidiazuron, NAA (Naphthaleneacetic Acid, GA3 (Gibberellic Acid, IBA (Indole-3-Butyric Acid and  A.C. (Activated Charcoal in different combinations and concentrations for each developmental step. The regeneration rate and the shoot elongation were higher in the WPM medium containing 0,1 mg.l-1 of BAP and GA3. The rooting of regenerated shoots was observed in IBA (0,5 mg.l-1 and Ativated Charcoal (1,5 g.l-1.

Transcriptomic and Hormonal Analyses Reveal that YUC-Mediated Auxin Biogenesis Is Involved in Shoot Regeneration from Rhizome in Cymbidium.

Science.gov (United States)

Liu, Yang; Zhang, Hai-Liang; Guo, He-Rong; Xie, Li; Zeng, Rui-Zhen; Zhang, Xiang-Qian; Zhang, Zhi-Sheng

2017-01-01

Cymbidium , one of the most important orchid genera in horticulture, can be classified into epiphytic and terrestrial species. Generally, epiphytic Cymbidium seedlings can be easily propagated by tissue culture, but terrestrial seedlings are difficult to propagate. To date, the molecular mechanisms underlying the differences in the ease with which terrestrial and epiphytic cymbidiums can be propagated are largely unknown. Using RNA-sequencing, quantitative reverse transcription PCR and enzyme-linked immunosorbent assay, Cymbidium 'Xiaofeng' (CXF), which can be efficiently micropropagated, and terrestrial Cymbidium sinense 'Qijianbaimo' (CSQ), which has a low regeneration ability, were used to explore the molecular mechanisms underlying the micropropagation ability of Cymbidium species. To this end, 447 million clean short reads were generated, and 31,264 annotated unigenes were obtained from 10 cDNA libraries. A total of 1,290 differentially expressed genes (DEGs) were identified between CXF and CSQ during shoot induction. Gene ontology (GO) enrichment analysis indicated that the DEGs were significantly enriched in auxin pathway-related GO terms. Further analysis demonstrated that YUC and GH3 family genes, which play crucial roles in the regulation of auxin/IAA (indole-3-acetic acid) metabolism, acted quickly in response to shoot induction culture in vitro and were closely correlated with variation in shoot regeneration between CXF and CSQ. In addition, the study showed that IAA accumulated rapidly and significantly during shoot induction in CXF compared to that in CSQ; in contrast, no significant changes in other hormones were observed between CXF and CSQ. Furthermore, shoot regeneration in CXF was inhibited by a yucasin-auxin biosynthesis inhibitor, indicating that increased IAA level is required for high-frequency shoot regeneration in CXF. In conclusion, our study revealed that YUC-mediated auxin biogenesis is involved in shoot regeneration from rhizome in

Transcriptomic and Hormonal Analyses Reveal that YUC-Mediated Auxin Biogenesis Is Involved in Shoot Regeneration from Rhizome in Cymbidium

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Yang Liu

2017-10-01

Full Text Available Cymbidium, one of the most important orchid genera in horticulture, can be classified into epiphytic and terrestrial species. Generally, epiphytic Cymbidium seedlings can be easily propagated by tissue culture, but terrestrial seedlings are difficult to propagate. To date, the molecular mechanisms underlying the differences in the ease with which terrestrial and epiphytic cymbidiums can be propagated are largely unknown. Using RNA-sequencing, quantitative reverse transcription PCR and enzyme-linked immunosorbent assay, Cymbidium ‘Xiaofeng’ (CXF, which can be efficiently micropropagated, and terrestrial Cymbidium sinense ‘Qijianbaimo’ (CSQ, which has a low regeneration ability, were used to explore the molecular mechanisms underlying the micropropagation ability of Cymbidium species. To this end, 447 million clean short reads were generated, and 31,264 annotated unigenes were obtained from 10 cDNA libraries. A total of 1,290 differentially expressed genes (DEGs were identified between CXF and CSQ during shoot induction. Gene ontology (GO enrichment analysis indicated that the DEGs were significantly enriched in auxin pathway-related GO terms. Further analysis demonstrated that YUC and GH3 family genes, which play crucial roles in the regulation of auxin/IAA (indole-3-acetic acid metabolism, acted quickly in response to shoot induction culture in vitro and were closely correlated with variation in shoot regeneration between CXF and CSQ. In addition, the study showed that IAA accumulated rapidly and significantly during shoot induction in CXF compared to that in CSQ; in contrast, no significant changes in other hormones were observed between CXF and CSQ. Furthermore, shoot regeneration in CXF was inhibited by a yucasin-auxin biosynthesis inhibitor, indicating that increased IAA level is required for high-frequency shoot regeneration in CXF. In conclusion, our study revealed that YUC-mediated auxin biogenesis is involved in shoot

[Identifying transcription factors involved in Arabidopsis adventious shoot regeneration by RNA-Seq technology].

Science.gov (United States)

Wang, Xingchun; Chen, Zhao; Fan, Juan; He, Miaomiao; Han, Yuanhuai; Yang, Zhirong

2015-04-01

Transcriptional regulation is one of the major regulations in plant adventious shoot regeneration, but the exact mechanism remains unclear. In our study, the RNA-seq technology based on the IlluminaHiSeq 2000 sequencing platform was used to identify differentially expressed transcription factor (TF) encoding genes during callus formation stage and adventious shoot regeneration stage between wild type and adventious shoot formation defective mutant be1-3 and during the transition from dedifferentiation to redifferentiation stage in wildtype WS. Results show that 155 TFs were differentially expressed between be1-3 mutant and wild type during callus formation, of which 97 genes were up-regulated, and 58 genes were down-regulated; and that 68 genes were differentially expressed during redifferentiation stage, with 40 genes up-regulated and 28 genes down-regulated; whereas at the transition stage from dedifferentiation to redifferention in WS wild type explants, a total of 231 differentially expressed TF genes were identified, including 160 up-regualted genes and 71 down-regulated genes. Among these TF genes, the adventious shoot related transcription factor 1 (ART1) gene encoding a MYB-related (v-myb avian myeloblastosis viral oncogene homolog) TF, was up-regulated 3 217 folds, and was the highest up-regulated gene during be1-3 callus formation. Over expression of the ART1 gene caused defects in callus formation and shoot regeneration and inhibited seedling growth, indicating that the ART1 gene is a negative regulator of callus formation and shoot regeneration. This work not only enriches our knowledge about the transcriptional regulation mechanism of adventious shoot regeneration, but also provides valuable information on candidate TF genes associated with adventious shoot regeneration for future research.

Influence of calcium content of tissue on hyperhydricity and shoot-tip necrosis of in vitro regenerated shoots of Lavandula angustifolia Mill.

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Marília Pereira Machado

2014-10-01

Full Text Available In the present study, the effects of two CaCl2.2H2O levels (440 and 1320 mg L-1 and two subcultures were evaluated on in vitro shoots of Lavandula angustifolia cv. Provence Blue. Ca2+ content of the apical, middle and basal portion of shoots was determined. Increasing CaCl2.2H2O level in the culture medium increased tissue Ca2+ content and decreased hyperhydricity. Shoot-tip necrosis also decreased with 1320 mg L-1 CaCl2.2H2O, but it did not occur in the second subculture. The middle and basal portion had higher Ca2+ content than apical portion. In non-hyperhydric tissues, there were smaller and more juxtaposed cells. Scanning electron microscopy of the leaves demonstrated that trichomes from in vitro leaf surface occurred in smaller quantities.

In vitro regeneration of a common medicinal plant, Ocimum sanctum ...

African Journals Online (AJOL)

Embracing micro-propagation method for large scale production of plantlets and also for protection of appropriate germplasm is a prerequisite that needs to be undertaken in order to develop a rapid in vitro regeneration protocol for Ocimum sanctum L. Shoot tips as well as nodal segments were subjected to numerous ...

Monochromic radiation through light-emitting diode (LED positively augments in vitro shoot regeneration in Orchid (Dendrobium sonia

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Vandita Billore

2017-07-01

Full Text Available Monochromatic lights emitted by light-emitting diodes (LEDs have generated great interest for efficient and controlled growth in vitro, especially of plants which are endangered or require specific intensity and wavelength of light. In the present study, we have evaluated the effect of monochromatic LEDs on in vitro morphogenesis: growth, proliferation of shoot cultures, and rooting of Dendrobium sonia. Different light sources viz. white LEDs (W, blue LEDs (B, yellow LEDs (Y and red LEDs (R were tested under photoperiod of 16 h of exposure and 8 h of dark. The frequency of morphogenesis depended on the wavelength of the applied monochromatic light. Higher wavelength monochromatic light (yellow light was observed to induce higher shoot proliferation (98%, early PLB (protocorm-like bodies formation, differentiation into green buds and shoot initiation as compared to red, blue and white light treatments. Yellow light also yielded higher number of shoots per explants (29 shoots/explant than red, blue and white light treatments. The results suggest that the monochromatic light sources stimulate morphogenic effects on in vitro culture of Dendrobium sonia, and that yellow light treatment can be used to enhance the efficiency of micropropagation.

Efficient regeneration of plants from shoot tip explants of ...

African Journals Online (AJOL)

Dendrobium densiflorum Lindl. is one of the horticulturally important orchids of Nepal due to its beautiful yellowish flower and medicinal properties. The present study was carried out for plant regeneration from shoot tip explants of D. densiflorum by tissue culture technique. The shoot tip explants of this species, obtained ...

High efficiency indirect shoot regeneration and hypericin content in ...

African Journals Online (AJOL)

USER

2010-04-12

Apr 12, 2010 ... opium poppy. Support of hormone-free medium on shoot regeneration from callus has also been underlined for sugarbeet (Doley and Saunders, 1989). In our study, obtained regenerants acclimatized readily, alived (94%) and are still continuing to grow in the greenhouse of our department. Cell or callus ...

Regeneración de brotes adventicios en hojas de guayaba (Psidium guajava L. cultivadas in vitro Adventitious shoot regeneration from in vitro cultured leaves of guava (Psidium guava L.

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Trujillo Sánchez Reinaldo

2004-12-01

Full Text Available La regeneración de brotes adventicios es una etapa clave para la aplicación de las técnicas de ingeniería gené­tica. El presente trabajo tuvo como objetivo el desarrollo de un procedimiento para la regeneración de brotes por organogénesis a partir de hojas de microesquejes o brotes de guayaba (Psidium guajava L. cultivados in vi­tro. Para ello se estudiaron algunos de los principales factores que afectan la regeneración de brotes, tales como la concentración de reguladores del crecimiento (citoquinina, el estado fisiológico del explante y la he­rida. En todos los experimentos se utilizaron como explantes hojas de vitroplantas de guayaba de la variedad Enana Roja Cubana EEA18-40 de diferentes subcultivos. El mejor resultado se alcanzó con una concentración de 0,75 mg/L de 6-bencilaminopurina como suplemento hormonal del medio MS. Se comprobó que para las hojas provenientes de microesquejes con mayor número de subcultivo y tomadas de la parte inferior de los brotes, el potencial morfogenético disminuye significativamente (p60% y el mayor número de brotes por explante promedio (>3 se alcanzó al realizar varias heridas en forma de punteaduras en el nervio central de las hojas. Este protocolo de regeneración constituye una importante herramienta que puede ser empleada para futuros estudios de transformación genética en esta especie. Palabras clave: cultivo de tejidos, explantes de hoja, reguladores de crecimiento, organogénesis, guayaba.Adventitious regeneration is a key step in the application of genetic engineering to the breeding programs of plants. In this work a method for adventitious shoot regeneration from leaves of micropropagated guava shoots has been developed and some of main factors to affect the shoot regeneration like, concentration of plant growth regulators (citoquinine, physiological state of explants and the wound are studied. Leaves from guava in vitro cultured of variety Cuban Red Dwarf 18-40 was used

In vitro somatic embryogenesis and plant regeneration of cassava.

Science.gov (United States)

Szabados, L; Hoyos, R; Roca, W

1987-06-01

An efficient and reproducible plant regeneration system, initiated in somatic tissues, has been devised for cassava (Manihot esculenta Crantz). Somatic embryogenesis has been induced from shoot tips and immature leaves of in vitro shoot cultures of 15 cassava genotypes. Somatic embryos developed directly on the explants when cultured on a medium containing 4-16 mg/l 2,4-D. Differences were observed with respect to the embryogenic capacity of the explants of different varieties. Secondary embryogenesis has been induced by subculture on solid or liquid induction medium. Long term cultures were established and maintained for up to 18 months by repeated subculture of the proliferating somatic embryos. Plantlets developed from primary and secondary embryos in the presence of 0.1 mg/l BAP, 1mg/l GA3, and 0.01 mg/l 2,4-D. Regenerated plants were transferred to the field, and were grown to maturity.

Effect of lanthanum on rooting of in vitro regenerated shoots of Saussurea involucrata Kar. et Kir.

Science.gov (United States)

Guo, Bin; Xu, Ling-Ling; Guan, Zhen-Jun; Wei, Ya-Hui

2012-06-01

In present study, the effect of lanthanum (La) on the rooting of regenerated shoots of Saussurea involucrata Kar. et Kir was analyzed. Rooting occurred from regenerated shoots inoculated on a medium supplemented with La, the plant rooting hormone indole-3-acetic acid (IAA), or both La and IAA together. The highest rooting efficiency (96%), root number/shoot (8.5), and root length (63 mm) were recorded in shoots cultured on medium containing 2.5 μM IAA combined with 100 μM La(3+). In order to elucidate the mechanism of rooting enhancement by La, we examined dynamic changes in antioxidant enzyme activities in plant tissue over time in culture. We found that the activities of peroxidase (POX) and superoxide dismutase (SOD) were significantly higher in plant tissue cultured in IAA plus La than in La or IAA alone. At the same time, the highest H(2)O(2) content was detected in plant tissue in the presence of 2.5 μM IAA plus 100 μM La(3+). In light of these data and previous results, we speculate that La enhanced IAA-induced rooting by acting as a mild abiotic stress to stimulate POX and SOD activities in plant cells. Then, IAA reacted with oxygen and POX to form the ternary complex enzyme-IAA-O(2) that dissociated into IAA radicals and O(2)(-). Subsequently, IAA-induced O(2)(-) readily converted to hydroxyl radical (HO·) via SOD-catalyzed dismutation. Finally, cell wall loosening and cell elongation occurred as a consequence of HO-dependent scission of wall components, leading to root growth. The treatment of IAA combined with La resulted in the highest plantlet survival (80%) compared to single treatments with IAA or La alone. These data suggest that rare earth elements enhance root morphogenesis and the growth of S. involucrata.

In vitro plant regeneration of Spartina argentinensis Parodi

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Mirian Susana Bueno

2012-07-01

; in vitro; organogénesis; variación somaclonal. Abstract: Spartina argentinensis Parodi is the dominant species of the temporally-flooded halophyte communities of the Santa Fe Province, Argentina. It occupies around 20,000 km2 and it is mainly used as low-cost impute forage for cattle production. The objective of this work was to develop a simple method for in vitro plant regeneration of S. argentinensis that could be used for fundamental and applied research. Leaf-basal segments from both young and mature plants, roots tips and immature inflorescences were used as explants. Culture media for calli, shoot, and root induction consisted of Murashige & Skoog salts supplemented with different plant growth regulators (2,4-D; BAP or ANA. Most of the explants (with the exception of root tips showed cell proliferation and calli formation after 30 days of culture. However, only immature inflorescences responded to shoot and root induction when calli were incubated on MS salts plus 2,4_D and BAP (0.1 and 0.01 mg.L-1, respectively, transferred to shoot inductionmedia (MS salts plus BAP, 0.5 mg.L-1 and then to root induction media (MS slts plus NAA 0.5 mg.L-1. Regenerated plants were evaluated for morphological abnormalities and lignin content. Historical analysis of regenerating calli showed that shoots and roots originated via organogenesis. A low proportion of regenerated plants resulted with deficiency in chlorophyll (albino plants and other morphological abnormalities. Interestingly, significant variations in the lignin content were detected between regenerated plants. The protocol described in this work could be used ordinarily for S. argentinensis in vitro plant regeneration and selecting somaclonal variants for breeding purposes. Key words: Spartina argentinensis; in vitro; organogenesis; somaclonal variation.

thidiazuron improves adventitious bud and shoot regeneration

African Journals Online (AJOL)

Prof. Adipala Ekwamu

Induction of adventitious buds and shoots from intact leaves and stem internode segments of two recalcitrant. Ugandan sweetpotato (Ipomoea batatas L.) cultivars was investigated in vitro on Murashige and Skoog (MS) medium, supplemented with 3 different levels (0.5, 2.0 and 4.0 µM) of Thidiazuron (TDZ). Shoots were.

Shoot Organogenesis and Plant Regeneration from Leaf Explants of Lysionotus serratus D. Don

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Qiansheng Li

2013-01-01

Full Text Available The gesneriaceous perennial plant, Lysionotus serratus, has been used in traditional Chinese medicine. It also has a great development potential as an ornamental plant with its attractive foliage and beautiful flowers. An efficient propagation and regeneration system via direct shoot organogenesis from leaf explant was established in this study. High active cytokinin (6-benzyladenine (BA or thidiazuron (TDZ was effective for direct organogenesis of initial induction. Murashige and Skoog (MS growth media containing 0.5 mg L−1 BA alone or with combination of 0.1 mg L−1  α-Naphthaleneacetic acid (NAA were the most effective for shoot proliferation. High BA concentration (1.0 mg L−1 in the media caused high percentage of vitrified shoots though they introduced high shoot proliferation rate. Histological observation indicated that adventitious shoot regeneration on the medium containing 0.5 mg L−1 BA alone occurred directly from leaf epidermal cells without callus formation. Regenerated shoots rooted well on medium containing half-strength MS medium with 0.5 mg L−1 indole-3-butyric acid (IBA and indole-3-acetic acid (IAA, and the plantlets successfully acclimatized and grew vigorously in the greenhouse with a 94.2% and 92.1% survival rate.

Effects of N6-benzylaminopurine and Indole Acetic Acid on In Vitro Shoot Multiplication, Nodule-like Meristem Proliferation and Plant Regeneration of Malaysian Bananas (Musa spp.)

Science.gov (United States)

Sipen, Philip; Davey, Michael R

2012-01-01

Different concentrations of N6-benzylaminopurine (BAP) and indole acetic acid (IAA) in Murashige and Skoog based medium were assessed for their effects on shoot multiplication, nodule-like meristem proliferation and plant regeneration of the Malaysian banana cultivars Pisang Mas, Pisang Nangka, Pisang Berangan and Pisang Awak. BAP at 1–14 mg L−1 with or without 0.2 mg L−1 IAA, or BAP at 7–14 mg L−1 with the same concentration of IAA, was evaluated for shoot multiplication from shoot tips and the proliferation of nodule-like meristems from scalps, respectively. Plant regeneration from scalps was assessed using 1 mg L−1 BAP and 0.2 mg L−1 IAA separately, or a combination of these two growth regulators. Data on shoot multiplication, the proliferation of nodule-like meristems with associated plant regeneration were recorded after 30 days of culture. A maximum of 5 shoots per original shoot tip was achieved on medium supplemented with BAP at 5 mg L−1 (Pisang Nangka), 6 mg L−1 (Pisang Mas and Pisang Berangan), or 7 mg L−1 (Pisang Awak), with 0.2 mg L−1 IAA. BAP at 11 mg L−1 with 0.2 mg L−1 IAA induced the most highly proliferating nodule-like meristems in the four banana cultivars. Plant regeneration from scalps was optimum in all cases on medium containing 1 mg L−1 BAP and 0.2 mg L−1 IAA. This is the first report on the successful induction of highly proliferating nodule-like meristems and plant regeneration from scalps of the Malaysian banana cultivars Pisang Mas, Pisang Nangka, Pisang Berangan and Pisang Awak. PMID:24575235

In Vitro Regeneration of Endangered Medicinal Plant Heliotropium kotschyi (Ramram).

Science.gov (United States)

Sadeq, Manal Ahmed; Pathak, Malabika Roy; Salih, Ahmed Ali; Abido, Mohammed; Abahussain, Asma

2016-01-01

Heliotropium kotschyi (Ramram) is an important endangered medicinal plant distributed in the Kingdom of Bahrain. Plant tissue culture technique is applied for ex situ conservation study. Nodal stem segments are cultured in modified MS media supplemented with various combination and concentration of plant growth regulators (PGRs). Plants are regenerated via shoot organogenesis from the nodal meristems. Plants are regenerated in three different steps: initial shoot development, shoot multiplication, and rooting. After 4 weeks of culture, 100 % explants respond to shoot initiation on the medium containing 8.88 μM BAP and 5.71 μM IAA. The highest frequency of shoot regeneration is observed in the same media after second subculture of shoots. The highest rooting frequency is observed in the presence of 2.85 μM IAA. After root development, the plantlets are transferred to pots filled with soil and 60 % of plants survived after 45 days. This plant regeneration protocol is of great value for rapid desert plant propagation program.

Cryopreservation of in vitro grown shoot tips and apical meristems of the forage legume Arachis pintoi.

Science.gov (United States)

Rey, Hebe Y; Faloci, Mirta; Medina, Ricardo; Dolce, Natalia; Mroginski, Luis; Engelmann, Florent

2009-01-01

A cryopreservation protocol using the encapsulation-dehydration procedure was established for shoot tips (2-3 mm in length) and meristems (0.3-0.5 mm) sampled from in vitro plantlets of diploid and triploid cytotypes of Arachis pintoi. The optimal protocol was the following: after dissection, explants were precultured for 24 h on establishment medium (EM), encapsulated in calcium alginate beads and pretreated in liquid EM medium with daily increasing sucrose concentration (0.5, 0.75, 1.0 M) and desiccated to 22-23 percent moisture content (fresh weight basis). Explants were frozen using slow cooling (1 C per min from 25C to -30C followed by direct immersion in liquid nitrogen), thawed rapidly and post-cultured in liquid EM medium enriched with daily decreasing sucrose concentrations (0.75, 0.50, 0.1 M). Explants were then transferred to solid EM medium in order to achieve shoot regeneration, then on Murashige and Skoog medium supplemented with 0.05 microM naphthalene acetic acid to induce rooting of shoots. With this procedure, 53 percent and 56 percent of cryopreserved shoot tips of the diploid and triploid cytotypes, respectively, survived and formed plants. However, only 16 percent of cryopreserved meristems of both cytotypes regenerated plants. Using ten isozyme systems and seven RAPD profiles, no modification induced by cryopreservation could be detected in plantlets regenerated from cryopreserved material.

Genotype dependent callus induction and shoot regeneration in ...

African Journals Online (AJOL)

SERVER

2007-07-04

Jul 4, 2007 ... tems have been used for improvement of sunflower, but these techniques are mainly limited by the tissue culture response of commercial varieties (Nestares et al., 2002). In sunflower, reports on shoot regeneration from hypocotyls, cotyledons, leaves and meristematic tissues of young plantlets are available ...

Shoot regeneration from cotyledonary leaf explants of jatropha curcas: A biodiesel plant

KAUST Repository

Kumar, Nitish Chandramohana

2010-03-07

A simple, high frequency, and reproducible method for plant regeneration through direct organogenesis from cotyledonary leaf explants of Jatropha curcas was developed using Murashige and Skoog (MS) medium supplemented with different concentrations of thidiazuron (TDZ) or 6-benzyl aminopurine (BAP). Medium containing TDZ has greater influence on regeneration as compared to BAP. The induced shoot buds were transferred to MS medium containing 10 lM kinetin (Kn), 4.5 lM BAP, and 5.5 lM a-naphthaleneacetic acid (NAA) for shoot proliferation. The proliferated shoots could be elongated on MS medium supplemented with different concentrations and combinations of BAP, indole-3-acetic acid (IAA), NAA, and indole-3-butyric acid (IBA). MS medium with 2.25 lM BAP and 8.5 lM IAA was found to be the best combination for shoot elongation. However, significant differences in plant regeneration and shoot elongation were observed among the genotypes studied. Rooting was achieved when the basal cut end of elongated shoots were dipped in half strength MS liquid medium containing dif- ferent concentrations and combinations of IBA, IAA, and NAA for 4 days, followed by transfer to growth regulators free half strength MS medium supplemented 0.25 mg l-1 activated charcoal. Elongated shoot treated with 15 lM IBA, 5.7 lM IAA, and 11 lM NAA resulted in highest percent rooting. The rooted plants could be established in soil with more than 90% survival rate. The method developed may be useful in improvement of J. curcas through genetic modification. © Franciszek Górski Institute of Plant Physiology, Polish Academy of Sciences, Kraków 2010.

Shoot regeneration from cotyledonary leaf explants of jatropha curcas: A biodiesel plant

KAUST Repository

Kumar, Nitish Chandramohana; Vijay Anand, K. G.; Reddy, Muppala P.

2010-01-01

A simple, high frequency, and reproducible method for plant regeneration through direct organogenesis from cotyledonary leaf explants of Jatropha curcas was developed using Murashige and Skoog (MS) medium supplemented with different concentrations of thidiazuron (TDZ) or 6-benzyl aminopurine (BAP). Medium containing TDZ has greater influence on regeneration as compared to BAP. The induced shoot buds were transferred to MS medium containing 10 lM kinetin (Kn), 4.5 lM BAP, and 5.5 lM a-naphthaleneacetic acid (NAA) for shoot proliferation. The proliferated shoots could be elongated on MS medium supplemented with different concentrations and combinations of BAP, indole-3-acetic acid (IAA), NAA, and indole-3-butyric acid (IBA). MS medium with 2.25 lM BAP and 8.5 lM IAA was found to be the best combination for shoot elongation. However, significant differences in plant regeneration and shoot elongation were observed among the genotypes studied. Rooting was achieved when the basal cut end of elongated shoots were dipped in half strength MS liquid medium containing dif- ferent concentrations and combinations of IBA, IAA, and NAA for 4 days, followed by transfer to growth regulators free half strength MS medium supplemented 0.25 mg l-1 activated charcoal. Elongated shoot treated with 15 lM IBA, 5.7 lM IAA, and 11 lM NAA resulted in highest percent rooting. The rooted plants could be established in soil with more than 90% survival rate. The method developed may be useful in improvement of J. curcas through genetic modification. © Franciszek Górski Institute of Plant Physiology, Polish Academy of Sciences, Kraków 2010.

An Integrated Strategy to Identify Key Genes in Almond Adventitious Shoot Regeneration

Science.gov (United States)

Plant genetic transformation usually depends on efficient adventitious regeneration systems. In almond (Prunus dulcis Mill.), regeneration of transgenic adventitious shoots was achieved but with low efficiency. Histological studies identified two main stages of organogenesis in almond explants that ...

Spine micromorphology of normal and hyperhydric Mammillaria gracilis Pfeiff. (Cactaceae) shoots.

Science.gov (United States)

Peharec, P; Posilović, H; Balen, B; Krsnik-Rasol, M

2010-07-01

Artificial conditions of tissue culture affect growth and physiology of crassulacean acid metabolism plants which often results in formation of hyperhydric shoots. In in vitro conditions Mammillaria gracilis Pfeiff. (Cactaceae) growth switches from organized to unorganized way, producing a habituated organogenic callus which simultaneously regenerates morphologically normal as well as altered hyperhydric shoots. In this study, influence of tissue culture conditions on morphology of cactus spines of normal and hyperhydric shoots was investigated. Spines of pot-grown Mammillaria plants and of in vitro regenerated shoots were examined with stereo microscope and scanning electron microscope. The pot-grown plants had 16-17 spines per areole. In vitro grown normal shoots, even though they kept typical shoot morphology, had lower number of spines (11-12) and altered spine morphology. This difference was even more pronounced in spine number (six to seven) and morphology of the hyperhydric shoots. Scanning electron microscopy analysis revealed remarkable differences in micromorphology of spine surface between pot-grown and in vitro grown shoots. Spines of in vitro grown normal shoots showed numerous long trichomes, which were more elongated on spines of the hyperhydric shoots; the corresponding structures on spine surface of pot-grown plants were noticed only as small protrusions. Scanning electron microscopy morphometric studies showed that the spines of pot-grown plants were significantly longer compared to the spines of shoots grown in tissue culture. Moreover, transverse section shape varies from elliptical in pot-grown plants to circular in normal and hyperhydric shoots grown in vitro. Cluster and correspondence analyses performed on the scanning electron microscope obtained results suggest great variability among spines of pot-grown plants. Spines of in vitro grown normal and hyperhydric shoots showed low level of morphological variation among themselves despite the

High Frequency Plant Regeneration System from Transverse Thin Cell Layer Section of In vitro Derived ‘Nadia’ Ginger Microrhizome

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Dikash Singh THINGBAIJAM

2014-03-01

Full Text Available An efficient and reproducible procedure is outlined for rapid in vitro multiplication of Zingiber officinale var. ‘Nadia’ through high frequency shoot proliferation from transverse thin cell layer (tTCL sections of in vitro derived microrhizome. In vitro derived microrhizome of size 500 μm in thickness was used as initial explants for induction of somatic embryos. Among the different phytohormones tested, tTCL explants shows maximum calli proliferation in medium containing 2 mg/L 2,4-Dichlorophenoxyacetic acid (88.30±0.11%. Reduced concentration of 2,4 Dichlorophenoxyacetic acid was supplemented with different cytokinins for regeneration of callus. Among the different medium tested, optimum redifferentiation of somatic embryos were observed in medium containing 0.2 mg/L 2,4 Dichlorophenoxyacetic acid and 6.0 mg/L BAP (141.08±0.25. Clump of regenerated plantlets were further subculture and transfer into microrhizome inducing medium containing high sucrose concentration (8%. Plantlets with well developed microrhizome were successfully acclimatized and eventually transferred to the field. The application of studying embryo section for regeneration of plants might be useful alternative to ginger improvement programme. Histological analysis showed formation of somatic embryos and regenerated adventitious shoot.

In-vitro regeneration of castor bean (ricinus communis, L) and physic nut (jatropha curcas, L.)

International Nuclear Information System (INIS)

Afful, N. T.

2008-06-01

Investigations were conducted on in vitro regeneration of physic nut (Jatropha curcas, 1.) and castor bean (Ricinus communis, 1.) using zygotic embryo culture, shoot tip and meristem culture as well as somatic embryogenesis. Zygotic embryos from different stages of fruit maturity cultured on Murashige and Skoog (MS) medium supplemented with benzylaminopurine (BAP), kinetin and 2-isopentyl (2iP) produced vigorous seedlings with roots. However the optimal concentration of cytokinin for seedling development depended on the type cytokinin as well as the maturity stage of the fruit. Embryos from yellow fruits of Jatropha had the highest germination (100%) on an MS medium supplemented with 0.5 mg/L kinetin or 1 mg/L BAP while in Ricinus embryos from black fruits had the highest percentage germination (97%) on an MS medium modified with I mg/L 2iP or 0.5 or I mg/L BAP. Decrease in embryo viability especially in Jatropha as the seed matured may be due to high oil content in matured (black) fruit. Oil content in both Jatropha and Ricinus increased as the fruit matured from green to black and this may have effect on germination of zygotic embryos. In vitro culture response of shoot tip or meristem was significantly influenced by the concentration of growth regulators in the culture medium as well as the plant species. In both plant species, 2iP resulted in highest percentage shoot regeneration. The optimal concentration of 2iP was 0.5 mg/L for Ricinus shoot tip and 1.5 or 2.0 mg/L for Jatropha, indicating genotypic difference between the species. However excessive callus formation and browning in Ricinus led to the loss of all the regenerants. Shoot regeneration from meristem explants was also influenced by growth regulators with 2iP again being the best. Cultures of cotyledon and leaf lobe explants on MS medium supplemented with 2, 4-D and picloram resulted in callus induction with the exception of cotyledons obtained from black fruits. However transfer of calli onto fresh

Adventitious shoot regeneration from leaf explants of the valuable ...

African Journals Online (AJOL)

The objective of this study was to develop an efficient protocol for adventitious shoot regeneration for Plectranthus barbatus Andrews using leaf explants. The explants were cultured on MS (Murashige and Skoog, 1962) medium containing various concentration of kinetin (KN), 6-benzylaminopurine (BAP) and thidiazuron ...

High Frequency Plant Regeneration System from Transverse Thin Cell Layer Section of In vitro Derived ‘Nadia’ Ginger Microrhizome

Directory of Open Access Journals (Sweden)

Dikash Singh THINGBAIJAM

2014-03-01

Full Text Available An efficient and reproducible procedure is outlined for rapid in vitro multiplication of Zingiber officinale var. ‘Nadia’ through high frequency shoot proliferation from transverse thin cell layer (tTCL sections of in vitro derived microrhizome. In vitro derived microrhizome of size 500 μm in thickness was used as initial explants for induction of somatic embryos. Among the different phytohormones tested, tTCL explants shows maximum calli proliferation in medium containing 2 mg/L 2,4-Dichlorophenoxyacetic acid (88.30±0.11%. Reduced concentration of 2,4 Dichlorophenoxyacetic acid was supplemented with different cytokinins for regeneration of callus. Among the different medium tested, optimum redifferentiation of somatic embryos were observed in medium containing 0.2 mg/L 2,4 Dichlorophenoxyacetic acid and 6.0 mg/L BAP (141.08±0.25. Clump of regenerated plantlets were further subculture and transfer into microrhizome inducing medium containing high sucrose concentration (8%. Plantlets with well developed microrhizome were successfully acclimatized and eventually transferred to the field. The application of studying embryo section for regeneration of plants might be useful alternative to ginger improvement programme. Histological analysis showed formation of somatic embryos and regenerated adventitious shoot.

Adventitious shoot regeneration from leaf explants of the valuable ...

African Journals Online (AJOL)

Jane

2011-08-10

Aug 10, 2011 ... 2Department of Plant Biology and Plant Biotechnology, St. Joseph's College, Tiruchirappalli – 620 002, South India. Accepted 28 March, 2011 .... Effect of cytokinins (KN and BAP) alone or in combination with NAA on direct shoot bud regeneration from leaf explants of P. barbatus. Plant growth regulator ...

Rosette growth of shoots in chrysanthemum (Dendranthema grandiflora Tzvelev) as a result of in vitro propagation of plants and gamma irradiation used for mutation induction

International Nuclear Information System (INIS)

Jerzy, M.; Zalewska, M.; Lema, J.

1999-01-01

Rosette growth of chrysanthemum shoots in 'Mrs. R.C. Puling' was observed after in vitro propagation with explants obtained from vernalised and non-vernalised stock plants. The phenomenon was also observed as a result of the exposure of leaf explants to gamma radiation used for in vitro regeneration of plants in mutation induction. The higher the irradiation dose, the more considerable the rosette growth. Following the 4th pinching of shoot tips, only elongating growth of plants was observed

Detection of sodium azide-induced mutagenicity in the regenerated shoots of artemisia annual L., using internal transcribed spacer (its) sequences of nrDNA

International Nuclear Information System (INIS)

Al-Qurainy, F.; Al-Hemaidi, F.M.; Khan, S.; Ali, M.A.; Tarroum, M.; Ashraf, M.

2011-01-01

Sodium azide (NaN/sub 3/) is a well known chemical mutagen which can effectively cause point mutation in plant genome. The mutagenicity by this potential mutagen was assessed in the regenerated mutant shoots of Artemisia annua using internal transcribed spacer (ITS) sequences of n rDNA. Insertions and/or deletions were detected in n rDNA-ITS sequences of all mutant shoots and compared with control ones using the ClustalX program. The regenerated shoots TS1 and TS2 had deleted bases, whereas TS3, TS4 and TS5 had insertions, because NaN/sub 3/ replaced the cytosine (C) by thymine (T) (C - T) (shoots; TS1 and TS4) and thymine (T) replaced by guanine (G) (T - G) (shoot; TS5), respectively. Artemisinin content was also measured in the regenerated six-week-old shoots of A. annua. All regenerated shoots had enhanced level of this compound as compared to that in the controls, being highest in the regenerated shoot TS3. (author)

Agrobacterium-mediated transformation of cotton (Gossypium hirsutum) shoot apex with a fungal phytase gene improves phosphorus acquisition.

Science.gov (United States)

Ma, Zhiying; Liu, Jianfeng; Wang, Xingfen

2013-01-01

Cotton is an important world economic crop plant. It is considered that cotton is recalcitrant to in vitro proliferation. Somatic embryogenesis and plant regeneration has been successful by using hypocotyl, whereas it is highly genotype dependent. Here, a genotype-independent cotton regeneration protocol from shoot apices is presented. Shoot apices from 3- to 5-day-old seedlings of cotton are infected with an Agrobacterium strain, EHA105, carrying the binary vector pC-KSA contained phytase gene (phyA) and the marker gene neomycin phosphotransferase (NPTII), and directly regenerated as shoots in vitro. Rooted shoots can be obtained within 6-8 weeks. Plants that survived by leaf painting kanamycin (kan) were -further analyzed by DNA and RNA blottings. The transgenic plants with increased the phosphorus (P) acquisition efficiency were obtained following the transformation method.

Improved recovery of cryotherapy-treated shoot tips following thermotherapy of in vitro-grown stock shoots of raspberry (Rubus idaeus L.).

Science.gov (United States)

Wang, Qiaochun; Valkonen, Jari P T

2009-01-01

Raspberry bushy dwarf virus (RBDV) can be efficiently eradicated from raspberry plants (Rubus idaeus) by a procedure combining thermotherapy and cryotherapy. However, the bottleneck of this procedure is that, following thermotherapy, cryopreserved shoot tips become chlorotic during regrowth and eventually die after several subcultures. In addition, survival of heat-treated stock shoots and recovery of cryopreserved shoot tips following thermotherapy are low. The present study focused towards improving regrowth of cryopreserved raspberry shoot tips following thermotherapy. Results showed that preconditioning stock shoots with salicylic acid (SA; 0.01-0.1 mM) markedly increased survival of stock shoots after 4 weeks of thermotherapy. Regrowth of cryopreserved shoot tips following thermotherapy was also significantly enhanced when SA (0.05-0.1 mM) was used for preconditioning stock shoots. Addition of either Fe-ethylenediaminetetracetic acid (Fe-EDTA, 50 mg per L) or Fe-ethylenediaminedi(o)hydroxyphenylacetic acid (Fe-EDDHA, 50 mg per L) to post-culture medium strongly promoted regrowth and totally prevented chlorosis of shoots regenerated from cryopreserved shoot tips following thermotherapy. Using the parameters optimized in the present study, about 80 percent survival of heat-treated stock shoots and about 33 percent regrowth of cryopreserved shoot tips following thermotherapy were obtained. Morphology of plants regenerated from cryopreserved shoot tips following thermotherapy was identical to that of control plants, based on observations of leaf shape and size, internode length and plant height. Optimization of the thermotherapy procedure followed by cryotherapy will facilitate the wider application of this technique to eliminate viruses which can invade meristems.

Rapid in vitro propagation system through shoot tip cultures of Vitex trifolia L.-an important multipurpose plant of the Pacific traditional Medicine.

Science.gov (United States)

Ahmed, Rafique; Anis, Mohammad

2014-07-01

A rapid and efficient plant propagation system through shoot tip explants was established in Vitex trifolia L., a medicinally important plant belonging to the family Verbenaceae. Multiple shoots were induced directly on Murashige and Skoog (MS) medium consisting of different cytokinins, 6-benzyladenine (BA), kinetin (Kin) and 2-isopentenyl adenine (2-iP), BA at an optimal concentration of 5.0 μM was most effective in inducing multiple shoots where 90 % explants responded with an average shoot number (4.4±0.1) and shoot length (2.0±0.1 cm) after 6 weeks of culture. Inclusion of NAA in the culture medium along with the optimum concentration of BA promoted a higher rate of shoot multiplication and length of the shoot, where 19.2±0.3 well-grown healthy shoots with an average shoot length of 4.4±0.1 cm were obtained on completion of 12 weeks culture period. Ex vitro rooting was achieved best directly in soilrite when basal portion of the shoots were treated with 500 μM indole-3-butyric acid for 15 min which was the most effective in inducing roots, as 95 % of the microshoots produced roots. Plantlets went through a hardening phase in a controlled plant growth chamber, prior to ex-vitro transfer. Micropropagated plants grew well, attained maturity and flowered with 92 % survival rate. The results of this study provide the first report on in vitro plant regeneration of Vitex trifolia L. using shoot tip explants.

Preculturing effect of thidiazuron on in vitro shoot multiplication and micropropagation round in Capparis decidua (Forsk.) an important multipurpose plant.

Science.gov (United States)

Bukhari, Najat A W; Siddique, Iram; Perveen, Kahkashan

2016-09-01

An efficient protocol was developed for clonal multiplication of an important shrub: Capparis decidua (Forsk.) Edgew, through in vitro shoot induction and multiplication from nodal explants. Pretreatment of nodal explants in a liquid Murashige and Skoog (MS) medium augmented with various thidiazuron (TDZ) concentrations at relatively high levels (5-100 μM) for different time duration (4, 8, 12 and 16 d), proved a significant approach for in vitro shoot production. After an initial exposure time to TDZ, nodal explants were inoculated onto a MS basal medium devoid of TDZ for further induction and proliferation. The highest regeneration rate (85%), average number of shoots/explant (8.7 ± 0.22) and maximum shoot length (3.9 ± 0.33 cm) were obtained from the nodal explants exposed to 50 μM TDZ for 8 d. The nodal explants excised from the proliferated cultures of TDZ (50 μM) for 8 d were used as explants and showed an enhancement rate after next three round of in vitro propagation. Best results for rooting was obtained by ex vitro treatment of shoots with 200 μM indole-3-butyric acid (IBA) for 20 min. as it produced an average of 5.7 ± 0.41 roots per microshoot with 4.4 ± 0.39 cm root length in 84% shoots. Different planting substrates was tested for maximum survival of hardening off micropropagated plantlets and soilrite proved most effective than others as 97.1 ± 7.21 plantlets survived. All micropropagated plants grew well in natural conditions and showed similar morphology to the mother plant.

Protocol optimization for in vitro shoot multiplication of Jackfruit ...

African Journals Online (AJOL)

Jemal

2017-01-11

Jan 11, 2017 ... Protocol optimization for in vitro shoot multiplication of ... shoot length and leaf number, whereby 2 mg/L BAP alone was found to be the best with a mean shoot .... Analysis of variance showed that the interaction between.

Factors affecting in vitro plant regeneration of the critically endangered Mediterranean knapweed ( Centaurea tchihatcheffii Fisch et. Mey)

Science.gov (United States)

Ozel, Cigdem Alev; Khawar, Khalid Mahmood; Mirici, Semra; Ozcan, Sebahattin; Arslan, Orhan

2006-10-01

Habitat destruction has resulted in the extinction of many plant species from the earth, and many more face extinction. Likely, the annual endemic Mediterranean knapweed ( Centaurea tchihatcheffii) growing in the Golbasi district of Ankara, Turkey is facing extinction and needs urgent conservation. Plant tissue culture, a potentially useful technique for ex situ multiplication, was used for the restoration of this ill-fated plant through seed germination, micropropagation from stem nodes, and adventitious shoot regeneration from immature zygotic embryos. The seeds were highly dormant and very difficult to germinate. No results were obtained from the micropropagation of stem nodes. However, immature zygotic embryos showed the highest adventitious shoot regeneration on Murashige and Skoog (MS) medium, containing 1 mg l-1 kinetin and 0.25 mg l-1 NAA. Regenerated shoots were best rooted on MS medium containing 1 mg l-1 IBA and transferred to the greenhouse for flowering and seed set. As such, the present work is the first record of in vitro propagation of critically endangered C. tchihatcheffii, using immature zygotic embryos, and is a step forward towards conservation of this indigenous species.

DNA Methylation and Methylation Polymorphism in Genetically Stable In vitro Regenerates of Jatropha curcas L. Using Methylation-Sensitive AFLP Markers.

Science.gov (United States)

Rathore, Mangal S; Jha, Bhavanath

2016-03-01

The present investigation aimed to evaluate the degree and pattern of DNA methylation using methylation-sensitive AFLP (MS-AFLP) markers in genetically stable in vitro regenerates of Jatropha curcas L.. The genetically stable in vitro regenerates were raised through direct organogenesis via enhanced axillary shoot bud proliferation (Protocol-1) and in vitro-derived leaf regeneration (Protocol-2). Ten selective combinations of MS-AFLP primers produced 462 and 477 MS-AFLP bands in Protocol-1 (P-1) and Protocol-2 (P-2) regenerates, respectively. In P-1 regenerates, 15.8-31.17 % DNA was found methylated with an average of 25.24 %. In P-2 regenerates, 15.93-32.7 % DNA was found methylated with an average of 24.11 %. Using MS-AFLP in P-1 and P-2 regenerates, 11.52-25.53 % and 13.33-25.47 % polymorphism in methylated DNA was reported, respectively. Compared to the mother plant, P-1 regenerates showed hyper-methylation while P-2 showed hypo-methylation. The results clearly indicated alternation in degree and pattern of DNA methylation; hence, epigenetic instability in the genetically stable in vitro regenerates of J. curcas, developed so far using two different regeneration systems and explants of two different origins. The homologous nucleotide fragments in genomes of P-1 and P-2 regenerates showing methylation re-patterning might be involved in immediate adaptive responses and developmental processes through differential regulation of transcriptome under in vitro conditions.

Plant regeneration from protoplasts ofVicia narbonensis via somatic embryogenesis and shoot organogenesis.

Science.gov (United States)

Tegeder, M; Kohn, H; Nibbe, M; Schieder, O; Pickardt, T

1996-11-01

Protoplasts ofVicia narbonensis isolated from epicotyls and shoot tips of etiolated seedlings were embedded in 1.4% sodium-alginate at a final density of 2.5×10(5) protoplasts/ml and cultivated in Kao and Michayluk-medium containing 0.5 mg/I of each of 2,4- dichlorophenoxyacetic acid, naphthylacetic acid and 6 -benzylaminopurine. A division frequency of 36% and a plating efficiency of 0.40-0.5% were obtained. Six weeks after embedding, protoplast-derived calluses were transferred onto gelrite-solidified Murashige and Skoog-media containing various growth regulators. Regeneration of plants was achieved via two morphologically distinguishable pathways. A two step protocol (initially on medium with a high auxin concentration followed by a culture phase with lowered auxin amount) was used to regenerate somatic embryos, whereas cultivation on medium containing thidiazuron and naphthylacetic acid resulted in shoot morphogenesis. Mature plants were recovered from both somatic embryos as well as from thidiazuron-induced shoots.

Improving adventitious shoot regeneration from cultured leaf ...

African Journals Online (AJOL)

The effect of various concentrations of thidiazuron (TDZ) with or without 2.7 μM of α-naphthalene acetic acid (NAA) on adventitious shoot formation of two Petunia hybrida cultivars was studied. Seeds from 'Daddy Blue' and 'Dreams White' cultivars were germinated in vitro. Expanded leaves from both seedlings and ...

Direct shoot organogenesis of Digitalis trojana Ivan., an endemic ...

African Journals Online (AJOL)

STORAGESEVER

2010-03-15

Mar 15, 2010 ... An efficient protocol for in vitro propagation of Digitalis trojana Ivan. was developed via adventitious shoot regeneration. Leaf explants were cultured on MS which were supplemented with different concentrations of NAA (0.1, 0.5, 1.0 mg/ml) and BAP (0.1, 0.5, 1.0, 3.0, 5.0 mg/ml) for shoot formation.

In vitro regeneration from petiole explants of non-toxic Jatropha curcas

KAUST Repository

Kumar, Nitish

2011-01-01

Jatropha curcas, a multipurpose shrub has acquired significant economic potential as biodiesel plant. The seeds or pressed cake is toxic due to the presence of toxic substances and is not useful as food/fodder despite having the best protein composition. A simple, efficient, and reproducible method for plant regeneration through direct organogenesis from petiole explants of non-toxic J. curcas was developed using Murashige and Skoog (MS) medium supplemented with different concentrations of thidiazuron (TDZ). The best induction of shoot buds (57.61%), and number of shoot buds (4.98) per explant were obtained when in vitro petiole explants were placed horizontally on MS medium supplemented with 2.27 mu M TDZ. The Induced shoot buds were transferred to MS medium containing 10 mu M kinetin (Kn), 4.5 mu M 6-benzyl aminopurine (BA), and 5.5 mu M alpha-naphthaleneacetic acid (NAA) for shoot proliferation and subsequent elongation was achieved on MS medium supplemented with 2.25 mu M BA and 8.5 mu M IAA. The elongated shoots could be rooted on half-strength MS medium with 15 mu M IBA, 11.4 mu M IAA and 5.5 mu M NAA with more than 90% survival rate. (C) 2010 Elsevier B.V. All rights reserved.

Plant regeneration through organogenesis and shoot proliferation in ...

African Journals Online (AJOL)

STORAGESEVER

2008-10-20

Oct 20, 2008 ... Effect of 2,4-D, NAA and BA (in MS medium) on Trichodesma indicum callus induction and shoot .... Structure/activity relationships of the fenotoxic potentcies of sixteen pyrrolizidine ... In vitro propagation of Cordia verbe- ...

Micropropagation of Asparagus by in vitro shoot culture.

Science.gov (United States)

Stajner, Nataša

2013-01-01

Asparagus officinalis is most extensively studied species within the genus Asparagus, which is well known as garden asparagus. This species is dioecious with unisexual flowers, which means that generative propagation gives roughly equal number of male and female plants. Male plants are high yielders and preferred commercially over female plants. Tissue culture techniques could efficiently promote vegetative propagation of male plants and pave the way for efficient plant breeding.This chapter describes an efficient micropropagation protocol for developing rapid growing in vitro Asparagus shoot cultures. The source of explants, inoculation, and shoot proliferation, followed by shoot propagation, rooting, and acclimatization is described. The optimal medium for Asparagus micropropagation described in this chapter is composed of MS macro- and microelements and a combination of auxins and cytokinins. Plant growth regulators NAA, kinetin, and BA were used in various concentrations. Three different media representing the whole micropropagation protocol of Asparagus are described; medium for shoot initiation, medium for shoot multiplication, and medium for root formation. By in vitro propagation of Asparagus, root initiation is difficult, but can be promoted by adding growth retardant ancymidol which also greatly promotes shoot development and suppresses callus formation.

Cytokinin induced shoot regeneration and flowering of Scoparia dulcis L. (Scrophulariaceae)-an ethnomedicinal herb.

Science.gov (United States)

Premkumar, G; Sankaranarayanan, R; Jeeva, S; Rajarathinam, K

2011-06-01

To develop an improved protocol for micropropagation of ethnomedicinally important Scoparia dulcis (S. dulcis) L. Explants were inoculated on MS basal medium supplemented with kinetin and 6-benzylaminopurine for shoot bud induction. To enhance the shoot induction, various auxins like 3-indoleacetic acid or 3-indolebutyric acid or α-naphthylacetic acid were tested along with 2.32 M KI and 4.44 µM BAP. The regenerated shoots were rooted in half strength MS medium supplemented with various concentrations of IAA, IBA or NAA. After roots were developed, the plantlets were transplanted to pots filled with vermiculate and sand and kept in growth chamber with 70%-80% humidity under 16 h photoperiod. After acclimatization, the plantlets were transferred to the garden and survival percentage was calculated. Data were statistically analyzed and means were compared using Duncan's multiple range test (Pdulcis. Shoot induction on young leaf explants was most successful in MS medium supplemented with combination of two cytokinins (2.32 µM KI and 4.44 µM BAP) 2.85 µM IAA, 10% CM and 1 483.79 µM adenine sulfate. A single young leaf explant was capable of producing 59 shoots after 13 days of culture. Flower was induced in medium supplemented with combination of KI and BAP. Cytokinins are the key factor to induce the direct shoot regeneration and flowering of S. dulcis.

Direct adventitious shoot bud formation on hypocotyls explants in Millettia pinnata (L.) Panigrahi- a biodiesel producing medicinal tree species

OpenAIRE

Nagar, Durga Singh; Jha, Suman Kumar; Jani, Jigar

2015-01-01

A reproducible protocol developed for in vitro regeneration of Milletia pinnata using hypocotyl segments. Multiple shoots were induced from hypocotyl explants through direct adventitious shoot bud regeneration. The proximal end of hypocotyls was responsive for shoot bud induction. Silver nitrate and adenine sulphate had a positive effect on shoot bud induction and elongation. The maximum response and number of shoot bud produced in media supplemented with 8.88 μM BAP with 108.6 μM adenine sul...

In vitro propagation of lnula royleana DC

Directory of Open Access Journals (Sweden)

Anna Stojakowska

2011-01-01

Full Text Available A micropropagation method, through axillary shoot proliferation, was elaborated for Inula royleana DC. (Asteraceae, a medicinal plant native of Himalaya. Primary explants (cotyledonary node explants and secondary explants (node explants of in vitro regenerated shoots of the plant, inoculated on MS medium supplemented with 0.1 μM NAA and 5.0 μM kinetin, regenerated 3.4 ± 1.2 and 5.1 ± 1.9 axillary shoots per explant, respectively. The regenerated shoots were easily rooting and adapting to growth in soil.

Effects of gamma-rays irradiation in seed of mungbean (vigna radiata (L.) wilczek) composition of media on shoot regeneration of explants from node of cotyledon

International Nuclear Information System (INIS)

Hutabarat, Dameria; H, Soeranto

1998-01-01

Study the effects of gamma-rays irradiation and composition and media on shoot regeneration of explants from node of cotyledon of mungbean. Wallet variety have been conducted. The explants derived of irradiated seeds of 10-20 Gy of gamma rays were planted in the 0.7% agar solution. One day after planting in the agar media the embryo axis of germinate seed were removed and the node of cotyledon were cultured in the regeneration media as examples. The results shown that shoot regeneration was influenced by media composition and the doses of gamma rays irradiation in seed. In the MURASHIGE and SKOOG medium which contain of BAP or 2-iP or Kinetin with 3 ppm concentrate respectively the explants could produced 100% of shoots. However, the highest. number of produced shoot (3 shoots) was showed in the medium which contained of BAP. The medium with I ppm concentrate od BAP could produced 100% shoot regeneration and the maximum number of shoots (4 shoots) per explant was showed in with 5 ppm. concentrate of BAP. The effectivity off BAP for shoot regeneration by enrichment of 12 ppm Ag 2 SO 4 in the media. Irradiation of 10-20 Gy gamma rays on seeds of mungbean walet variety could improved shoot regeneration of explants from node cotyledon. (author)

In vitro regeneration of solanum aethiopicum L. (scarlet eggplant), an african vegetable crop with potential ornamental value

Science.gov (United States)

Successful in vitro regeneration of plantlets was obtained from shoot tips of five Solanum aethiopicum (African eggplants) accessions evaluated in two media, M1 and M2. The M1 medium consisted of Murashige and Skoog (MS) basal salt mixture supplemented with 20 g/L sucrose, 0.75 g/L MgCl2, and 2 g/L ...

In vitro regeneration of Drosera burmannii Vahl.: a carnivorous plant of north-east India.

Science.gov (United States)

Yanthan, J Sureni; Kehie, Mechuselie; Kumaria, Suman; Tandon, Pramod

2017-06-01

An efficient in vitro regeneration protocol has been developed from shoot tips of Drosera burmannii Vahl., a carnivorous plant of north-east India. Various plant growth regulators were used to study their efficacy in the induction of multiple shoots and roots. Of the various treatments, the maximum number of shoots (28.8 ± 1.5) and roots (9.7 ± 0.6) was observed in one-fourth strength standard medium (MS with 50 mg/l citric acid and 10 mg/l ascorbic acid) supplemented with 4 mg/l 6-benzylaminopurine (BAP) and 4 mg/l α-naphthalene acetic acid (NAA) followed by 26.8 ± 1.4 shoots in one-fourth strength SM fortified with 4 mg/l kinetin (KN) and 4 mg/l NAA. The well-developed plantlets with shoots and roots were potted in small plastic glasses filled with a mixture of sand and farmyard manure (3:1); these plantlets when transferred to a glasshouse for hardening and acclimatization showed 90% survival.

Genotype, explant, medium, light and radiation effects on the in vitro plant regeneration in alfalfa (Medicago Sativa L.)

International Nuclear Information System (INIS)

El-Fiki, A.A.; Abdel-Hameed, A.A.M.; Sayed, A.I.H.

2005-01-01

The relative importance of genotype, explants, radiation, medium and light and their interactions for in vitro plant regeneration via somatic embryogenesis in alfalfa (Medicago sativa L.) has been studied. Shoot and leaf explants of two commercially grown Egyptian cultivars, Al-Wadi Al-Gadid and Siwa Tarkibi, were used in the study. The effect of gamma radiation doses 40, 80, 120 and 160 Gy were negative on plant regeneration, in spite of increase with some treatments. The best results of plant regeneration were obtained with dose 40 Gy with control light regime (16 h) on MS + 0.5 mg NAA + 1.5 mg BAP in both shoot and leaf explants of cv. Al-Wadi. The shoot explant of cv. Siwa was sensitive for gamma radiation dose 40 Gy while affirmative effect was obtained in leaf explant on MS + 1.0 mg NAA + 0.5 mg BAP with control light regime. However, dose 80 Gy showed the best results on MS + 0.5 mg NAA + 0.5 mg BAP in shoot and leaf explants of both cultivars, with control light regime in shoot explant and dark/light (DL) and dark/dark (DD) in leaf explant of cv. Al-Wadi, while with light/dark (LD) in shoot explant and control light regime in leaf explant of cv. Siwa. On the other hand, the highest plant regeneration ratio observed with dose 120 Gy were on 1.5 mg NAA + 0.5 mg BAP with control light regime in shoot and leaf explants of cv. Al-Wadi but on 0.5 mg NAA + 0.5 mg BAP with control and dark/light (DL) light regime in shoot and leaf explants of cv. Siwa. Whereas, the radiation dose 160 Gy showed severe effect on plant regeneration in both cultivars but highest percentage was observed on MS + 0.5 mg NAA + 0.5 mg BAP with dark/light (DL) in shoot explant, MS + 0.5 mg NAA + 1.5 mg BAP with control light regime in leaf explant of cv. Al-Wadi, MS + 0.5 mg NAA + 1.5 mg BAP in shoot explant and MS + 0.5 mg NAA + 0.5 mg BAP in leaf explant with dark/light (DL) in cv. Siwa. However, the effects of the same doses on callus growth showed that the highest callus weight was

In vitro propagation, encapsulation, and genetic fidelity analysis of Terminalia arjuna: a cardioprotective medicinal tree.

Science.gov (United States)

Gupta, Amit K; Harish; Rai, Manoj K; Phulwaria, Mahendra; Agarwal, Tanvi; Shekhawat, N S

2014-07-01

The present study described an improved and reproducible in vitro regeneration system for Terminalia arjuna using nodal segment explants obtained from a mature plant. Shoot tips excised from in vitro proliferated shoots were encapsulated in 3 % sodium alginate and 100 mM CaCl2[Symbol: see text]2H2O for the development of synthetic seeds which may be applicable in short-term storage and germplasm exchange of elite genotype. Shoot multiplication was significantly influenced by a number of factors, namely types and concentrations of plant growth regulators, medium composition, repeated transfer of mother explants, subculturing of in vitro regenerated shoot clumps, agar concentrations, and temperature. Maximum numbers of shoots (16.50 ± 3.67) were observed on modified Murashige and Skoog (MMS) medium containing 0.5 mg l(-1) of benzylaminopurine (BAP) and 0.1 mg l(-1) of naphthalene acetic acid (NAA). To shortening the regeneration pathway, rooting of micropropagated shoots under in vitro condition was excluded and an experiment on ex vitro rooting was conducted and it was observed that the highest percentage of shoots rooted ex vitro when treated with indole-3-butyric acid (IBA, 250 mg l(-1)) + 2-naphthoxy acetic acid (NOA, 250 mg l(-1)) for 5 min. The well-developed ex vitro rooted shoots were acclimatized successfully in soilrite under greenhouse conditions with 80 % survival of plants. Randomly amplified polymorphic DNA (RAPD) analysis confirmed that all the regenerated plants were genetically identical to the mother plant, suggesting the absence of detectable genetic variation in the regenerated plantlets. To the best of our knowledge, this is the first report on synthetic seed production as well as ex vitro rooting and genetic fidelity assessment of micropropagated shoots of T. arjuna.

Enhanced in vitro multiple shoot induction in elite Pakistani guava ...

African Journals Online (AJOL)

Elite guava (Psidium guajava L.) strains of cv. Safeda were explored in vitro for multiple shoot induction. Shoot induction was enhanced up to 83% with 3.5 to 4.25 shoots per single node cutting and shoot tip explants, respectively, using higher levels of benzyl amino purine (BAP) in Murashige and Skoog (MS) medium.

Optimization of culture conditions (sucrose, pH, and photoperiod) for in vitro regeneration and early detection of somaclonal variation in ginger lime (Citrus assamensis).

Science.gov (United States)

Yaacob, Jamilah Syafawati; Mahmad, Noraini; Mat Taha, Rosna; Mohamed, Normadiha; Mad Yussof, Anis Idayu; Saleh, Azani

2014-01-01

Various explants (stem, leaf, and root) of Citrus assamensis were cultured on MS media supplemented with various combinations and concentrations (0.5-2.0 mg L(-1)) of NAA and BAP. Optimum shoot and root regeneration were obtained from stem cultures supplemented with 1.5 mg L(-1) NAA and 2.0 mg L(-1) BAP, respectively. Explant type affects the success of tissue culture of this species, whereby stem explants were observed to be the most responsive. Addition of 30 gL(-1) sucrose and pH of 5.8 was most optimum for in vitro regeneration of this species. Photoperiod of 16 hours of light and 8 hours of darkness was most optimum for shoot regeneration, but photoperiod of 24 hours of darkness was beneficial for production of callus. The morphology (macro and micro) and anatomy of in vivo and in vitro/ex vitro Citrus assamensis were also observed to elucidate any irregularities (or somaclonal variation) that may arise due to tissue culture protocols. Several minor micromorphological and anatomical differences were observed, possibly due to stress of tissue culture, but in vitro plantlets are expected to revert back to normal phenotype following full adaptation to the natural environment.

Optimization of Callus Induction and Regeneration in Two Fenugreek Landraces as a Medicinal Plant during in vitro Condition

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Hasan Hasani Jifroudi

2017-10-01

Full Text Available Introduction: Fenugreek (Trigonella foenum- graecum is a medicinal plant extensively distributed in most regions of the world. Fenugreek is an annual plant from the family of papilionaceae, leguminosae. Fenugreek leaves and seeds have been used extensively to prepare extracts and powders for medicinal uses. Its root, leaf and seed contain a number of important medicinal compounds such as polysaccharide, galactomannan, different saponins such as diosgenin, yamogenin, mucilage, volatile oil and alkaloids such as choline and trigonelline. Plant tissue culture is fundamental to most aspects of biotechnology of plants. Establishment of an efficient callus induction and direct regeneration protocol is an essential prerequisite in harnessing the advantage of cell and tissue culture for genetic improvement. For the successful application of the tissue culture technique in plant breeding, callus induction and plant regeneration potential of each plant must be determined. The present study was performed in order to determine the optimum concentration of plant growth regulators (IBA + TDZ for producing of in vitro plantlet using cotyledon and hypocotyl as an explant for two different Iranian genotypes (Ardestani and Neyshabouri. Materials and Methods: In this investigation, Ardestani and Neyshabouri genotypes were used for callus induction and direct shoot regeneration. The medium used in this investigation was MS (Murashige and Skoog basal medium. Then seeds were germinated on MS medium. For callus induction and direct shoot regeneration, cotyledon and hypocotyl explants were excised from 8-day-old sterile seedlings and cultured on MS medium containing various concentrations of IBA and TDZ. In this experiment, two combinations (TDZ + IBA were used. In the first composition, IBA had four levels (0, 0.1, 0.3, 0.5 mg l-1 and TDZ had five levels (0, 0.2, 0.4, 0.6, 0.8 mg l-1 and in the second composition, IBA had four levels (0, 0.05, 0.1, 0.15 mg l-1 and TDZ

Agrobacterium-mediated transformation and direct shoot regeneration in Iranian tomato (Solanum lycopersicum L.) cultivar Falat- CH

International Nuclear Information System (INIS)

Kauser, N.; Khan, S.

2016-01-01

Falat CH is an important commercial tomato cultivar being used in Iran. In this article an optimized protocol with increased transformation and regeneration rate for this tomato variety is reported. Several explants including cotyledon, leaf and hypocotyl were evaluated for direct shoot formation and the effect of various combinations of BAP, Zeatin, IAA and IBA were studied. It is the first report on two cytokinins BAP and Zeatin in various combinations to evaluate the synergetic effect of cytokinins on direct shoot regeneration. The synergetic combination of 1.5mg/l BAP, 0.5 mg/l Zeatin and 0.2 mg/l IAA was considered as the best treatment which resulted in higher plant regeneration rates from all of the explants over previous reported methods. Using the best regeneration treatment obtained, the HBsAg gene was transferred into the tomato explants using Agrobacterium mediated transformation technique Percent of the putative transgenic plants regenerated was 68%. PCR of putative transformed plants showed that 87.1% of regenerated plants amplified nptII and HBsAg gene when specifically designed primers were used giving a final transformation rate of 34.85%. (author)

Evaluación y selección de un protocolo para la regeneración in vitro de la variedad de tomate Unapal-Arreboles Evaluation and selection of a protocol for in vitro regeneration of tomato variety Unapal-Arreboles

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Hernando Ramírez

2009-01-01

Full Text Available Se compararon tres medios de cultivo para seleccionar el apropiado para la regeneración in vitro de la variedad de tomate UNAPAL-Arreboles. Se evaluó la producción de callos/explante, brotes/explante y plántulas /explante. El análisis de varianza y la prueba de Duncan permitieron concluir que el medio M3 presentó los mejores promedios para las variables evaluadas.The main objective of this research was to carried out the comparison of three culture medium of major use for in vitro regeneration of tomato, to select the more appropriate medium for in vitro regeneration of tomato, variety UNAPAL-Arreboles. The analysis of variance and media comparison was made based on the callus, shoots and plantlets production. The highest efficiency of callus, shoots and plantlets production was obtained with the M3 media.

Shoot regeneration of callus culture from irradiated sheed of piper nigrum L by gamma rays

International Nuclear Information System (INIS)

Ishak; Hutabarat, D.

1988-01-01

Shoot regeneration was obtained from callus that induced by irradiated seed with 25 and 50 Gy of gamma-rays and then on M.S. medium containing NAA 1 ppm and 2-ip 0.5 ppm. Irradiated seed with a dose of 25 Gy produced normal root and failed to produce shoot, but rice callus. Irradiated seed with a dose of 50 Gy pruduce callus only. Shoot differentiation occured after the callus were cultured on M.S., medium containing 2-ip 1 ppm and Kinetin 2.5 ppm. (authors). 9 refs, 3 figs

In vitro Embryogenesis Derived from Shoot Tips in Mass Propagation of Two Selected-Clones of Phalaenopsis

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Budi WINARTO

2016-09-01

Full Text Available Phalaenopsis is of high economic value and market demand in Indonesia; however, orchid products are mostly imported from other countries. ‘Kristina Dwi’ (KD 69.274 and ‘Dedeh’ (D 802.28 are two selected clones with high potential utilized and developed commercially. To support their commercialization, a reliable in vitro propagation protocol is essential.  In the current study, an in vitro mass propagation protocol for KD 69.274 and D 802.28 clones was successfully established using shoot tips as explant sources. A high number of embryos, up to 8.2 embryos per explant, with 58.5% explant regeneration, and 3.5 regenerated-explants in average were regenerated from shoot tips of KD 69.274 clone cultured on  half-strength Murashige and Skoog (MS medium, with full strength micro, Fe-chellate and vitamin containing 0.5 mg/L thidiazuruon (TDZ and 0.25 mg/L N6-benzyladenine (BA. The initial embryos were proliferated by culturing embryos individually on half-strength MS medium with 0.13 mg/L TDZ and 0.25 mg/L BA and resulted in high embryo regeneration up to 91.4%, with 10.2 embryos per explant and no embryo browning. The embryos were multiplied under periodical subcultures of 3 months each, resulting in gradual increasing number of embryos from the first subculture till the fifth subculture, with 23.6 embryos produced, then declined afterward. The embryos were easily germinated on half-strength MS medium with full strength of vitamin and hormone free, with 73.9% embryo germination and 14.9 germinated embryos. Healthy plantlets were stimulated on the same medium with 2 g/L activated charcoal (AC and successfully acclimatized on Cycas rumphii bulk, with 88.3% survival plantlets. Finally, it can be summarized that a new in vitro mass propagation protocol, as new alternative choice for Phalaenopsis propagation, was successfully established.

Assessment of the potentiality of TDZ on multiple shoot induction in Bauhinia tomentosa L., a woody legume.

Science.gov (United States)

Naz, Ruphi; Anis, M; Aref, I M

2012-12-01

An efficient and reproducible protocol for in vitro multiplication of Bauhinia tomentosa L. was developed. Multiple shoots were regenerated from cotyledonary node and stem nodal segments excised from in vitro raised seedlings on Murashige and Skoog (MS) medium supplemented with different concentrations (0.1, 0.3, 0.5, 0.8 and 1.0 μM) of thidiazuron (TDZ). The maximum response (62.6%) was recorded on MS medium amended with 0.8 μM TDZ. A long exposure to TDZ for 8 weeks showed abnormalities such as fasciation and compact shoots formation. To avoid adverse effects of prolonged exposure to TDZ in long-term establishment, the culture were transferred to TDZ free MS medium for further multiplication and elongation. The highest number of shoots and shoot length were recorded at the end of fourth subculture passage. Ex vitro rooting was achieved when the basal cut end of regenerated shoots were dipped in 200 μM indole-3-butyric acid (IBA) for half an hour followed by their transplantation in plastic pots filled with sterile Soilrite™ where 60% plantlets grew well and all expressed normal development.

In vitro regeneration of cocona (Solanum sessiliflorum, Solanaceae) cultivars for commercial production.

Science.gov (United States)

Schuelter, A R; Grunvald, A K; Amaral, A T; da Luz, C L; Luz, C L; Gonçalves, L M; Stefanello, S; Scapim, C A

2009-08-11

Cocona (Solanum sessiliflorum Dunal) is a solanaceous shrub native to the Amazon region that produces an edible fruit. This species has numerous advantages, particularly a high nutritional value and productivity. However, due to irregular germination and rapid loss of seed viability, there are few plantations for production on a large scale. Development of alternative propagation strategies is essential for the production of homogeneous seedlings of genotypes with superior agronomic performance. We developed techniques for in vitro regeneration of the cocona varieties Santa Luzia and Thaís for large-scale production of healthy plantlets. Twenty days after seeding, seedling segments germinated in vitro were used as explant sources. Three successive experiments were performed: one to test the effect of the explant source and combinations of two growth regulators, auxin (indole acetic acid, IAA) and kinetin (KIN), on the morphogenetic response; another to investigate the effect of the combination of growth regulators on the morphogenetic response of hypocotyl segments, and another to evaluate how sucrose concentration affects the development of adventitious shoots. The best shoot induction was obtained using hypocotyl segments and stem apices, while rhizogenesis was greatest in leaves with a petiole. The number of adventitious shoots per explant on hypocotyl segments increased with 10 and 20 mg/L KIN, combined with 0.02 mg/L IAA in the variety Santa Luzia. Sucrose combined with these growth regulator levels increased the average number of calli; these were optimally produced when 45 g/L sucrose and 0.01 mg/L IAA + 20 mg/L KIN were applied. Only sucrose concentration influenced shoot proliferation in the two S. sessiliflorum varieties, with a maximum at 17.5 g/L.

Effects of GhWUS from upland cotton (Gossypium hirsutum L.) on somatic embryogenesis and shoot regeneration.

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Xiao, Yanqing; Chen, Yanli; Ding, Yanpeng; Wu, Jie; Wang, Peng; Yu, Ya; Wei, Xi; Wang, Ye; Zhang, Chaojun; Li, Fuguang; Ge, Xiaoyang

2018-05-01

The WUSCHEL (WUS) gene encodes a plant-specific homeodomain-containing transcriptional regulator, which plays important roles during embryogenesis, as well as in the formation of shoot and flower meristems. Here, we isolated two homologues of Arabidopsis thaliana WUS (AtWUS), GhWUS1a_At and GhWUS1b_At, from upland cotton (Gossypium hirsutum). Domain analysis suggested that the two putative GhWUS proteins contained a highly conserved DNA-binding HOX domain and a WUS-box. Expression profile analysis showed that GhWUSs were predominantly expressed during the embryoid stage. Ectopic expression of GhWUSs in Arabidopsis could induce somatic embryo and shoot formation from seedling root tips. Furthermore, in the absence of exogenous hormone, overexpression of GhWUSs in Arabidopsis could promote shoot regeneration from excised roots, and in the presence of exogenous auxin, excised roots expressing GhWUS could be induced to produce somatic embryo. In addition, expression of the chimeric GhWUS repressor in cotton callus inhibited embryogenic callus formation. Our results show that GhWUS is an important regulator of somatic embryogenesis and shoot regeneration. Copyright © 2018 Elsevier B.V. All rights reserved.

In vitro shoot regeneration of fenugreek (Trigonella foenum-graecum ...

African Journals Online (AJOL)

Jane

2010-10-18

Oct 18, 2010 ... sprouts are good source of protein, mineral and vitamin C. (Khan et al. ... hypocotyl explants were isolated from 8 - 10 days old germinated seedlings and ..... TJV (1993). Transformation and regeneration of two cultivars of pea.

In vitro propagation, ex vitro rooting and leaf micromorphology of Bauhinia racemosa Lam.: a leguminous tree with medicinal values.

Science.gov (United States)

Sharma, Udit; Kataria, Vinod; Shekhawat, N S

2017-10-01

A micropropagation system for Bauhinia racemosa Lam. was developed involving axillary shoot proliferation and ex vitro rooting using nodal explants obtained from mature tree. MS medium with 3.0 mg l -1 BA (6-benzyladenine) was optimum for shoot bud induction. For shoot multiplication, mother explants were transferred repeatedly on medium containing low concentration of BA (0.75 mg l -1 ). Number of shoots was increased up to two passages and decreased thereafter. Shoot multiplication was further enhanced on MS medium containing 0.25 mg l -1 each of BA and Kin (Kinetin) with 0.1 mg l -1 of NAA (α-naphthalene acetic acid). Addition of 0.004 mg l -1 TDZ (thidiazuron) increased the rate of shoot multiplication and 21.81 ± 1.26 shoots per culture vessel were obtained. In vitro regenerated shoots were rooted under ex vitro conditions treated with 400 mg l -1 IBA (indole-3-butyric acid) for 7 min on sterile soilrite. After successful hardening in greenhouse, ex vitro rooted plants were transferred to the field conditions with ≈85% of survival rate. Micromorphological changes were observed on leaf surface i.e. development of vein density and trichomes and stomatal appearance, when plants were subjected to environmental conditions. This is the first report on in vitro regeneration of B. racemosa from mature tree.

The study of ascorbate peroxidase, catalase and peroxidase during in vitro regeneration of Argyrolobium roseum.

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Habib, Darima; Chaudhary, Muhammad Fayyaz; Zia, Muhammad

2014-01-01

Here, we demonstrate the micropropagation protocol of Argyrolobium roseum (Camb.), an endangered herb exhibiting anti-diabetic and immune-suppressant properties, and antioxidant enzymes pattern is evaluated. Maximum callogenic response (60 %) was observed from leaf explant at 1.0 mg L(-1) 1-nephthalene acetic acid (NAA) and 0.5 mg L(-1) 6-benzyl aminopurine (BA) in Murashige and Skoog (MS) medium using hypocotyl and root explants (48 % each). Addition of AgNO3 and PVP in the culture medium led to an increase in callogenic response up to 86 % from leaf explant and 72 % from hypocotyl and root explants. The best shooting response was observed in the presence of NAA, while maximum shoot length and number of shoots were achieved based on BA-supplemented MS medium. The regenerated shoots were rooted and successfully acclimatized under greenhouse conditions. Catalase and peroxidase enzymes showed ascending pattern during in vitro plant development from seed while ascorbate peroxidase showed descending pattern. Totally reverse response of these enzymes was observed during callus induction from three different explants. During shoot induction, catalase and peroxidase increased at high rate while there was a mild reduction in ascorbate peroxidase activity. Catalase and peroxidase continuously increased; on the other hand, ascorbate peroxidase activity decreased during root development and acclimatization states. The protocol described here can be employed for the mass propagation and genetic transformation of this rare herb. This study also highlights the importance and role of ascorbate peroxidase, catalase, and peroxidase in the establishment of A. roseum in vitro culture through callogenesis and organogenesis.

Cryopreservation of in vitro -grown shoot tips of apricot ( Prunus ...

African Journals Online (AJOL)

In vitro grown apricot (Prunus armeniaca L.) cv. El-Hamawey shoot tips were successfully cryopreserved using an encapsulation-dehydration procedure. Shoot tips were encapsulated in calcium-alginate beads before preculture on woody plant (WP) medium supplemented with different sucrose concentrations; 0.1, 0.3, 0.5, ...

Effects of NAA and BAP, double-layered media, and light distance on in vitro regeneration of Nelumbo nucifera Gaertn. (lotus), an aquatic edible plant.

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Mahmad, Noraini; Taha, Rosna Mat; Othman, Rashidi; Saleh, Azani; Hasbullah, Nor Azlina; Elias, Hashimah

2014-01-01

In vitro direct regeneration of Nelumbo nucifera Gaertn. was successfully achieved from immature explants (yellow plumule) cultured on a solid MS media supplemented with combinations of 0.5 mg/L BAP and 1.5 mg/L NAA which resulted in 16.00 ± 0.30 number of shoots per explant and exhibited a new characteristic of layered multiple shoots, while normal roots formed on the solid MS basal media. The double-layered media gave the highest number of shoots per explant with a ratio of 2 : 1 (liquid to solid) with a mean number of 16.67 ± 0.23 shoots per explant with the formation of primary and secondary roots from immature explants. In the study involving light distance, the tallest shoot (16.67 ± 0.23 mm) obtained from the immature explants was at a light distance of 200 mm from the source of inflorescent light (1000 lux). The plantlets were successfully acclimatized in clay loam soil after 8 months being maintained under in vitro conditions.

In vitro propagation of chungah (caralluma tuberculata n.e. brown)

International Nuclear Information System (INIS)

Rehman, R.

2014-01-01

In vitro propagation of Caralluma tuberculata (Chungah) was developed from shoot tip and meristem explants. C. tuberculata is an imperative medicinal plant comprising antidiabetic and anticancer properties. The explants were inoculated on Murashige and Skoog (MS) medium containing different plant growth regulators. Presence of BA or Kin alone in the MS medium did not favor regeneration of shoot from both explants. However, addition of 2,4-dichlorophenoxy acetic acid (2,4-D), gibberellic acid (GA3) and thidiazuron (TDZ) along with 6-benzyl amino purine (BA) or kinetin (Kin) in the medium exhibited significant percentage response, number of shoots per explant and shoot length. Maximum shooting response (53.3+-5.77% from meristem and shoot tip explants each) with highest number of shoots per explant (5.33+-2.08 and 5.6+-2.52 from meristem and shoot tip explants, respectively) were observed at 13.32 micro mol BA along with 2.26 micro mol 2,4-D, 2.89 micro mol GA3 and 9.08*10-3 micro mol TDZ. Replacing BA with kin showed less shoot regeneration response and number of shoots per explant, however, shoots length markedly increased in the presence of Kin. The regenerated plants were successfully rooted and acclimatized in ex vitro condition. The protocol described here can be used for fast multiplication of this endangered herb and genetic transformation. (author)

In Vitro Propagation of Sambong (Blumea balsamifera Linn.).

Science.gov (United States)

Soriano, Thelma L; Cangao, Evangelina C

2016-01-01

Terminal shoot tips of sambong (Blumea balsamifera Linn.) are cultured to initiate and regenerate shoots on Murashige and Skoog (MS) medium containing 1.0 mg/L benzyl adenine (BA). After 1 month, shoots, usually 4.5 cm long are separated and subcultured for multiplication. Regenerated shoots, about 6 cm long are rooted on MS medium supplemented with 1.0 mg/L naphthalene acetic acid (NAA). Exposure of shoots to high humidity for the first 2 weeks and equal proportion (1:1:1) of sterile sand, compost, and coir dust as potting mix favors the development of whole sambong plants. Young shoots from in vitro-derived sambong plants could also be used for propagation.

Resposta à regeneração e crescimento de brotos in vitro de Kielmeyera coriacea quando influenciado por diferentes concentrações dos sais e de sacarose In vitro regeneration and growth response of Kielmeyera coriacea shoots when affected by salts and sucrose concentrations

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José Eduardo Brasil Pereira Pinto

1996-04-01

Full Text Available Avaliaram-se diferentes concentrações de sacarose e sais do MS (MURASHIGE & SKOOG, J 962 na regeneração e crescimento de brotos de Kielmeyera coriacea in vitro. A redução e o aumento da quantidade de sacarose partindo de 30 e 45g/L, respectivamente, causou um decréscimo no número total de brotos. O uso integral ou da metade das concentrações dos sais do MS proporcionou maiores taxas de multiplicação. A maior porcentagem de brotos com mais de 1,0cm de comprimento pôde ser obtida empregando-se 30 g/l de sacarose associado a concentração 1/1 do MS. A porcentagem média de brotos decresceu linearmente com a diluição salina.Trials were carried out to test sucrose and salt concentrations added to growth medium on regeneration and growth response of Kielmeyera coriacea shoots. The reduction and increase of sucrose of 30 and 45g/l, rerspectively, caused a decrease in total number of shoots. A higher shoots percentage of with more than 1.0cm, occurred on media with 30g/L of sucrose. The use of total force (1/1 and half strength (1/2 of MS salt proportioned a higher shoot ratio and growth.

Plant regeneration from cotyledons of mature soybean (Glycine max L.) Wilis cultivar using gamma rays

International Nuclear Information System (INIS)

Hutabarat, D.; Ratna, R.

1999-01-01

Soybean Wilis cultivar was efficiently regenerated in vitro via somatic embryogenesis. Cotyledonary explants were excised from mature germinating seeds. Seeds were germinated on agar solution and on B5 medium enriched with 5 ppm BA, 0.25 ppm BA, 0.25 ppm IBA and 500 ppm casein hydrolyzate. Cotyledonary nodes from both germinating seeds were excised and cultured on B5 medium enriched with 5 ppm BA, 0.25 ppm IBA and 500 ppm casein hydrolyzate. Age of seedlings had a remarkable influence on shoot regeneration. Cotyledon from seeds germinated on agar solution with light gave better result in shoot regeneration compare with those germinated in darkness. The highest number of regenerants per explants (5 shoots) was produced by cotyledon from seeds germinated on B5 medium enriched with 5 ppm IBA and 500 ppm casein hydrolyzate in darkness. The seeds of soybean were exposed to gamma-rays doses 10 Gy then germinated on B5 medium enriched with 5 ppm BA, 0.25 ppm IBA and 500 ppm casein hydrolyzate did not improve the number of plant regeneration. Only 5-day-old seedlings from seeds were exposed to gamma-rays dose 30 Gy could improve the number of shoot regeneration, one of the cotyledonary node treated produced 21 regeneration shoots

Micropropagation of Vaccinium sp. by in vitro axillary shoot proliferation.

Science.gov (United States)

Litwińczuk, Wojciech

2013-01-01

The Vaccinium genus contains several valuable fruit and ornamental species, among others: highbush blueberry (Vaccinium × corymbosum L.), cranberry (Vaccinium macrocarpon Ait.), and lingonberry (Vaccinium vitis-idaea L.). In some most popular and valuable cultivars, the conventional propagation methods, exploiting hard or soft wood cuttings, are inefficient. The demand for nursery plants could be fulfilled only by micropropagation. In principle cultivars are propagated in vitro through similar three-stage method, based on subculture of shoot explants on different culture media supplemented with IAA (0-4 mg/L) and 2iP (5-10 mg/L), and rooting shoots in vivo. The obtained plantlets are transferred to peat substrate and grown in the glasshouse until the end of growing period. The development of adventitious shoots should be monitored and controlled during in vitro stages. Many clones have specific requirements for growing conditions and/or are recalcitrant.

IN VITRO REGENERATION OF THREE CHRYSANTHEMUM (Dendrathema grandiflora VARIETIES “VIA” ORGANOGENESIS AND SOMATIC EMBRYOGENESIS

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Elizabeth Hodson de Jaramillo

2008-09-01

Full Text Available Chrysanthemum (Dendrathema grandiflora has a high demand in the Colombian and international cut flower markets.Since commercial production of this ornamental species is strongly affected by fungal diseases such as chrysanthemumwhite rust (Puccinia horiana, high doses of fungicides are being used posing increased environmental and commercialcosts. Assessment of in vitro regeneration systems from leaf discs was a first step in developing a plant genetic transformationprotocol to obtain fungi-resistant plants. Leaf discs of White Albatross, Yellow Albatross, and Escapade varieties wereestablished in vitro on MS medium supplemented with NAA (0 - 4.83 μM and BAP (0 - 13.32 μM alone and incombination. Leaf discs were also cultured on MumB medium containing 2,4-D (0 - 4.52 μM for 7, 14, and 21 days priorto their transferral to a 2,4-D free MumB medium. Regenerated shoots were individualized, rooted, and hardened. Resultsshow that MS with 4.83 μM NAA + 4.44 μM BAP and 4.83 μM NAA + 13.32 μM BAP induce organogenesis, and MumBwith 2.26 μM 2,4-D induces somatic embryogenesis on all three varieties, with exposition periods to 2,4-D of 14 days forWhite Albatross and 21 days for Yellow Albatross and Escapade. Shoot development from somatic embryos was observedin the three varieties when cultured on a 2,4-D free MumB medium. Spontaneous rooting was recorded in 85% of the shootsthus facilitating hardening and successful transfer to soil.

Direct adventitious shoot bud formation on hypocotyls explants in Millettia pinnata (L.) Panigrahi- a biodiesel producing medicinal tree species.

Science.gov (United States)

Nagar, Durga Singh; Jha, Suman Kumar; Jani, Jigar

2015-04-01

A reproducible protocol developed for in vitro regeneration of Milletia pinnata using hypocotyl segments. Multiple shoots were induced from hypocotyl explants through direct adventitious shoot bud regeneration. The proximal end of hypocotyls was responsive for shoot bud induction. Silver nitrate and adenine sulphate had a positive effect on shoot bud induction and elongation. The maximum response and number of shoot bud produced in media supplemented with 8.88 μM BAP with 108.6 μM adenine sulphate and 11.84 μM silver nitrate. Elongated shoots were harvested and successful rooting of microshoots achieved on MS media supplemented with 9.84 μM IBA, with 81.1 % rooting. Remaining shoot buds sub-cultured for further multiplication and elongation. Each subculture produced eight to nine elongated microshoots up to four subcultures. The rooted microshoots were successfully hardened and transferred to field.

Regeneration of multiple shoots from transgenic potato events facilitates the recovery of phenotypically normal lines: assessing a cry9Aa2 gene conferring insect resistance

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Jacobs Jeanne ME

2011-10-01

Full Text Available Abstract Background The recovery of high performing transgenic lines in clonal crops is limited by the occurrence of somaclonal variation during the tissue culture phase of transformation. This is usually circumvented by developing large populations of transgenic lines, each derived from the first shoot to regenerate from each transformation event. This study investigates a new strategy of assessing multiple shoots independently regenerated from different transformed cell colonies of potato (Solanum tuberosum L.. Results A modified cry9Aa2 gene, under the transcriptional control of the CaMV 35S promoter, was transformed into four potato cultivars using Agrobacterium-mediated gene transfer using a nptII gene conferring kanamycin resistance as a selectable marker gene. Following gene transfer, 291 transgenic lines were grown in greenhouse experiments to assess somaclonal variation and resistance to potato tuber moth (PTM, Phthorimaea operculella (Zeller. Independently regenerated lines were recovered from many transformed cell colonies and Southern analysis confirmed whether they were derived from the same transformed cell. Multiple lines regenerated from the same transformed cell exhibited a similar response to PTM, but frequently exhibited a markedly different spectrum of somaclonal variation. Conclusions A new strategy for the genetic improvement of clonal crops involves the regeneration and evaluation of multiple shoots from each transformation event to facilitate the recovery of phenotypically normal transgenic lines. Most importantly, regenerated lines exhibiting the phenotypic appearance most similar to the parental cultivar are not necessarily derived from the first shoot regenerated from a transformed cell colony, but can frequently be a later regeneration event.

High Frequency Multiple Shoot Induction of Curculigo orchioides Gaertn.: Shoot Tip V/S Rhizome Disc

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K. S. Nagesh

2008-09-01

Full Text Available Curculigo orchioides Gaertn. is an endangered medicinal plant with anticancer properties. The rhizome and tuberous roots of the plant have been used extensively in India in indigenous medicine. Due to its multiple uses, the demand for Curculigo orchioides is constantly on the rise; however, the supply is rather erratic and inadequate. Destructive harvesting, combined with habitat destruction in the form of deforestation has aggravated the problem. The plant is now considered ‘endangered’ in its natural habitat. Therefore, the need for conservation of this plant is crucial. Here, we describe a successful protocol for multiple shoot induction of C. orchioides using shoot tip and rhizome disc. We find that proximal rhizome discs are optimal for high frequency shoot bud formation than shoot tip and distal rhizome disc. We observed a synergistic effect between 6-benzylaminopurine (BAP and kinetin (KN (each at 1 mg/L on the regeneration of shoot buds from proximal rhizome disc than shoot tip explant. Optimum root induction was achieved on half-strength MS liquid medium supplemented with 1 mg/L of indole-3-butyric acid (IBA. The in vitro raised plantlets were acclimatized in green house and successfully transplanted to natural condition with 90% survival.

Studies on Callus Induction and Regeneration of Medicinal Plant Chicory (Cichorium intybus L. from Leaf and Petiole Explants

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H. Hadizadeh

2016-07-01

Full Text Available Introduction: Chicory (Cichorium intybus L. belongs to Asteraceae family is commonly known as witloof chicory. The leaves and the roots of this medicinal plant are edible and commonly used as salad. Some varieties are also cultivated as coffee substitute after roasting the roots. All parts of the plant contain these volatile oils, with the majority of the toxic components concentrated in the plant's root. In folk medicine, the plant is used for the treatment of diarrhea, spleen enlargement, fever, and vomiting. Antihepatotoxic activity on damaged rat’s liver sections and anti-bacterial activity of this crop has been recently reported. In vitro regeneration from leaf explants with various hormonal combinations has been reported previously. Moreover, in vitro regeneration of Chicory from cotyledon explants using different combinations of plant growth regulators has been studied. Also, a protocol for the regeneration of plantlets from leaf and petiole explants of witloof chicory has been developed. The aim of the present investigation was optimization of callus induction and shoot regeneration from leaf and petiole tissues of Chicory (Esfahan genotype. Materials and Methods: In this investigation, Esfahan genotype was used for callus induction and direct shoot regeneration. Seeds were first washed with running tap water for 30 min then seeds were surface sterilized by dipping in 70% ethanol for 90 s and rinsed with sterile distilled water, followed by immersing in 5% sodium hypochlorite solution for 25 min and thereafter rinsed for 30 min with sterile distilled water. The basal medium used in this investigation was MS. For shoot regeneration, leaf and petiole explants (5 mm segments were excised from 4-week-old sterile seedlings and cultured on MS medium containing different combinations of NAA / BA and KIN / BA in two separate experiments. Experiments were performed factorial based on completely randomized design. Cultures were incubated at 25�

In vitro regeneration of Turkish dwarf chickling (Lathyrus cicera L ...

African Journals Online (AJOL)

STORAGESEVER

2008-06-17

Jun 17, 2008 ... Key words: Immature zygotic embryos, dwarf chickling, Lathyrus cicera L., axillary shoot regeneration, ... are widely used as animal feed in green and dry form or ..... Hanbury CD, White CL, Mullan BP, Siddique KHM (2000).

Zinc tolerance and accumulation in stable cell suspension cultures and in vitro regenerated plants of the emerging model plant Arabidopsis halleri (Brassicaceae).

Science.gov (United States)

Vera-Estrella, Rosario; Miranda-Vergara, Maria Cristina; Barkla, Bronwyn J

2009-03-01

Arabidopsis halleri is increasingly employed as a model plant for studying heavy metal hyperaccumulation. With the aim of providing valuable tools for studies on cellular physiology and molecular biology of metal tolerance and transport, this study reports the development of successful and highly efficient methods for the in vitro regeneration of A. halleri plants and production of stable cell suspension lines. Plants were regenerated from leaf explants of A. halleri via a three-step procedure: callus induction, somatic embryogenesis and shoot development. Efficiency of callus proliferation and regeneration depended on the initial callus induction media and was optimal in the presence of 1 mg L(-1) 2,4-dichlorophenoxyacetic acid, and 0.05 mg L(-1) benzylaminopurine. Subsequent shoot and root regeneration from callus initiated under these conditions reached levels of 100% efficiency. High friability of the callus supported the development of cell suspension cultures with minimal cellular aggregates. Characterization of regenerated plants and cell cultures determined that they maintained not only the zinc tolerance and requirement of the whole plant but also the ability to accumulate zinc; with plants accumulating up to 50.0 micromoles zinc g(-1) FW, and cell suspension cultures 30.9 micromoles zinc g(-1) DW. Together this work will provide the experimental basis for furthering our knowledge of A. halleri as a model heavy metal hyperaccumulating plant.

Plant Regeneration and Cellular Behaviour Studies in Celosia cristata Grown In Vivo and In Vitro

Science.gov (United States)

Taha, Rosna Mat; Wafa, Sharifah Nurashikin

2012-01-01

Tissue culture studies of Celosia cristata were established from various explants and the effects of various hormones on morphogenesis of this species were examined. It was found that complete plant regeneration occurred at highest percentage on MS medium supplemented with 2.0 mg/L NAA and 1.5 mg/L BAP, with the best response showed by shoot explants. In vitro flowering was observed on MS basal medium after six weeks. The occurrence of somaclonal variation and changes in cellular behavior from in vivo and in vitro grown plants were investigated through cytological studies and image analysis. It was observed that Mitotic Index (MI), mean chromosome numbers, and mean nuclear to cell area ratio of in vitro root meristem cells were slightly higher compared to in vivo values. However, in vitro plants produced lower mean cell areas but higher nuclear areas when compared to in vivo plants. Thus, no occurrence of somaclonal variation was detected, and this was supported by morphological features of the in vitro plants. PMID:22593677

Direct regeneration and efficient in vitro root development studies in lentil (lens culinaris medik)

International Nuclear Information System (INIS)

Sultana, T.; Majeed, N.; Naqvi, S.

2016-01-01

Lentil is a self-pollinating annual crop with increasing demand all over the world due to its high protein content and easy digestibility. However, like many other crops lentil too needs improvement for which conventional as well as biotechnological tools are to be employed. This study was aimed at development of tissue culture protocol especially targeting improved root development to ensure their establishment in soil in order to use their potential towards genetic manipulation. Two Pakistani lentil cultivars, Masoor-2002 and Manshera-89 were used to obtain cotyledonary nodes, epicotyl and hypocotyl explants. The explants were cultured on shoot regeneration medium containing different concentration of kinetin, BAP and tyrosine with the addition of GA3, with or without charcoal for shoot development. Masoor-2002, showed the highest frequency of shoot development on MS medium containing 5.5 mg/L tyrosine, 0.25 mg/L kinetin, 1.0 mg/L BAP, 0.1 mg/L GA3, using cotyledonary node as explant. The addition of 2 g/L of charcoal in shoot medium resulted in healthier plants, but the number of shoots were reduced. Regarding the effect of age of explants on regeneration frequency, cotyledonary nodes of age 4-6 days had higher regeneration potential. Well-developed shoots were shifted to rooting medium containing different concentration of auxin with or without charcoal. Healthier and more roots were observed on medium containing 4 mg/L IAA with addition of 2 g/L charcoal. Plants were better established (70% survival) in a soil mix containing perlite, vermiculite and peat moss in 1:1:1 ratio. (author)

In vitro propagation of Stevia rebaudina plants using multiple shoot culture.

Science.gov (United States)

Nepovím, A; Vanek, T

1998-12-01

A multiple shoot culture was induced from nodal segments on MS medium containing half concentration of macroelements, 1% sucrose, and supplemented with NAA (0.01 mg/l). A bioreactor with hormone-free MS medium (300 ml) was inoculated with 1.5 g of the multiple shoot culture and cultivated for a month. The cultivating process of the multiple shoot culture in the bioreactor and the transfer into ex vitro conditions took about 8-9 weeks and produced approx. 600 new seedlings, that could be transferred from greenhouse to field conditions.

Field evaluation of regenerated plants by somatic embryogenesis from shoots apexes of axillary buds in ´Navolean’ (Musa spp., AAB.

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Jorge López

2005-04-01

Full Text Available The use of shoots apexes from axilary buds for callus induction with embryogenic structures in plantain ‘Navolean’ (Group AAB permitted to develop a plant regeneration method through out somatic embryogenesis. In order to know the phenotypic variants that may be produced with the previously mentioned method , 1000 plants were planted in field conditions in comparison to those coming from somatic embryos obtained from multibuds as initial explants and organogenesis-derived plants (shoot tipsand conventionally derived plants (corms, during two growing cycles. The main morphological characters and yield components were evaluated. The total frequency of somaclonal variation during the first growing cycle in plants coming from somatic embryos obtained from shoots apexes from axilary buds as initial explants were 1.1%, and 8,6% in regenerated plants from somatic embryos obtained from multi-buds as initial explants. Later, in this same growing cycle, plants regenerated from somatic embryos (both sources showed a similar performance between them and they were significantly superior in all evaluated variants in comparison to corm-derived plants. In the second growing cycle, significant differences were not observed in yield components of suckers from evaluated plants, in spite of the propagation method used. With regard to somaclonal variation, the best performance was obtained with shoots apexes from axilary buds as explants. Finally, the feasibility of using the new method was shown. Key words: embryogenic cell suspensions, somaclonal variation

In vitro establishment of Vasconcellea x helbornii (Badillo Badillo shoots

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Mónica Jadán Guerrero

2016-04-01

Full Text Available Babaco [Vasconcellea x helbornii (Badillo Badillo] is a specie of commercial importance in Ecuador. The present work aimed to establish in vitro shoots of axillary buds from mother plants of babaco kept in greenhouse. Carbendazim and the biostimulant (GERMO-TB01 were applied to the plants. For the disinfection of the explants, three concentrations of Sodium Hypochlorite (1, 1.5 and 2% were evaluated during two times (5 and 10 minutes. In addition, the effect of the use of Gentamicin 50 mg l-1 and Streptomycin 25 mg l-1 in the culture medium was determined. The best results were achieved by using 1.5% Sodium Hypochlorite for 10 minutes and immersion in a solution with both antibiotics for 3 hours. A 68.5% in vitro establishment of the shoots was achieved at 21 days of culture. The results will contribute to in vitro mass propagation of this hybrid.   Key words: antibiotic mixture, babaco, disinfection, micropropagation

Plant regeneration of Brassica oleracea subsp. italica (Broccoli) CV ...

African Journals Online (AJOL)

STORAGESEVER

2009-06-03

Jun 3, 2009 ... Department of Agriculture Technology, Faculty of Agriculture, Universiti Putra Malaysia, 43400 Serdang, Selangor Darul. Ehsan, Malaysia. Accepted 20 March, 2009. Hypocotyls and shoot tips were used as explants in in vitro plant regeneration of broccoli (Brassica oleracea subsp.italica) cv. Green Marvel.

Thidiazuron: A potent cytokinin for efficient plant regeneration in Himalayan poplar (Populus ciliata Wall. using leaf explants

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Gaurav Aggarwal

2012-11-01

Full Text Available Populus species are important resource for certain branches of industry and have special roles for scientific study on biological and agricultural systems. The present investigation was undertaken with an objective of enhancing the frequency of plant regeneration in Himalayan poplar (Populus ciliata Wall.. The effect of Thiadizuron (TDZ alone and in combination with adenine and α-Naphthalene acetic acid (NAA were studied on the regeneration potential of leaf explants. A high efficiency of shoot regeneration was observed in leaf (80.00% explants on MS basal medium supplemented with 0.024 mg/l TDZ and 79.7 mg/l adenine. Elongation and multiplication of shoots were obtained on Murashige and Skoog (MS basal medium, containing 0.5 mg/l 6. Benzyl aminopurine (BAP + 0.2mg/l Indole 3-acetic acid (IAA + 0.3 mg/l Gibberellic acid (GA3. High frequency root regeneration from in vitro developed shoots was observed on MS basal medium supplemented with 0.10 mg/l Indole 3-butyric acid(IBA. Maximum of the in vitro rooted plantlets were well accomplished to the mixture of sand: soil (1:1 and exhibited similar morphology with the field plants. A high efficiency plant regeneration protocol has been developedfrom leaf explants in Himalayan poplar (Populus ciliata Wall..

Essential Oil Composition of the Different Parts and In Vitro Shoot Culture of Eryngium planum L.

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Anna Kurowska

2011-08-01

Full Text Available The essential oils obtained by hydrodistillation from the different parts (inflorescence, stalk leaves, rosette leaves and root as well as from in vitro shoot culture of Eryngium planum L. were analyzed by GC-FID-MS in respect to their chemical composition. The different parts of E. planum and in vitro shoots showed different yields. The part with higher amount was the inflorescences, followed by the stalk leaves and in vitro shoots, rosette leaves and finally roots. The essential oils obtained from rosette leaves and in vitro-derived rosettes had totally different composition. Quantitative differences were also found between compounds of intact plant organs. The main components of stalk leaf oil and rosette leaf oil were monoterpene (limonene, α- and β-pinene and sesquiterpene hydrocarbons. In inflorescence oil cis-chrysanthenyl acetate (43.2% was accompanied by other esters (propionate, butanoate, hexanoate and octanoate and numerous oxygenated sesquiterpenes. Root oil and in vitro shoot oil contained mainly (Z-falcarinol and 2,3,4-trimethylbenzaldehyde. This is the first report on the chemical composition of this species.

In vitro regeneration of five wheat genotypes from immature zygotic embryos

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Khokhar, M.I.; Iqbal, M.Z.

2016-01-01

This study examined the ability to induce callus from immature zygotic embryos of five wheat genotypes (Lu 26, WH 543, Zamindar 80, BT-002 and Seher-06) in response to 2, 4 and 6 mg/L of 2,4-dichlorophenoxy acetic acid (2,4-D). Callus induction was most effective (41% averaged across the 5 genotypes) in the presence of 2 mg/L 2,4-D. Callus induction was highest in Lu 26 (34%) followed by WH 543 (33%). Highest percentage shoot formation (33%) from callus was possible on Murashige and Skoog (1962) medium containing 300 mg casein hydrolysate. BT-002 responded best to shoot formation (26%) followed by WH 543 (24%). Under these optimal conditions, callus could form within 7.4 days and shoots within 20.87 days (fastest growth averaged across the 5 genotypes). Zamindar-80 responded best by taking fewest days to initiate callus formation (7.88 days) while Lu 26 took the least amount of time to form shoots (23.25 days). This study provides a rapid and efficient, as well as cultivar-independent protocol for the indirect formation of shoots from callus, the first such report for WH 543, Zamindar 80, BT-002 and Seher-06. This protocol may be a useful protocol for transgenic wheat plants that are derived from the genetic transformation of callus, either by particle bombardment or Agrobacterium-mediated transformation, to produce, for example, insect- or herbicide-resistant plants, since a rapid and effective regeneration protocol is an essential first step for the successful regeneration of transgenic plants. (author)

Organ and plantlet regeneration of Menyanthes trifoliata through tissue culture

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Urszula Adamczyk-Rogozińska

2014-01-01

Full Text Available The conditions for the regeneration of plants through organogenesis from callus tissues of Menyanthes trifoliata are described. The shoot multiplication rate was affected by basal culture media, the type and concentration of cytokinin and subculture number. The best response was obtained when caulogenic calli were cultured on the modified Schenk and Hildebrandt medium (SH-M containing indole-3-acetic acid (IAA 0,5 mg/l and 6-benzyladenine (BA 1 mg/l or zeatin (2 mg/l. Under these conditions ca 7 shoots (mostly 1 cm or more in length per culture in the 5th and 6th passages could be developed. In older cultures (after 11-12 passages there was a trend for more numerous but shorter shoot formation. All regenerated shoots could be rooted on the SH-M medium supplemented with 0.5 mg/l IAA within 6 weeks; 80% of in vitro rooted plantlets survived their transfer to soil.

In vitro regeneration of Basella alba L

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Edney, Norris Allen; Rizvi, Muhammad A.; Rizvi, Narjis F.

1989-01-01

Basella alba L. is a tropical vine used as a vegetable in some Asian and African countries. It has potential as a nontraditional crop for small family farms. A short day plant, it blooms during the fall, provided the temperatures are mild. In the southeastern U.S., the short days of fall are associated with subfreezing temperatures, and plants are killed before blooming. Attempts were made to regenerate the plant using tissue culture techniques. Several trials were conducted with different media, hormones, and explants. It was found that nodal segments on Gamborg medium regenerated shoots. Interaction studies of auxins and cytokinins indicated that its endogeneous auxin content might be high because callus proliferated in almost all treatments and roots initiated even when the medium was not supplemented with an auxin.

Shoot regeneration of limau purut (citrus hystrix) using shoot tip: assessment of calcium gluconate and silver nitrate in overcoming premature leaf senescence

International Nuclear Information System (INIS)

Eng, W.H.; Aziz, M.A.; Sinniah, U.R.

2014-01-01

This study was carried out to establish an optimum In vitro shoot multiplication system using shoot tip explants derived from 7 week-old seedlings of Citrus hystrix. In the first experiment, shoot tips were cultured on Murashige and Skoog (MS) medium supplemented with 0-13.33 mu M 6-benzylaminopurine (BAP) for 8 weeks. Shoot tips cultured on 2.22 mu M BAP produced the highest mean number of shoots (3.42 shoots) but the shoots had low number of leaves (1.14 leaves) due to the occurrence of premature leaf senescence and callus formation. Meanwhile, the medium devoid of BAP produced the lowest mean number of shoots (1.50 shoots) but highest mean number of leaves (5.41 leaves) indicating that BAP was likely responsible for the premature leaf senescence. In order to overcome the occurrence of premature leaf senescence on medium with BAP, a second experiment was carried out whereby shoot tips were cultured on medium containing 2.22 micro M BAP fortified with 2.00, 4.00 and 6.00 mM calcium gluconate (Ca-glu) and a control treatment with 2.22 mu M BAP. The shoot and leaf numbers were increased with the addition of 4.00 and 6.00 mM Ca-glu. The presence of Ca-glu reduced premature leaf senescence and callus formation to some extent. In the third experiment, the addition of silver nitrate (AgNO/sub 3/) at 10-80 micro M in media with 2.22 micro M BAP and 2.22 micro M BAP + 4 mM Ca-glu could totally overcome premature leaf senescence and callus formation. Media supplemented with 2.22 mirco M BAP + 4 mM Ca-glu + 20 micro M AgNOsub 3/ significantly induced among the highest mean number of shoots and highest mean number of leaves per shoot. (author)

Plant regeneration of non-toxic Jatropha curcas—impacts of plant growth regulators, source and type of explants

KAUST Repository

Kumar, Nitish

2011-01-28

Jatropha curcas is an oil bearing species with multiple uses and considerable economic potential as a biofuel plant, however, oil and deoiled cake are toxic. A non-toxic variety of J. curcas is reported from Mexico. The present investigation explores the effects of different plant growth regulators (PGRs) viz. 6-benzyl aminopurine (BAP) or thidiazuron (TDZ) individually and in combination with indole-3-butyric acid (IBA), on regeneration from in vitro and field-grown mature leaf explants, in vitro and glasshouse-grown seedlings cotyledonary leaf explants of non-toxic J. curcas. In all the tested parameters maximum regeneration efficiency (81.07%) and the number of shoot buds per explants (20.17) was observed on 9.08 μM TDZ containing Murashige and Skoog’s (MS) medium from in vitro cotyledonary leaf explants. The regenerated shoot buds were transferred to MS medium containing 10 μM kinetin (Kn), 4.5 μM BAP and 5.5 μM α-naphthaleneacetic acid (NAA) for shoot proliferation. The proliferated shoots could be elongated on MS medium supplemented with 2.25 μM BAP and 8.5 μM IAA. Rooting was achieved when the basal cut end of elongated shoots were dipped in half strength MS liquid medium containing different concentrations and combinations of IBA, IAA and NAA for four days followed by transfer to growth regulators free half strength MS medium supplemented 0.25 mg/l activated charcoal. The rooted plants could be established in soil with more than 90% survival rate.

Plant regeneration of non-toxic Jatropha curcas—impacts of plant growth regulators, source and type of explants

KAUST Repository

Kumar, Nitish; Vijay Anand, K. G.; Reddy, Muppala P.

2011-01-01

Jatropha curcas is an oil bearing species with multiple uses and considerable economic potential as a biofuel plant, however, oil and deoiled cake are toxic. A non-toxic variety of J. curcas is reported from Mexico. The present investigation explores the effects of different plant growth regulators (PGRs) viz. 6-benzyl aminopurine (BAP) or thidiazuron (TDZ) individually and in combination with indole-3-butyric acid (IBA), on regeneration from in vitro and field-grown mature leaf explants, in vitro and glasshouse-grown seedlings cotyledonary leaf explants of non-toxic J. curcas. In all the tested parameters maximum regeneration efficiency (81.07%) and the number of shoot buds per explants (20.17) was observed on 9.08 μM TDZ containing Murashige and Skoog’s (MS) medium from in vitro cotyledonary leaf explants. The regenerated shoot buds were transferred to MS medium containing 10 μM kinetin (Kn), 4.5 μM BAP and 5.5 μM α-naphthaleneacetic acid (NAA) for shoot proliferation. The proliferated shoots could be elongated on MS medium supplemented with 2.25 μM BAP and 8.5 μM IAA. Rooting was achieved when the basal cut end of elongated shoots were dipped in half strength MS liquid medium containing different concentrations and combinations of IBA, IAA and NAA for four days followed by transfer to growth regulators free half strength MS medium supplemented 0.25 mg/l activated charcoal. The rooted plants could be established in soil with more than 90% survival rate.

Evaluation of the capacity for direct regeneration of maize inbreds of the Lancaster selection group

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K. V. Derkach

2013-11-01

Full Text Available In connection with the necessity of bringing elite maize inbreds of the Lancaster germplasm group, which have potential for cultivation in Ukraine, into the system of genetic tranformation, the aim of this investigation is to identify the ability of maize inbreds of this group to regenerate by direct organogenesis and to determine the optimal mineral basis for their nutritional environment using segments of the node area of shoots. As explantats we used sterile 4-day old seedlings of 4 maize inbreds of Lancaster germplasm and model inbred Chi31 exotic germplasm. The seedlings were obtained by germination of sterile seeds in Petri dishes between two layers of moist sterile filter paper at a temperature of 27 ºC in dark conditions. A single 1 cmlong segment was cut from each from each seedling, running from 0.5 cmbefore the node to 0.5 cmafter the node. A cut was made in each segment of the node in order to create a wounded surface. Explantats were planted in a nutrient environment with mineral bases of MS or N6, modified by the addition of 10 mg/l silver nitrate, 100 mg/l casein hydrolyzate, 690 mg/l L-proline, 30 g/l sucrose, 1.0 mg/l 2,4-dychlorphenoksiacetic acid and 0,1 mg/l abscisic acid. Cultivation was carried out at 25–27 ºC in the light. Direct hemogenesis in this environment on the 14th day of cultivation in vitro reached 100% for each line. This meant that all researched lines of Lancaster germplasm and the model line showed a high capacity for direct regeneration through direct hemogenesis, which does not depend on the composition of the mineral content of their nutritional environment. Callus formation was observed in all genotypes on the 14th day of cultivation in vitro and the extent of its formation increased during the following month of cultivation. The callus formation was observed only at the site of the wounded surface. The calluses were transparent. Although green areas appeared in these calluses, they were

STUDIES REGARDING CULTURE MEDIUM INFLUENCE ON IN VITRO REGENERATION FROM WHEAT IMATURE EMBRYOS

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M. DANCI

2008-05-01

Full Text Available Surnamed „embryos’ saving method”, embryos culture is an in vitro technique used for over half of the century for saving the distant hybridization products, that would have degenerate in other conditions. Immature embryos culture is used for initiation of in vitro cultures imposed by the impossibility of using other explants for some of the plant species. Wheat is one of the crops that immature embryos culture technique is suitable for. This methods principle is based on aseptic embryos excision and their inoculation to an adequate culture medium. In vitro culture results depend in a greater manner of the basic culture medium and the hormonal balance used. Immature embryos isolated from two Romanian wheat cultivars – Dropia and Lovrin 41 – were inoculated for callus production on two types of basic media added with 2,4 D. The selected calluses were transferred on MS basic medium and several parameters were registered. Both cultivars emphasized a good callusing capacity, in a different percentage depending on the culture media used, such as 71,09 – 94,45%.. big differences between the cultivars regarding embriogenic callus frequency, shooting callus frequency and regenerated plants percentage were registered.

Regeneration and genetic transformation of cowpea (Vigna unguiculata Walp.)

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Filippone, E; Colucci, G; Ciardi, F; Monti, L [Department of Agronomy and Plant Genetics, Univ. of Naples Federico 11, Portici (Italy)

1997-07-01

Regeneration of cowpea (Vigna unguiculata Walp.) was achieved through massive bud formation induced in apical and lateral meristems by the herbicide Thidiazuron (TDZ). The effect of TDZ (5, 10, or 20 {mu}M) was tested in vitro on four different cowpea genotypes. Thidiazuron, even at the highest concentration, had no effect on seed germination. After one month of culture, multiple bud cluster formation was observed in all genotypes tested; about 80% of shoot apices regenerated multiple buds, whilst only 34% of cotyledonary nodes behaved in the same way. Histology of regenerating multiple bud clusters revealed that regeneration initiated from pre-existing meristems in the apex and cotyledonary node. Thidiazuron at 10 {mu}M appeared to be the best concentration to produce clusters with high number of buds, ranging from 5 to 10. Shoot elongation occurred only on MS medium without TDZ. On the same medium, 75% of elongated shoots rooted. For genetic transformation of cowpea, a direct DNA transfer methods in plants under in vivo conditions was tested by electroporation of plasmid DNA into the nodal meristematic cells. Some transformed plants were obtained, and produced T{sub 1} transformed progenies; their transgenic nature was confirmed by Southern analysis. (author). 21 refs, 2 figs, 3 tabs.

Regeneration and genetic transformation of cowpea (Vigna unguiculata Walp.)

International Nuclear Information System (INIS)

Filippone, E.; Colucci, G.; Ciardi, F.; Monti, L.

1997-01-01

Regeneration of cowpea (Vigna unguiculata Walp.) was achieved through massive bud formation induced in apical and lateral meristems by the herbicide Thidiazuron (TDZ). The effect of TDZ (5, 10, or 20 μM) was tested in vitro on four different cowpea genotypes. Thidiazuron, even at the highest concentration, had no effect on seed germination. After one month of culture, multiple bud cluster formation was observed in all genotypes tested; about 80% of shoot apices regenerated multiple buds, whilst only 34% of cotyledonary nodes behaved in the same way. Histology of regenerating multiple bud clusters revealed that regeneration initiated from pre-existing meristems in the apex and cotyledonary node. Thidiazuron at 10 μM appeared to be the best concentration to produce clusters with high number of buds, ranging from 5 to 10. Shoot elongation occurred only on MS medium without TDZ. On the same medium, 75% of elongated shoots rooted. For genetic transformation of cowpea, a direct DNA transfer methods in plants under in vivo conditions was tested by electroporation of plasmid DNA into the nodal meristematic cells. Some transformed plants were obtained, and produced T 1 transformed progenies; their transgenic nature was confirmed by Southern analysis. (author). 21 refs, 2 figs, 3 tabs

In vitro propagation of olive (Olea europaea L.) by nodal segmentation of elongated shoots.

Science.gov (United States)

Lambardi, Maurizio; Ozudogru, Elif Aylin; Roncasaglia, Romano

2013-01-01

Olive (Olea europaea L.), long-living, ever-green fruit tree of the Old World, has been part of a traditional landscape in the Mediterranean area for centuries. Both the fruits consumed after processing and the oil extracted from the fruits are among the main components of the Mediterranean diet, widely used for salads and cooking, as well as for preserving other food. Documentations show that the ancient use of this beautiful tree also includes lamp fuel production, wool treatment, soap production, medicine, and cosmetics. However, unlike the majority of the fruit species, olive propagation is still a laborious practice. As regards traditional propagation, rooting of cuttings and grafting stem segments onto rootstocks are possible, former being achieved only when the cuttings are collected in specific periods (spring or beginning of autumn), and latter only when skilled grafters are available. In both the cases, performance of the cultivars varies considerably. The regeneration of whole plants from ovules, on the other hand, is used only occasionally. Micropropagation of olive is not easy mainly due to explant oxidation, difficulties in explant disinfection, and labor-oriented establishment of in vitro shoot cultures. However today, the progress in micropropagation technology has made available the complete protocols for several Mediterranean cultivars. This chapter describes a micropropagation protocol based on the segmentation of nodal segments obtained from elongated shoots.

Antioxidant potential in regenerated tissues of medicinally important atropa accuminata

International Nuclear Information System (INIS)

Khan, F. A.; Abbasi, B. H.; Shinwari, Z. K.; Shah, S. H.

2017-01-01

Due to random exploitation from natural resources, an efficient regeneration system of medicinally important but rare plant species, Atropa acuminata for conservation was inevitable. Leaf explants were incubated on MS medium with different level of various plant growth regulators (PGRs) alone and in combination for callus induction and induced organogenesis. After 4 weeks of culture, callus induction was recorded with the highest frequency with 1.0 mg/l thidiazuron (TDZ) supplement. After 5 weeks of subsequent sub-culturing, optimum shoot induction frequency of 89% was achieved with 1.0 mg/l TDZ and 1.0 mg/l a-naphthaleneacetic acid (NAA) supplement. Highest number of shoots/explant (8.2) were recorded on MS medium with 2.0 mg/l 6-benzyladenine (BA)+1.0 mg/l NAA supplement. Shoots in elongation medium was recorded 5.8 cm long in two medium i.e., 1.0 mg/l TDZ supplement and 1.0 mg/l TDZ+1.0 mg/l NAA supplement. Successful In vitro rooting was induced on MS medium with all applied level of indole butyric acid (IBA). The regenerated shoots with well developed roots were successfully acclimatized in sterilized soil and transferred to greenhouse conditions. Furthermore higher activity for detoxifying DPPH free radical was shown by regenerated shoots in this medicinally important plant species. (author)

An Efficient In Vitro Regeneration System for Ornamental Ginger (Hedychium spp.)

Science.gov (United States)

An improved and efficient regeneration protocol was established for Hedychium via somatic embryogenesis. The plant material used consisted of 11 species and 9 cultivars of Hedychium. The explants consisted of young leaves taken from lateral or terminal shoots of mature greenhouse grown plants. These...

In-vitro regeneration of sugarcane (saccharum officinarum L.)

International Nuclear Information System (INIS)

Azu, E.

2009-01-01

Sugarcane (Saccharum officinarum L.) cultivars Barbados 46 (B46), Natal Coimbatore 339 (NC0339), Ragna, local sugarcane cultivar (LSC) and Kenana (Kn) series sourced from Kyebi, Akorley, Subriso, Jejeti and the University of Ghana Agriculture Research Station, Kpong, were evaluated for sterilisation, multiplication and rooting. An efficient double sterilisation protocol was achieved by immersing axillary buds pre-treated with Goldazim (active ingredient carbendazim) in 0.2% mercuric chloride for 7 minutes followed by 0.1 % mercuric chloride for 3 minutes. At this optimal sterilisation regime, 70% of the cultured buds were decontaminated. However, post sterilisation survival required the incorporation of 0.002g/L and 0.001g/L of amphotericin Band cefotaxime respectively in the culture medium indicating that the contaminants were endophytic. Evaluation for multiple shoot induction was conducted using Murashige and Skoog (1962) basal salts (MS) medium supplemented with 25g/L sucrose, 2mg/L IAA, 2mg/L GA 3 , 3g/L activated charcoal and varying concentrations (mg/L) of BAP or kinetin. The response of the cultivars to shoot induction varied with Ragna producing the highest number of shoots (2.17) on 5mg/L BAP indicating genotypic differences. Naphthalene acetic acid (NAA) at a concentration of 5mg/L induced the highest frequency (65%) of roots in all the cultivars tested. Post-flask acclimatization and survival of plantlets was high and independent of the age at which regenerated plantlets were transferred to the plant barn for weaning. This procedure could therefore be useful for regenerating sugarcane plantlets as well as provide target tissues for genetic transformation studies (au).

Effects of gibberellin mutations on in vitro shoot bud regeneration of ...

African Journals Online (AJOL)

hormone in regulating plant growth and development. It is interesting to know the effects of gibberellin and its signalling pathway on plant regeneration. In this report Arabidopsis thaliana landsberg (wild type), ga1-3 (gibberellin biosynthesis deficiency mutant), gai (gibberellin insensitive mutant), penta mutant (lacking GA1, ...

Antioxidant activity influenced by in vivo and in vitro mutagenesis in ...

African Journals Online (AJOL)

The antioxidant potential (1,1-diphenyl-2-picrylhydrazyl (DPPHº)-scavenging activity) of in vitro regenerated and induced mutant sugarcane (Saccharum officinarum L.) was investigated. Efficient callus induction and shoot regeneration were induced in bud explants when incubated on Murashige and Skoog (MS) medium ...

Optimization of in vitro regeneration and Agrobacterium tumefaciens-mediated transformation with heat-resistant cDNA in Brassica oleracea subsp. italica cv. Green Marvel.

Science.gov (United States)

Ravanfar, Seyed Ali; Aziz, Maheran Abdul; Saud, Halimi Mohd; Abdullah, Janna Ong

2015-11-01

An efficient system for shoot regeneration and Agrobacterium tumefaciens-mediated transformation of Brassica oleracea cv. Green Marvel cultivar is described. This study focuses on developing shoot regeneration from hypocotyl explants of broccoli cv. Green Marvel using thidiazuron (TDZ), zeatin, and kinetin, the optimization of factors affecting Agrobacterium-mediated transformation of the hypocotyl explants with heat-resistant cDNA, followed by the confirmation of transgenicity of the regenerants. High shoot regeneration was observed in 0.05-0.1 mg dm(-3) TDZ. TDZ at 0.1 mg dm(-3) produced among the highest percentage of shoot regeneration (96.67 %) and mean number of shoot formation (6.17). The highest percentage (13.33 %) and mean number (0.17) of putative transformant production were on hypocotyl explants subjected to preculture on shoot regeneration medium (SRM) with 200 µM acetosyringone. On optimization of bacterial density and inoculation time, the highest percentage and mean number of putative transformant production were on hypocotyl explants inoculated with a bacterial dilution of 1:5 for 30 min. Polymerase chain reaction (PCR) assay indicated a transformation efficiency of 8.33 %. The luciferase assay showed stable integration of the Arabidopsis thaliana HSP101 (AtHSP101) cDNA in the transgenic broccoli regenerants. Three out of five transgenic lines confirmed through PCR showed positive hybridization bands of the AtHSP101 cDNA through Southern blot analysis. The presence of AtHSP101 transcripts in the three transgenic broccoli lines indicated by reverse transcription-PCR (RT-PCR) confirmed the expression of the gene. In conclusion, an improved regeneration system has been established from hypocotyl explants of broccoli followed by successful transformation with AtHSP101 for resistance to high temperature.

Localization of QTLs for in vitro plant regeneration in tomato.

Science.gov (United States)

Trujillo-Moya, Carlos; Gisbert, Carmina; Vilanova, Santiago; Nuez, Fernando

2011-10-20

Low regeneration ability limits biotechnological breeding approaches. The influence of genotype in the regeneration response is high in both tomato and other important crops. Despite the various studies that have been carried out on regeneration genetics, little is known about the key genes involved in this process. The aim of this study was to localize the genetic factors affecting regeneration in tomato. We developed two mapping populations (F2 and BC1) derived from a previously selected tomato cultivar (cv. Anl27) with low regeneration ability and a high regeneration accession of the wild species Solanum pennellii (PE-47). The phenotypic assay indicated dominance for bud induction and additive effects for both the percentage of explants with shoots and the number of regenerated shoots per explant. Two linkage maps were developed and six QTLs were identified on five chromosomes (1, 3, 4, 7 and 8) in the BC1 population by means of the Interval Mapping and restricted Multiple QTL Mapping methods. These QTLs came from S. pennellii, with the exception of the minor QTL located on chromosome 8, which was provided by cv. Anl27. The main QTLs correspond to those detected on chromosomes 1 and 7. In the F2 population, a QTL on chromosome 7 was identified on a similar region as that detected in the BC1 population. Marker segregation distortion was observed in this population in those areas where the QTLs of BC1 were detected. Furthermore, we located two tomato candidate genes using a marker linked to the high regeneration gene: Rg-2 (a putative allele of Rg-1) and LESK1, which encodes a serine/threonine kinase and was proposed as a marker for regeneration competence. As a result, we located a putative allele of Rg-2 in the QTL detected on chromosome 3 that we named Rg-3. LESK1, which is also situated on chromosome 3, is outside Rg-3. In a preliminary exploration of the detected QTL peaks, we found several genes that may be related to regeneration. In this study we have

Localization of QTLs for in vitro plant regeneration in tomato

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Nuez Fernando

2011-10-01

Full Text Available Abstract Background Low regeneration ability limits biotechnological breeding approaches. The influence of genotype in the regeneration response is high in both tomato and other important crops. Despite the various studies that have been carried out on regeneration genetics, little is known about the key genes involved in this process. The aim of this study was to localize the genetic factors affecting regeneration in tomato. Results We developed two mapping populations (F2 and BC1 derived from a previously selected tomato cultivar (cv. Anl27 with low regeneration ability and a high regeneration accession of the wild species Solanum pennellii (PE-47. The phenotypic assay indicated dominance for bud induction and additive effects for both the percentage of explants with shoots and the number of regenerated shoots per explant. Two linkage maps were developed and six QTLs were identified on five chromosomes (1, 3, 4, 7 and 8 in the BC1 population by means of the Interval Mapping and restricted Multiple QTL Mapping methods. These QTLs came from S. pennellii, with the exception of the minor QTL located on chromosome 8, which was provided by cv. Anl27. The main QTLs correspond to those detected on chromosomes 1 and 7. In the F2 population, a QTL on chromosome 7 was identified on a similar region as that detected in the BC1 population. Marker segregation distortion was observed in this population in those areas where the QTLs of BC1 were detected. Furthermore, we located two tomato candidate genes using a marker linked to the high regeneration gene: Rg-2 (a putative allele of Rg-1 and LESK1, which encodes a serine/threonine kinase and was proposed as a marker for regeneration competence. As a result, we located a putative allele of Rg-2 in the QTL detected on chromosome 3 that we named Rg-3. LESK1, which is also situated on chromosome 3, is outside Rg-3. In a preliminary exploration of the detected QTL peaks, we found several genes that may be related

Vegetative regeneration

Science.gov (United States)

George A. Schier; John R. Jones; Robert P. Winokur

1985-01-01

Aspen is noted for its ability to regenerate vegetatively by adventitious shoots or suckers that arise on its long lateral roots. It also produces sprouts from stumps and root collars; but they are not common. In a survey of regeneration after clearcutting mature aspen in Utah. Baker (1918b) found that 92% of the shoots originated from roots, 7% from root collars, and...

The high frequency of variegated forms after in vitro mutagenesis in Saintpaulia ionantha Wendl.

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Marek Gaj

2014-01-01

Full Text Available The leaf-explants of Saintpaulia ionantha Wendl. var.'miniature' were treated by different doses of MNH and cultured on shoot regeneration medium. A strong toxic effect of some MNH doses on explant survival during the first two subcultures was noticed. The explants surviving treatment regenerated shoots with the efficiency comparable to the control. The high number of shoots regenerated from mutagenised leaves showed chlorophyll chimerism (so-called variegated forms. The use of 5 mM MNH for 1.5 or 2 h was found very effective, as 100% of survived explants regenerated variegated shoots. Besides hundreds of variegated forms also leaf-shape and flower-colour variants were observed in MNH-treated culture. Somaclonal variation was not observed in the control culture. The results indicate the great efficiency of in vitro applied MNH for in-duction of morphological variants of Saintpaulia, and especially variegated forms.

Establishment of an Efficient and Reproducible Regeneration System for Potato Cultivars Grown in Pakistan

International Nuclear Information System (INIS)

Iqbal, A.; Rizwan, A.; Mukhtar, Z.; Mansoor, S.; Asad, S.; Khalid, Z. M.

2016-01-01

The present study was carried out to assess the effect of growth regulators in three different combinations on mass propagation of currently grown three potato cultivars cv. Desiree, Kuroda and Cardinal. Varietal response on In vitro regeneration under different hormonal combinations. For callus induction, internodes of potato cultivars were cultured on modified Murashige and Skoog (MS) medium, added with different growth hormonal combinations. Callusing frequency for all these treatments and cultivars were recorded and in callus induction medium (CIM1) explants showed significantly higher callus formation as compared to two other combinations. For shoot induction, calli were cultured on Murashige and Skoog (MS) modified medium, supplemented with different hormonal combinations. Shoot induction medium (SIM3) gave best shoot induction frequency as compared to other media combinations. On the same media, an average number of shoots per explant were obtained for cultivar Desiree which is significantly different from the other two media combinations. Overall, the In vitro regeneration and multiplication potential was highest in the variety Cardinal followed by Kuroda and Desiree. The interaction between different hormonal combinations and varietal response for all the parameters showed significant differences. (author)

The influence of different hormone concentration and combination on callus induction and regeneration of Rauwolfia serpentina L. Benth.

Science.gov (United States)

Salma, U; Rahman, M S M; Islam, S; Haque, N; Jubair, T A; Haque, A K M F; Mukti, I J

2008-06-15

The influence of media composition on callus induction and subsequent regeneration of Rauwolfia serpentina L. Benth has been studied. High frequency (96.43%) callus induction was obtained when nodal segments from in vitro raised shoots were cultured on MS medium supplemented with 0.5 mg L(-1) BA and 2.0 mg L(-1) NAA. The callus differentiated into adventitious shoots when it was subcultured on MS medium supplemented with 2.0 mg L(-1) BA with 0.2 mg L(-1) NAA. Regenerated shoots were best rooted on half-strength MS medium with 1.0 mg L(-1) each of IBA and IAA.

Alginate-encapsulation of shoot tips of jojoba [Simmondsia chinensis (Link) Schneider] for germplasm exchange and distribution.

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Kumar, Sunil; Rai, Manoj K; Singh, Narender; Mangal, Manisha

2010-12-01

Shoot tips excised from in vitro proliferated shoots derived from nodal explants of jojoba [Simmondsia chinensis (Link) Schneider] were encapsulated in calcium alginate beads for germplasm exchange and distribution. A gelling matrix of 3 % sodium alginate and 100 mM calcium chloride was found most suitable for formation of ideal calcium alginate beads. Best response for shoot sprouting from encapsulated shoot tips was recorded on 0.8 % agar-solidified full-strength MS medium. Rooting was induced upon transfer of sprouted shoots to 0.8 % agar-solidified MS medium containing 1 mg l(-1) IBA. About 70 % of encapsulated shoot tips were rooted and converted into plantlets. Plants regenerated from encapsulated shoot tips were acclimatized successfully. The present encapsulation approach could also be applied as an alternative method of propagation of desirable elite genotype of jojoba.

In vitro binding of germanium to proteins of rice shoots

International Nuclear Information System (INIS)

Matsumoto, Hideaki; Takahashi, Eiichi

1976-01-01

The possibility of in vitro binding between proteins of rice shoots and germanium (Ge) was investigated. The proteins in mixtures of aqueous extracts of rice shoots and radioactive germanium ( 68 GeO 2 ) were fractionated. The binding of radioactivity to the proteins was observed even after 5 successive fractionation steps from the original mixtures. At the final fractionation step using polyacrylamide gel electrophoresis, a constant proportionality between protein concentration and associated radioactivity was found in most samples although not all. These results indicate that the binding of 68 Ge to proteins is not due to the simple adsorption by proteins. (auth.)

In vitro propagation of persimmon (Diospyros kaki Thunb.).

Science.gov (United States)

Giordani, Edgardo; Naval, Mar; Benelli, Carla

2013-01-01

Persimmon (Diospyros kaki Thunb.) is a temperate fruit tree species diffused in all continents. The traditional propagation method adopted by the nursery industry is based on budding/grafting scion cultivars on seedlings from D. kaki, Diospyros lotus, and Diospyros virginiana, the most important species used as rootstock, reproduced by seeds since they are not easy to root. Furthermore, most of nonastringent cultivars of persimmon are not compatible with D. lotus, a rootstock largely utilized because of its hardiness and frost resistance. The main in vitro tissue culture techniques, developed for persimmon, deal with direct regeneration (from dormant buds and root tips), and indirect regeneration through callus from dormant buds, apexes, and leaves. The bottlenecks of micropropagation are (1) the recalcitrance of many cultivars to in vitro establishment, (2) the low multiplication ratio of D. kaki compared to other fruit tree species, (3) the very low rooting ability of ex novo microcuttings both from direct and indirect regeneration, (4) the high sensitivity to transplant from in vitro to in vivo conditions. The development of reliable in vitro regeneration procedures is likely to play a key role for production of both clonal rootstocks and self-rooted cultivars. The general protocol for micropropagation of persimmon reported here is based on the establishment of winter dormant buds in vitro, shoot development, multiplication and elongation, and shoot rooting, using cytokinins (BA or zeatin) in a MS media along with an auxinic pretreatment for rooting induction.

Effect Of Gamma Rays And Growth Regulators On Explants Excised From In Vitro Shoots And Greenhouse Seedlings, Of Pepper (Capsicum Annum L.)

International Nuclear Information System (INIS)

Maarouf, A. A.; Kassem, M.

2004-01-01

This experiment was conducted on pepper (Capsicum annum L.) to compare the ability of the in vitro explants with those of greenhouse grown seedlings on shoot proliferation and callus formation and their ability to form plantlets and the effect of gamma irradiation and growth regulators on the shoot tip, hypocotyls and leaf tissue was used as laboratory explants, leaf tissue nodes and internodes were taken from greenhouse seedlings. 6- benzyla-minopurine (BAP) in different concentrations was combined with Indoleacertic acid (IAA) to know their effect on shoot proliferation, 2,4 - Dichlorophenoxy acetic acid (2,4- D) was used for callus formation, and use stimulation effect of gamma irradiation, potassium nitrat (KNO 3 ), Thidaiazurom (TDZ) and casine hydrolysate (CH) for plantlet formation. The results showed that the highest percentage of callus was obtained by in vitro hypocotyls and greenhouse grown nodes followed by in vitro leaf tissue thereafter greenhouse leaf tissue. The shoot tips were the lowest efficient explants in producing callus in both in vitro and greenhouse ones. The highest percentage of shooting resulted from shoot tip, hypocotyls and leaf tissue of in vitro explants, followed by shoot tip, nodes and internodes of greenhouse grown explants and the lowest percentage was recorded by leaf tissue. Highest percentage of shoot number was obtained form greenhouse grown shoot tip followed by in vitro shoot tip, hypocotyls and leaf tissue of greenhouse grown seedlings the internodes were the lowest efficient in producing shoots. The highest success in plantlet formation was caused by TDZ followed by gamma irradiation and the other treatments were equaled. (Authors)

In vitro regeneration of selected commercial Tanzanian open ...

African Journals Online (AJOL)

Yomi

2012-03-15

Mar 15, 2012 ... regenerated in vitro using immature zygotic embryos as ex-plants. Callus induction .... Ears were harvested between 12 to 16 days post pollination depending on .... After 28 days of culture on CMM, a mass of dedifferentiated cells .... showed high regeneration ability compared to Situka M-1 that had good ...

Heritability of regeneration in tissue cultures of sweet potato (Ipomoea batatas L.).

Science.gov (United States)

Templeton-Somers, K M; Collins, W W

1986-03-01

A population of open-pollinated progeny from 12 parents, and the 12 parents, was surveyed for in vitro growth and regeneration characteristics. Four different tissue culture procedures involving different media and the use of different explants to initiate the cultures were used. Petiole explants from young leaves were used as explants for initiation of callus cultures. These were evaluated for callus growth rate, friability, and callus color and texture, before transferring to each of three different regeneration media for evaluation of morphogenetic potential. Small shoot tips also were used to initiate callus cultures, which were evaluated for the same growth characteristics and transferred to growth-regulator free regeneration media. Regeneration occurred through root or shoot regeneration or through embryogenesis. Tissue culture treatment effects, as well as genotypic effects, were highly significant in determining: the types of callus produced, callus growth rates, color and texture on the two types of media used for the second and third subcultures. The family x treatment interaction was generally not statistically significant, affecting only callus color. Estimates of narrow sense heritability for callus growth rate in both the second and third subcultures were high enough (0.35 and 0.63, respectively) for the evaluation of parental lines for selection procedures. These characteristics were also the only early culture callus traits that were consistently correlated with later morphogenesis of the cultures. They were negatively correlated with root or shoot regeneration. The occurence of somatic embryogenesis was not correlated with early callus growth characteristics. Genetic and treatment effects were highly significant in the evaluation of morphogenetic potential, through root or shoot regeneration, or through embryogenesis. Regeneration of all types was of low frequency for all procedures, expressed in ≦ 11% of the cultures of the total population.

Rapid multiplication of Dalbergia sissoo Roxb.: a timber yielding tree legume through axillary shoot proliferation and ex vitro rooting.

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Vibha, J B; Shekhawat, N S; Mehandru, Pooja; Dinesh, Rachana

2014-01-01

An efficient and improved method for in vitro propagation of mature tree of Dalbergia sissoo, an ecologically and commercially important timber yielding species, has been developed through axillary shoot proliferation. Bud breaking occurred from nodal shoot segments derived from rejuvenated shoots produced during early spring from a 20-25-year-old lopped tree, on MS medium containing 8.88 μM benzylaminopurine (BAP). Multiple shoots differentiated (20-21shoots/node) on re-culture of explants on half-strength agar gelled amended MS medium with a combination of 2.22 μM of BAP and 0.002 μM of thidiazuron (TDZ) with 1.0 mM each of Ca(NO3)2, K2SO4, KCl, and NH4(SO4)2. The maximum shoot multiplication (29-30 shoots/node) was achieved on subculturing in the above mentioned but liquid medium. Furthermore, the problem of shoot tip necrosis and defoliation observed on solid medium were overcome by the use of liquid medium. Ex vitro rooting was achieved on soilrite after basal treatment of microshoots with 984 μM of indole-3-butyric acid (IBA) for 2 min. About 90 % microshoots were rooted on soilrite within 2-3 weeks under the greenhouse conditions. From 20 nodal shoot segments, about 435 hardened plants were acclimatized and transplanted. This is the first report for rapid in vitro propagation of mature trees of D. sissoo on liquid medium followed by ex vitro rooting.

Photoperiod and growth regulators on in vitro shoot induction in Heliconia latispatha

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Antonio Anderson de Jesus Rodrigues

2016-12-01

Full Text Available Considering the growing economic importance of tropical flowers and the advantages of techniques applied to the in vitro cultivation of these plants, it is necessary to carry out studies to evaluate growth in species such as Heliconia latispatha. The aim of this study therefore, was to evaluate in vitro shoot induction for different concentrations of BAP and NAA and as a function of the photoperiod. Explants from zygotic embryos were inoculated in MS medium containing different concentrations of BAP (0.0, 0.5, 1.0, 1.5, 2.0 or 2.5 mg L-1, with the cultures kept in a growth room at a temperature of 24.0 ± 2.0° C, under a photoperiod of 12 and 16 hours of light and a light intensity of 30 μmol m-2 s-1. At 21, 28, 35, 42 and 49 days after inoculation, the number of shoots per explant was evaluated. The treatment at the BAP concentration that gave the best multiplication rate (2.5 mg L-1 was set, and was tested in a further trial with different concentrations of NAA (0.0, 0.2, 0.4, 0 6, 0.8 or 1.0 mg L-1 under the same conditions as the previous experiment. The experimental designs were completely randomised, with five replications, and analysed in a 6 x 2 factorial. The data were submitted to analysis of variance and regression. No significant differences were seen in relation to the photoperiod or its interaction with the cytokinin and auxin under test. Multiplication was greater in the presence of 2.5 mg L-1 BAP, which gave a rate of 1.25 shoots/explant at 49 days of in vitro culture. The association of this BAP dosage with 1.0 mg L-1 NAA was even more efficient, producing 1.83 shoots per explant at 30 days of growth. The use of BAP together with NAA is beneficial to the induction of shoots in H. latispatha.

In Vitro Propagarion and Cryopreservation of Important Grape Cultivars (Vitis Vinifera L. and Rootstocks

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F. CELEBI TOPRAK

2014-06-01

Full Text Available Grape (Vitis vinifera L. is among the most important species that is cultivated almost all around the world. There are over one thousand varieties that are grown for raisin, fresh consumption and wine making purposes. The grape germplasm resources are generally maintained as whole plants under field conditions. The traditional way of germplasm preservation is very risky due to natural uncertainties. In vitro technologies can help producing healthy propagation materials free from viroids, viruses, bacteria, phytoplasmas, fungi, and nematodes. When combined with cryopreservation technologies in vitro preservation systems can allow safe protection and propagation of valuable Vitis genetic resources. In this study, 12 commercial cultivar and two rootstock materials were tested for the applicability of long term preservation by in vitro clonal propagation and cryopreservation techniques. Axillary shoot tips collected from newly emerging shoots were placed in Magenda boxes containing 30 g/l sucrose on MS medium and cultured in a growth chamber adjusted to 16 h ligth/25o C and 8 h dark/17o C. All grape genotypes tested responded well to this application and produced healthy root and shoots. Shoot explants from these in vitro stocks were subcultured in every three months for one year. Apical dome explants excised from in vitro grape plants were stored in liquid nitrogen for cryopreservation. Genotypes varied in their responses to cryopservation treatment. Five genotypes showed shoot or callus formation. Regenerated shoots continued to grow and produced normal shoots and roots, but no plants could be developed from calli. Flow cytometry analysis of regenerants from continuous subculture and cryopreservation did not show any chromosome number abnormalities. In vitro micropropagation is an excellent choice for a long-term conservation of grape germplasm, which allows access to actively growing plant materials without seasonal restriction. Such cultures are

Plant regeneration of natural tetraploid Trifolium  Hum pratense L

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HATICE ÇÖLGEÇEN

2008-01-01

Full Text Available The regeneration of natural tetraploid T. pratense, originated from Erzurum-Turkey, is reported in this study. This plant has low seed setting and hard seed problems due to polyploidy. Hypocotyl, cotyledon, apical meristems, epicotyl and young primary leaves were inoculated on MS and PC-L2 media containing different concentrations of BAP and NAA as growth regulators. The best shoot formation has been observed on explants initiated from apical meristem placed on PC-L2 medium that includes 2 mg dm-3 BAP and 1 mg dm-3 NAA. 94.4% of the shoots originated from calli were rooted on PC-L2 medium with 1 mg dm-3 NAA. In vitro organogénesis has been accomplished in the natural tetraploid T. pratense regenerated plants successively transferred to the field

In vitro propagation, carotenoid, fatty acid and tocopherol content of Ajuga multiflora Bunge.

Science.gov (United States)

Sivanesan, Iyyakkannu; Saini, Ramesh Kumar; Noorzai, Rafi; Zamany, Ahmad Jawid; Kim, Doo Hwan

2016-06-01

The effect of plant growth regulators on shoot proliferation from shoot tip explants of Ajuga multiflora was studied. The highest number of shoots (17.1) was observed when shoot tip explants were cultured on Murashige and Skoog (MS) medium fortified with 8.0 µM 6-Benzyladenine (BA) and 2.7 µM α-naphthaleneacetic acid (NAA). The mean number of shoots per explant was increased 1.6-fold in liquid medium as compared with semi-solid medium. Maximum rooting (100 %) with an average of 7.2 roots per shoot was obtained on MS basal medium. Rooted plantlets were successfully acclimatised in the greenhouse with 100 % survival rate. Composition of carotenoids, fatty acids and tocopherols was also studied from leaves of greenhouse-grown plants and in vitro-regenerated shoots of A. multiflora. The greatest amounts of carotenoids, fatty acids and tocopherols were obtained from leaves of in vitro-regenerated shoots cultured on MS basal medium, followed by leaves of greenhouse-grown plants and leaves of in vitro-regenerated shoots cultured on MS basal medium with 2.0 µM BA or thidiazuron. The most abundant carotenoid in A. multiflora leaves was all-E-lutein (89.4-382.6 μg g -1  FW) followed by all-E-β-carotene (32.0-156.7 μg g -1  FW), 9'-Z-neoxanthin (14.2-63.4 μg g -1  FW), all-E-violaxanthin (13.0-45.9 μg g -1  FW), all-E-zeaxanthin (1.3-2.5 μg g -1  FW) and all-E-β-cryptoxanthin (0.3-0.9 μg g -1  FW). α-Tocopherol was the predominant tocopherol in A. multiflora leaves. Linolenic acid (49.03-52.59 %) was detected in higher amounts in A. multiflora leaf samples followed by linoleic acid (18.95-21.39 %) and palmitic acid (15.79-18.66 %).

Some factors affecting the in vitro culture of banana

International Nuclear Information System (INIS)

Zadi, T.A.N.; Khan, N.H.; Rehman, Z.U.

2006-01-01

Factors affecting in vitro regeneration of shoots in shoot tip explant cultures of banana cultivar 'Basrai', such as solid and liquid media, growth regulators, vitamins, and antioxidants were studied. Three-quarters strength of MS liquid medium supplemented with 17.75 micro m 6-benzyladenine (BA), 11.42 micro M indole-3-acetic acid (IAA) and 205 micro M adenine sulphate induced the formation of mean number of 12.3 shoots, with the mean length of 3.0 cm, after three weeks of culture. Maximum shoot multiplication (14.33) occurred in liquid medium containing 22.19 micro M BA. Addition of 2.0% activated charcoal (AC) to the liquid medium improved quality of the regenerated plants with expanded and glossy leaves, though the number of shoots was reduced (13.66). Profuse formation of roots was characteristically induced by AC. Addition of citric acid (CA) to the medium caused decline in morphogenetic expression of the cultures. (author)

In vitro clonal propagation of the neem tree ( Azadirachta indica A ...

African Journals Online (AJOL)

In vitro clonal propagation of the neem tree (Azadirachta indica A. Juss.) M Shahin-uz-zaman, M Ashrafuzzaman, MS Haque, LN Luna. Abstract. A study was conducted with root and shoot tip explants of neem to develop an efficient protocol of regeneration. Shoot tips and root tips from 10 - 20 days old seedlings of neem ...

Effect of light on contents of coumarin compounds in shoots of Ruta graveolens L. cultivated in vitro

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Halina Ekiert

2014-01-01

Full Text Available Shoots of Ruta graveolens L. (Rutaceae were cultivated in stationary liquid culture under different light conditions: constant artificial light (900 lx, darkness, constant artificial light (900 ix following irradiation with UV-C light. The contents of five furanocoumarins: psoralen, bergapten, xanthotoxin, isopimpinellin and imperatorin, as well as biogenetic precursor of these metabolites, umbelliferone, were determined by HPLC method in shoots cultivated in vitro and in overground parts of plants growing in open air. It was shown that light conditions, tested in these experiments, significantly influenced contents of the metabolites in shoots cultivated in in vitro culture. Total content of the coumarin compounds in shoots cultivated under constant artificial light (900 lx was equal or higher than in plants growing under natural conditions. Therefore, it is suggested that stationary liquid shoot culture of R. graveolens. can be an alternative source for obtaining biologically active furanocoumarins.

Acetylcholine suppresses shoot formation and callusing in leaf explants of in vitro raised seedlings of tomato, Lycopersicon esculentum Miller var. Pusa Ruby.

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Bamel, Kiran; Gupta, Rajendra; Gupta, Shirish C

2016-06-02

We present experimental evidence to show that acetylcholine (ACh) causes decrease in shoot formation in leaf explants of tomato (Lycopersicon esculentum Miller var Pusa Ruby) when cultured on shoot regeneration medium. The optimum response was obtained at 10(-4) M ACh-enriched medium. ACh also causes decrease in percentage of cultures forming callus and reduces the callus mass. Inhibitors of enzymatic hydrolysis of ACh, neostigmine and physostigmine, also suppresses callogenesis and caulogenesis. On the other hand, the breakdown products of Ach, choline and acetate, do not alter the morphogenic response induced on the shoot regeneration medium. Neostigmine showed optimal reduction in shoot formation at 10(-5) M. The explants cultured on neostigmine augmented medium showed decline in the activity of ACh hydrolyzing enzyme acetylcholinesterase. ACh and neostigmine added together showed marked reduction in callus mass. These results strongly support the role of ACh as a natural regulator of morphogenesis in tomato plants.

Acetylcholine suppresses shoot formation and callusing in leaf explants of in vitro raised seedlings of tomato, Lycopersicon esculentum Miller var. Pusa Ruby

Science.gov (United States)

Bamel, Kiran; Gupta, Rajendra; Gupta, Shirish C.

2016-01-01

ABSTRACT We present experimental evidence to show that acetylcholine (ACh) causes decrease in shoot formation in leaf explants of tomato (Lycopersicon esculentum Miller var Pusa Ruby) when cultured on shoot regeneration medium. The optimum response was obtained at 10−4 M ACh-enriched medium. ACh also causes decrease in percentage of cultures forming callus and reduces the callus mass. Inhibitors of enzymatic hydrolysis of ACh, neostigmine and physostigmine, also suppresses callogenesis and caulogenesis. On the other hand, the breakdown products of Ach, choline and acetate, do not alter the morphogenic response induced on the shoot regeneration medium. Neostigmine showed optimal reduction in shoot formation at 10−5 M. The explants cultured on neostigmine augmented medium showed decline in the activity of ACh hydrolyzing enzyme acetylcholinesterase. ACh and neostigmine added together showed marked reduction in callus mass. These results strongly support the role of ACh as a natural regulator of morphogenesis in tomato plants. PMID:27348536

Studies on seed germination and in vitro shoot multiplication of ...

African Journals Online (AJOL)

Yomi

2011-12-21

Dec 21, 2011 ... vitro seed germination and plantlet regeneration of this plant. ... Key words: Germination, gibberellic acid, growth regulators, node explants, Satureja ..... Abscisic Acid: A. Seed Maturation and Antistress Signal, 3rd ed. Sinauer ...

Reproducible and expedient rice regeneration system using in vitro ...

African Journals Online (AJOL)

Inevitable prerequisite for expedient regeneration in rice is the selection of totipotent explant and developing an apposite combination of growth hormones. Here, we reported a reproducible regeneration protocol in which basal segments of the stem of the in vitro grown rice plants were used as ex-plant. Using the protocol ...

In vitro propagation via organogenesis and synthetic seeds of Urginea altissima (L.f.) Baker: a threatened medicinal plant.

Science.gov (United States)

Baskaran, Ponnusamy; Kumari, Aloka; Van Staden, Johannes

2018-01-01

Efficient in vitro propagation systems via organogenesis and synthetic seeds were developed for the first time for conservation and commercial propagation from leaf or longitudinal thin cell layer (lTCL) leaf or shoot-tip explants of Urginea altissima . Various plant growth regulators and phloroglucinol were used in semi-solid and liquid Murashige and Skoog (MS) medium to establish multiplication of shoots and roots for in vitro regeneration. Of the various treatments, the highest number of shoots (17.4 per lTCL leaf explant) was obtained on liquid MS medium supplemented with 10 µM meta -Topolin ( m T) and 2 µM benzyladenine followed by transferal to semi-solid MS media. The shoot tips were encapsulated with liquid MS medium plus 3% (w/v) sodium alginate and 100 mM calcium chloride. Adventitious shoot regeneration (91.0%; 12.6 shoots per synthetic seed) of synthetic seeds was achieved on semi-solid MS medium supplemented with 10 µM m T and 2 µM naphthaleneacetic acid (NAA) after 15 days of storage in darkness at 25 ± 2 °C. Regenerated shoots rooted (9.8 roots per shoot; 6.5 cm long) efficiently when transferred to 5 µM indole-3-butyric acid and 2.5 µM NAA. All the plantlets were successfully acclimatized (100%) in a vermiculite:soil (1:1 v/v) mixture in the greenhouse.

Regeneração in vitro de urucum (Bixa orellana L. a partir de diferentes tipos de explantes In vitro regeneration of annatto (Bixa orellana L. from various explants

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Jane Fiuza Rodrigues Portela de Carvalho

2005-12-01

Full Text Available Este trabalho teve como objetivo avaliar a regeneração in vitro de plantas de urucum (Bixa orellana L. a partir de diferentes tipos de explantes. Para definir o meio de cultura adequado para indução de brotações, diferentes concentrações e, ou, combinações da auxina AIA e das citocininas BAP e ZEA foram testadas. As melhores respostas de regeneração para segmentos de hipocótilo, nós cotiledonares e hipocótilos invertidos foram observadas em meios suplementados de ZEA (2,28 µM e AIA (0,30 µM, ZEA (4,56 µM e ZEA (4,56 µM, respectivamente. O meio de enraizamento mais eficaz foi o MS, com a metade de sua concentração salina e 5 µM de AIB. Análises citológicas, realizadas antes da aclimatação, confirmaram a estabilidade cromossômica das plantas cultivadas in vitro, não sendo detectado variação com relação ao número de cromossomos metafásicos (2n = 14.The present work aimed the establishment of a regeneration protocol for annatto (Bixa orellana L. from different juvenile explants. In order to promote shoot induction, different concentrations and/or combinations of IAA and the cytokinins BAP and ZEA were assessed. Better regeneration responses were achieved when segmented hypocotyl, cotiledonary nodes and inverted hypocotyl were cultured onto MS-based medium supplemented with ZEA (2.28 µM and IAA (0.30 µM, ZEA (4.56 µM or ZEA (4.56 µM, respectively. Rooting of elongated shoots displayed higher frequencies when half-strength MS medium with IBA (5 µM was used. No genetic variation was detected among regenerants as revealed by cytological analysis based on metaphasic chromosome countings (2n = 14.

Response of Syngonium podophyllum L. ‘White Butterfly’ shoot cultures to alternative media additives and gelling agents, and flow cytometric analysis of regenerants

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JAIME A. TEIXEIRA DA SILVA

2015-05-01

Full Text Available Abstract. Teixeira da Silva JA. 2015. Response of Syngonium podophyllum L. ‘White Butterfly’ shoot cultures to alternative media additives and gelling agents, and flow cytometric analysis of regenerants. Nusantara Bioscience 7: 26-32. Syngonium podophyllum L. (arrowhead vine is a popular leafy indoor pot plant whose tissue culture has been established, primarily through in vitro shoot culture, but several interesting aspects have not yet been explored. In this study, cv. ‘White Butterfly’ was used to investigate the response of shoot formation to alternative gelling agents and media additives. Gellan gum (Gelrite® at 2 g/L resulted in greater leaf production, plantlet fresh weight and higher chlorophyll content (SPAD value than all other gelling agents tested, including agar, Bacto agar, phytagel, oatmeal agar, potato dextrose agar, barley starch and corn starch, when on a basal Hyponex® (NPK = 6.5: 6: 19; 3 g/L medium. Several alternative liquid medium additives tested (low and full fat milk, Coca-Cola®, coffee, Japanese green, Oolong and Darjeeling teas negatively impacted plant growth, stunted roots and decreased chlorophyll content (SPAD value of leaves. Plant growth on medium with refined sucrose or table sugar responded similarly. Poor growth was observed when crude extract from a high rebaudioside-containing stevia (Stevia rebaudiana Bertoni line - an artificial sweetener - was used. Leaf tissue from the control did not show any endopolyploidy but low levels of endopolyploidy (8C were detected in some treatments.

A fasciclin-like arabinogalactan-protein (FLA mutant of Arabidopsis thaliana, fla1, shows defects in shoot regeneration.

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Kim L Johnson

Full Text Available BACKGROUND: The fasciclin-like arabinogalactan-proteins (FLAs are an enigmatic class of 21 members within the larger family of arabinogalactan-proteins (AGPs in Arabidopsis thaliana. Located at the cell surface, in the cell wall/plasma membrane, they are implicated in many developmental roles yet their function remains largely undefined. Fasciclin (FAS domains are putative cell-adhesion domains found in extracellular matrix proteins of organisms from all kingdoms, but the juxtaposition of FAS domains with highly glycosylated AGP domains is unique to plants. Recent studies have started to elucidate the role of FLAs in Arabidopsis development. FLAs containing a single FAS domain are important for the integrity and elasticity of the plant cell wall matrix (FLA11 and FLA12 and FLA3 is involved in microspore development. FLA4/SOS5 with two FAS domains and two AGP domains has a role in maintaining proper cell expansion under salt stressed conditions. The role of other FLAs remains to be uncovered. METHOD/PRINCIPAL FINDINGS: Here we describe the characterisation of a T-DNA insertion mutant in the FLA1 gene (At5g55730. Under standard growth conditions fla1-1 mutants have no obvious phenotype. Based on gene expression studies, a putative role for FLA1 in callus induction was investigated and revealed that fla1-1 has a reduced ability to regenerate shoots in an in vitro shoot-induction assay. Analysis of FLA1p:GUS reporter lines show that FLA1 is expressed in several tissues including stomata, trichomes, the vasculature of leaves, the primary root tip and in lateral roots near the junction of the primary root. CONCLUSION: The results of the developmental expression of FLA1 and characterisation of the fla1 mutant support a role for FLA1 in the early events of lateral root development and shoot development in tissue culture, prior to cell-type specification.

Plant regeneration from petiole segments of some species in tissue culture

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Krystyna Klimaszewska

2013-12-01

Full Text Available The regeneration ability of 21 plant species belonging to 14 families was tested. The method of tissue culture in vitro was applied, on basic MS medium with an addition of growth regulators from the auxin and cytokinin groups. From among the investigated plant groups Peperomia scandens and Caladium × hortulanum were capable of plant regeneration, Passiilora coerulea regenerated shoots, Hedera helix, Begonia glabra, Coleus blumei, Fuchsia hybrida, Passiflora suberosa and Peperomia eburnea formed callus and roots, Kalanchoe blossfeldiana, Pelargonium grandiflorum, P. peltatum, P. radula, Coleus shirensis and Magnolia soulangeana produced callus, Philodendron scandens, Rhododendron smirnovii, Hibiscus rosa-sinensis, Coprosma baueri, Cestrum purpureum and Solanum rantonnetii did not exhibit any regeneration reactions.

Effect of different in vitro culture extracts of black pepper (Piper nigrum L.) on toxic metabolites-producing strains.

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Ahmad, Nisar; Abbasi, Bilal Haider; Fazal, Hina

2016-03-01

In the present study, the effect of different in vitro cultures (callus, in vitro shoots) and commercially available peppercorn extract was investigated for its activity against toxic metabolite-producing strains (Escherichia coli, Pseudomonas aeroginosa, Salmonella typhi, Bacillus subtilis, Bacillus cereus, Staphylococcus aureus, and Candida albicans). These in vitro cultures were extracted with ethanol, hexane, and chloroform, and the antipathogenic activity was determined by well-diffusion method. Hexane extract of callus showed 22 mm zone of inhibition against B. cereus, 23 mm against S. aureus, while regenerated shoots and seeds have shown 24.3 and 26 mm zones of inhibition. The ethanolic extracts of regenerated Piper shoots have shown 25 mm activity against S. aureus, 21 mm against B. cereus, and 16 mm in the case of C. albicans in comparison with standard antibiotics. Peppercorn extracts in chloroform and ethanol had shown activities against B. cereus (23.6 mm) and B. subtilis (23.5 mm). During in vitro organogenesis and morphogenesis, cells and tissues produced a comparable phytochemicals profile like mother plant. Morphogenesis is critically controlled by the application of exogenous plant-growth regulators. Such addition alters the hormonal transduction pathways, and cells under in vitro conditions regenerate tissues, which are dependant on the physiological state of cells, and finally enhance the production of secondary metabolites. To the best of our knowledge, this is the first report to compare the antimicrobial potential of in vitro regenerated tissues and peppercorn with standard antibiotics. In conclusion, most of the extracts showed pronounced activities against all the pathogenic microbes. This is a preliminary work, and the minimum inhibitory concentration values needs to be further explored. Regenerated tissues of P. nigrum are a good source of biologically active metabolites for antimicrobial activities, and callus culture presented itself as

In vitro propagation and conservation of Satureja avromanica Maroofi-an indigenous threatened medicinal plant of Iran.

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Mozafari, Ali Akbar; Vafaee, Yavar; Karami, Edris

2015-07-01

An efficient and rapid in vitro propagation system for Satureja avromanica, a rare and endangered folk medicinal plant of Iran was developed through the shoot tip and leaf disc explants. Nodal and leaf explants from wild plants were established on MS and WPM media supplemented with BA, BAP and TDZ (0, 0.1, 0.5, 1, 1.5, 2, 5 and 10 mgl(-1)) alone or by application of BA and TDZ (0, 2, 5 and 10 mgl(-1)) in combination with IBA and 2,4-D (0, 0.1, 0.5 and 1 mgl(-1)), respectively. Based on results, the highest mean shoot number (6.21) was obtained on MS medium supplemented with 2 mgl(-1) BA. Regarding the shoot elongation, MS supplemented with 2 mgl(-1) TDZ and MS containing 5 mgl(-1) BA showed the longest shoots (4.82 and 4.39 cm, respectively) after 6 weeks of culture. As a matter of fact, increasing all three tested cytokinins levels led to enhancement of explant response frequency and regenerated shoot number. On the other side, WPM medium supplemented with 0.1 mgl(-1) IBA was found suitable for rooting of regenerated shoots. RAPD molecular analysis revealed genetic stability of in vitro raised plants. In conclusion, individual application of BA, BAP and TDZ were in favor of S. avromanica direct shoot regeneration while treatment media with a combination of IBA and BA as well as 2,4-D and TDZ resulted in callogenesis in most explants. Finally, the in vitro raised plantlets were acclimatized and successfully established in the greenhouse conditions. Our developed protocol can be employed for the large-scale micropropagation and conservation of S. avromanica as a threatened medicinal plant.

Efficient plant regeneration of bittersweet (Solanum dulcamara L., a medicinal plant

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Arzu Ucar Turker

2011-01-01

Full Text Available Solanum dulcamara L. (bittersweet is a medicinal plant that has been used to treat skin diseases, warts, tumors, felons, arthritis, rheumatism, bronchial congestion, heart ailments, ulcerative colitis, eye inflammations, jaundice and pneumonia. A reliable in vitro culture protocol for bittersweet was established. Explants (leaf and petiole segments were cultured on Murashige and Skoog minimal organics (MSMO medium with various plant growth regulator combinations. Leaf explants formed more shoots than petiole explants. Plant regeneration was observed through indirect organogenesis with both explants. Best shoot proliferation was obtained from leaf explants with 3 mg/l BA (benzyladenine and 0.5 mg/l IAA (indole-3-acetic acid. Regenerated shoots were transferred to rooting media containing different levels of IAA (indole-3-acetic acid, IBA (indole-3-butyric acid, NAA (naphthalene acetic acid or 2,4-D (2,4 dichlorophenoxyacetic acid. Most shoots developed roots on medium with 0.5 mg/l IBA. Rooted explants were transferred to vermiculate in Magenta containers for acclimatization and after 2 weeks, they were planted in plastic pots containing potting soil and maintained in the plant growth room.

Micropropagation and validation of genetic and biochemical fidelity among regenerants of Nothapodytes nimmoniana (Graham) Mabb. employing ISSR markers and HPLC.

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Prakash, Lokesh; Middha, Sushil Kumar; Mohanty, Sudipta Kumar; Swamy, Mallappa Kumara

2016-12-01

An in vitro protocol has been established for clonal propagation of Nothapodytes nimmoniana which is an important source of Camptothecin (CPT). Elite source was identified based on the chemical potency to accumulate the optimum level of CPT. Different types and concentrations of plant growth regulators were used to study their effect on inducing multiple shoots from the explants regenerated from embryos of N. nimmoniana. Of these, a combination of N6-benzyladenine (0.2 mg L -1 ) and Indole-3-butyric acid (IBA) (0.1 mg L -1 ) proved optimum for differentiating multiple shoots in 90.6 % of the cultures with an average of 10.24 shoots per explant obtained within 8 weeks of inoculation. Nearly, 92 % of the excised in vitro shoots rooted on half strength Murashige and Skoog (MS) medium containing 0.05 % activated charcoal, supplemented with 1-naphthaleneacetic acid and IBA at 0.1 mg L -1 each. The micropropagated plants were evaluated for their genetic fidelity by employing inter simple sequence repeats (ISSR) markers. Ten individuals, randomly chosen from a population of 145 regenerants, were compared with the donor plant. The regenerated plants were also evaluated for their chemical potency using high-performance liquid chromatography (HPLC) analysis of CPT content. The true-to-type nature of the micropropagated plants was confirmed based on their monomorphic banding profiles with that of the mother plants using ISSR markers. Besides, HPLC evaluation of the CPT content confirmed the existence of chemical uniformity among the regenerated plants and the elite mother plant.

Improved efficiency of plant regeneration from protoplasts of eggplant Solanum melongena L.

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Guri, A; Izhar, S

1984-12-01

Eggplant (Solanum melongena L.) mesophyll protoplasts were obtained from in vitro growing plants of line 410 and cv. 'Classic'. Relatively high (15%) plating efficiency was achieved using petri dishes with alternate quadrants containing reservoir medium (R medium + 1% activated charcoal) and culture medium. Shoot regeneration occurred within 6 weeks following initiation of protoplast culture.

In vitro regeneration of hybrid plantlets of cashew (Anacardium ...

African Journals Online (AJOL)

Embryos from immature nuts of cashew (Anacardium occidentale L.) were cultured in vitro to regenerate improved hybrid plantlets. Explants (embryo) were excised from developing F1 hybrid immature nuts derived from diallel cross and harvested at 2-, 4-, 6- and 8-weeks after pollination (WAPo) for in vitro culture.

Influence of salicylic acid on in vitro propagation and salt tolerance ...

African Journals Online (AJOL)

Salicylic acid (SA) has been reported to improve in vitro regeneration as well as induce abiotic stress tolerance in plants. The effects of varying SA concentrations (0, 0.5, and 1 mM) on in vitro shoot apices of two Hibiscus species, Hibiscus moscheutos (cv 'Luna Red') and Hibiscus acetosella, grown under various salt ...

In vitro micropropagation of Dracaena sanderiana Sander ex Mast: An important indoor ornamental plant.

Science.gov (United States)

Aslam, Junaid; Mujib, Abdul; Sharma, Maheshwar Prasad

2013-01-01

A protocol has been developed for in vitro plant regeneration from a nodal explant of Dracaena sanderiana Sander ex Mast. Nodal explant showed high callus induction potentiality on MS medium supplemented with 6.78 μM 2,4-dichlorophenoxyacetic acid (2,4-D) followed by 46.5 μM chlorophenoxy acetic acid (CPA). The highest frequency of shoot regeneration (85%) and number of shoots per explant (5.6) were obtained on medium supplemented with 7.84 μM N(6)-benzylaminopurine (BA). Rooting was high on MS solid compared to liquid medium when added with 7.38 μM indole-3-butyric acid (IBA). Fifty percent of the roots were also directly rooted as microcuttings on soil rite, sand and peat mixture (1:1:1). In vitro and ex vitro raised plantlets were used for acclimatization. More than 90% of the plantlets was successfully acclimatized and established in plastic pots. Ex vitro transferred plantlets were normal without any phenotypic aberrations.

Clonal fidelity of chrysanthemum regenerated from long term cultures

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Jevremović Slađana

2006-01-01

Full Text Available Morphological characteristics of flowers of long term regenerated chrysanthemum, cv. "White Spider", after ten years of micropropagation are investigated. Shoot cultures are established and maintained more than ten years by stem segment culture on MS medium supplemented with BAP and NAA (1.0, 0.1 mgL-1, respectively. Rooting of shoots (100 % has done on MS medium without hormones and it was very successful after ten years, as well as, after two or eight years of micropropagation. Acclimation of rooted chrysanthemum plantlets at greenhouse conditions was excellent and after appropriate photoperiod "in vitro" plants flowered 90.3 % and have the same flower color, shape and size as mother plants. Flower color changes of "in vitro" plants are observed during another flowering cycle one year after acclimatization. Observed variations of chrysanthemum flowers could be attributed to epigenetic factors.

Multiple shoot regeneration of cotton (Gossypium hirsutum L.) via ...

African Journals Online (AJOL)

user

2011-03-14

Mar 14, 2011 ... Induction of multiple shoots of cotton (Gossypium hirsutum L.) plant in two commercial varieties (Sahel and Varamin) using shoot apex was done. Explants were isolated from 3 - 4 days old seedlings, then they were cultured on a shoot induction media, modified MS nutrient agar with combinations: 1- ...

Isatin as an auxin source favoring floral and vegetative shoot regeneration from calli produced by thin layer explants of tomato pedicel

Science.gov (United States)

Applewhite, P. B.; K-Sawhney, R.; Galston, A. W.

1994-01-01

Thin layer explants taken from the pedicels and peduncles of flowering tomato plants yielded calli with great organogenetic potential. Of the 15 cultivars tested, 7 regenerated roots, shoots and eventually entire fruit-bearing plants. Calli grown on modified Murashige-Skoog medium responded to varied auxins and cytokinins with different morphogenetic patterns. Thus, naphthaleneacetic acid yielded root-producing calli, while the auxin precursor isatin (indole 2,3-dione) caused the production of calli with vegetative and floral shoots, rarely yielding roots. This may be related to isatin's slow, steady conversion to an active auxin (Plant Physiol 41:1485-1488, 1966) in contrast with naphthaleneacetic acid's immediate presentation of a high level of active auxin. The highest incidence of vegetative shoot (100%) and flower (50%) formation was obtained with 10 micromoles isatin and 3 micromoles zeatin. A few of the flowers developed into ripe fruits. The high frequency of induction of vegetative shoots and flowers before roots with isatin suggests its utility in micropropagation from plant tissue cultures.

Effect of irradiance and spectral composition of radiation on in vitro shoot proliferation in Malus domestica

International Nuclear Information System (INIS)

Noè, N.; Eccher, T.; Bonini, L.

1997-01-01

Four clones of Malus domestica cv. Golden Delicious - namely Smoothee, Crielaard, Reinders and Golden B - were cultured in vitro from single-node microcuttings placed on solid medium under irradiance (PPFD) of 50 micromol m -2 s -1 . After 9 months an average shoot proliferation of 5.3 was achieved; Crielaard showed the highest rate (7.1), followed Golden B (5.4), Smoothee and Reinders (4.4). Proliferating shoots were then exposed to higher PPFD (80 micromol m -2 s -1 ) and different spectral composition of radiation using PMMA-B and PMMA-R/FR filters. High PPFD decreased the average proliferation rate to 4.5, in particular in Crielaard and Golden B, while it increased proliferation in Reinders. When a PMMA-R/FR filter was interposed, the mean proliferation rate slightly increased. PMMA-B filters decreased the overall proliferation rate to 3.0; only in Crielaard it was increased, but shoots were very small. Thus PPFD and spectral composition influenced in vitro shoot proliferation and growth and the responses were different among the clones. (author)

EFFECT OF BAP ON PROLIFERATION OF Schizolobium amazonicum Huber ex Ducke (PARICA SHOOTS, IN VITRO

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Iracema Maria Castro Coimbra Cordeiro

2004-06-01

Full Text Available Parica is a native tropical forest plant pertaining to the ofcaesalpinaceae family. It is species considered of great potential, due its wide use. Aiming reproducing the species, in vitro, stem segments of plantlets established in vitro were excised and inoculated on solid MS supplemented with 0; 1,0; 1,5; 2,0; 2,5 and 3,0 mg.L-1 of BAP with 3% of sucrose, 0,1% of PVP and one control. The activities had been carried out in the Laboratory of Genetic Resources and Biotechnology of the Embrapa Oriental, Amazon, and Belem (PA. The cultures had been under sun photo period for 16 hours 25+ 1°C of temperature. After three weeks of cultivation the explants were evaluated, counting the number and length of explants shoots. The treatment with 3 mg.L-1 BAP presented greater number of shoot proliferation with 2,14 for explants. The shoots were transferred to MS medium in the same conditions of culture for rooting.

Plants Regeneration Derived From Various on Peanut on Mutant Lines

International Nuclear Information System (INIS)

Dewi, Kumala; Masrizal; Mugiono

1998-01-01

The study of calli, greenspot formation and shoot regeneration on peanut mutant lines has ben conducted by MS media. Three explants derived from shoot tips, embryo and seeding root of two mutant lines a/20/3 and D/25/3/2 were used in this experiment. the explants were cultured on modified MS media enriched by vitamins, growth regulation, amino acids for fourth teen calli were transferred on regeneration media. The ability of calli formation and plant regeneration of each explant and genotypes of plants was varied. Greenspot and shoot formation were observed seventh days after the calli transferred on regeneration media. It is shown that the ability of calli, greenspot and shoot formation of each explants and genotypes was varied. the high ability of calli, greenspot and shoot formation were found in explant derived from shoot tip and embryo. Seedling root explant has lower ability in calli formation, while greenspot and shoot was formatted. The ability of calli, greenspot and shoot formation on A/20/3 mutant line was better than D/25/3/2 mutant line. (author)

Studies on seed germination and in vitro shoot multiplication of ...

African Journals Online (AJOL)

Yomi

2011-12-21

Dec 21, 2011 ... vitro seed germination and plantlet regeneration of this plant. ... medical books, a few varieties of herbs are described ... Seed viability was determined by tetrazolium (TZ) test. (Perry ..... Dictionary of Drugs and Difficult Terms of 'The Book of ... establishment of stem cuttings of oregano, pepermint and balm.

Shoot and root morphogenesis from Eucalyptus grandis x urophylla ...

African Journals Online (AJOL)

Eucalyptus grandis x urophylla plantlets were regenerated via indirect organogenesis. Histological assessment of their development focused on identifying the calli, the differentiation of shoots from the calli and the shoot-root junction from the nascent shoots. Vascular tissue formation within the callus preceded that of ...

Shoot organogenesis in oleaster (Elaeagnus angustifolia L.)

African Journals Online (AJOL)

STORAGESEVER

2009-02-04

Feb 4, 2009 ... Regenerated plantlets were acclimatized and successfully transplanted to soil. Key words: shoot organogenesis, callus, ... saline and alkaline soils (Economou and Maloupa, 1995). Its fruits have been used as a .... cally inert compounds as reported by Kaminek (1992). Direct shoot organogenesis without ...

Can protoplast production from in vitro cultured shoots of Tanacetum vary during the season?

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M. KESKITALO

2008-12-01

Full Text Available Two different experiments were carried out to study the production of protoplasts and the variation of protoplast yield from in vitro cultured shoot tips of tansy (Tanacetum vulgare L. and pyrethrum (Tanacetum cinerariifolium (Trevir. Schiltz-Bip. In the first experiment, light had more pronouced effect for tansy than for pyrethrum. When the donor tissues of tansy were cultured under high light intensity the leaves contained anthocyanin and became brown during enzyme maceration. In contrast, donor tissues cultured under low light intensity produced leaves without anthocyanin. Depending on the light intensity of donor tissues, on average 5.8 - 6.8 x 106 and 3.4 - 4.3 x 106 protoplasts were isolated from one gram of mesophyll leaves of tansy and pyrethrum, respectively. In the second experiment, the production of protoplasts from tansy and pyrethrum varied seasonally. The most successful season for the production of protoplasts from in vitro cultured shoot tips was between December and April, when also the highest number of protoplasts could be isolated. It was not possible to state whether Tanacetum species have rhythms, which could cause physiological or chemical changes for the in vitro grown shoot tips. However, some external or internal, possible seasonal-dependent stimuli may have caused variation in the number of protoplasts isolated from tansy and pyrethrum and favoured protoplast production during winter and spring.

Plant regeneration of bananas Ambon kuning and Barangan mutant lines were carried out by using female organ and shoot-tip as explants source

International Nuclear Information System (INIS)

Dewi, Azri K; Ishak

1998-01-01

Plant regeneration of bananas Ambon Kuning and Barangan mutant lines were carried out by using female organ and shoot-tip as explants source. Female organ was taken from heart of banana stem, while shoot-tip taken from sucker in banana plantation at Pasar Jumat, Jakarta. Those explants were cultured on MS medium containing 3 mg/l BAP, 0.5 mg/l IAA and supplemented by 100 tyrosin and 80 mg/l adenin hemisulphate. Observation showed that 180 and 42 buds were obtained from JBR 02 mutant lines respectively, while 84 and 79 buds for JAK 01 and JAK 02 respectively. The highest shoot formation was 1.013 shoots were obtained from BRC variety and lowest one was JBR 01 mutant line. statistical data analysis indicated that shoot formation between BRC variety and another mutant lines were significant difference using LSD test at level 0.05. Plantlet formation derived from female organ as well as shoot-tip showed that BRC variety produced number of plantlets per bottle was higher that another one. (author)

Regeneration of the Barley Zygote in In Vitro Cultured Ovules

DEFF Research Database (Denmark)

Holme, Inger B; Brinch-Pedersen, Henrik; Lange, Mette

2010-01-01

In vitro cultures of zygotes and small embryos carry a lot of potential for studying plant embryogenesis and are also highly relevant for plant biotechnology. Several years ago we established an in vitro ovule culture technique for barley that allows the regeneration of plants from zygotes (Holm et...... culture ability in immature embryo culture i.e. Femina, Salome and Corniche. Barley spikes were emasculated and hand pollinated 3 days after emasculation. In barley, fertilization takes place one hour after pollination and ovules with fertilized egg cells could therefore be isolated one hour after...... pollination. Ovules were grown for 3 weeks on a culture medium where after embryos could be isolated and transferred to regeneration medium. An average of 1.2 green plantlets per ovule could be regenerated from 50 % of the isolated ovules. No genotypic differences were found on embryo induction...

Efficient in vitro propagation of Artemisia nilagirica var. nilagirica (Indian wormwood) and assessment of genetic fidelity of micropropagated plants.

Science.gov (United States)

Shinde, Smita; Sebastian, Joseph Kadanthottu; Jain, Jyothi Ramesh; Hanamanthagouda, Manohar Shirugumbi; Murthy, Hosakatte Niranjana

2016-10-01

A reliable protocol has been established for in vitro propagation of Artemisia nilagirica var. nilagirica (Indian wormwood), a valuable medicinal plant from India. A highly proliferating organogenic callus was obtained on Murashige and Skoog (MS) medium supplemented with 2.5 µM IAA when nodal explants were cultured on MS medium supplemented with various growth regulators. Further, highest regeneration frequency (83.3 %) of adventitious shoots was observed, when the callus was sub-cultured on MS medium supplemented with 6-benzylaminopurine (BAP; 2.5 µM) along with 7.5 µM 2-isopentenyl adenine (2-iP). An optimal of 10.16 ± 2.24 shoots were regenerated on medium supplemented with 2.5 µM BAP + 7.5 µM 2-iP. Quarter strength MS medium supplemented with 10 µM IBA was effective for rooting of the shoots. Ex-vitro plants were normal and were established successfully. Cytological and molecular marker studies showed that regenerated plants showed genetic stability in micro-propagated plants.

Effect of cytokinins on in vitro multiplication, volatiles composition and rosmarinic acid content of Thymus leucotrichus Hal. shoots.

Science.gov (United States)

Bekircan, Tuba; Yaşar, Ahmet; Yıldırım, Sercan; Sökmen, Münevver; Sökmen, Atalay

2018-03-01

An efficient in vitro multiplication protocol was designed to Thymus leucotrichus , a subshrub and perennial herb growing naturally in the Northwest of Turkey. Of all basal media studied, Murashige and Skoog medium was found to be superior to the others, providing higher shoot formation and the maximum shoot length. Varying concentrations of cytokinins, i.e., 6-benzyladenine, thidiazuron, 2-isopentenyladenine and kinetin were supplemented in the nutrient media to observe their effects on shoot development and biomass. Rosmarinic acid content and volatile compositions of both naturally growing plants and in vitro multiplied plantlets were also evaluated. 6-benzyladenine (1.0 mg/L) and kinetin (0.5 mg/L) were found to be optimum for shoot number and shoot elongation, respectively. Thidiazuron (1.0 mg/L) was superior for biomass production. Rosmarinic acid content of in vitro multiplied plants was found to be higher than that of wild plants, reaching a maximum with 0.5 mg/L 2-isopentenyladenine, which yielded 10.15 mg/g dry weight. The highest thymol content was obtained with 1.0 mg/L kinetin (55.82%), while thidiazuron (0.1 mg/L) increased carvacrol production (12.53%). Overall, Murashige and Skoog medium supplemented with 1.0 mg/L kinetin was determined to be the most favorable medium studied.

Effect of 60Co γ irradiation with seed and shoot-tip of Brassica campestris L. var on its culture in vitro

International Nuclear Information System (INIS)

Liao Feixiong; Yu Rangcai; Pan Ruichi

2003-01-01

The survival rate in vitro of shoot-tips of Brassica campestris L. var from seeds irradiated by 60 Co γ-rays decreased with the increase of dose. Irradiation inhibited proliferation of shoot-tip, induction of callus from cotyledons and differentiation of the callus. The age of explant contributed to the effect of irradiation in the culture. Irradiation stimulated the proliferation of shoot-tip with dose less than 50 Gy. Based on the effect of irradiation in the tissue culture, the effective dose recommended was about 200 Gy for seeds, 50-100 Gy for pre-soaked germinating seeds and 40-70 Gy for shoot-tips in vitro, respectively

Growing Arabidopsis in vitro: cell suspensions, in vitro culture, and regeneration.

Science.gov (United States)

Barkla, Bronwyn J; Vera-Estrella, Rosario; Pantoja, Omar

2014-01-01

An understanding of basic methods in Arabidopsis tissue culture is beneficial for any laboratory working on this model plant. Tissue culture refers to the aseptic growth of cells, organs, or plants in a controlled environment, in which physical, nutrient, and hormonal conditions can all be easily manipulated and monitored. The methodology facilitates the production of a large number of plants that are genetically identical over a relatively short growth period. Techniques, including callus production, cell suspension cultures, and plant regeneration, are all indispensable tools for the study of cellular biochemical and molecular processes. Plant regeneration is a key technology for successful stable plant transformation, while cell suspension cultures can be exploited for metabolite profiling and mining. In this chapter we report methods for the successful and highly efficient in vitro regeneration of plants and production of stable cell suspension lines from leaf explants of both Arabidopsis thaliana and Arabidopsis halleri.

Isolation and propagation of mutations in dahlia by in vitro culture

International Nuclear Information System (INIS)

Asahira, T.; Yamagata, H.; Inagaki, M.; Osuga, S.

1975-01-01

The present study was undertaken to search for a successful method for in vitro culture of mutated tissues in dahlia. Preceding the objective, the features of induced mutations and the effects of cutting propagation in dahlia were investigated, and the tissues easily regenerating plantlets in vitro were searched following the examination of effective condition of medium. Induction of mutations: Tuberous roots of two cultivars, 'Kosei' and 'Sunlight', were irradiated with 1,000 - 2,000 R of X-rays. Chlorophyll and flower-color mutations were successfully induced in both cultivars, but the frequency differed with genotypic constitution. The maximum frequency was observed at leaves and shoots on or from the fourth to fifth nodes from the base of plant. The use of M 1 tuberous roots seemed a way for isolating mutations though not so much efficient. Tissue culture: In vitro cultured basal parts of ray florets, exactly the ovary, differentiated shoots. No shoot formation occurred in receptacle and leaf cultures, while roots were differentiated in leaf culture. Supplements of auxin and adenine to the medium besides cytokinin appeared to be necessary for inducing shoots. It is a serious problem in the tissue culture of dahlia that a large number of explants are endogenously comtaminated with bacteria. Taking into consideration low rates of surviving and regenerating explants, it seems difficult at present for dahlia to conclude whether or not the tissue culture may become efficient in mutation breeding as compared with cutting propagation. (author)

Development of salt tolerant potato and sugarcane through in vitro techniques

International Nuclear Information System (INIS)

Asad, S.; Iqbal, M.J.; Saif-Ur-Rasheed, M.; Zafar, Y.; Malik, K.A.

1997-01-01

Improvement of sugarcane and potato in Pakistan is hampered by their intricate flowering behavior under natural day-length conditions. The improvement of these crops for their salt tolerance can be carried out by tissue culture mediated techniques. To induce variation in sugarcane, five-week old white yellow nodular embryogenic calli were irradiated with 5, 20, 40 and 60 Gy gamma rays. After one month, the calli were cultured on regeneration media, and plant hardening procedures were optimized. Irradiated and non-irradiated calli were subjected to various levels of salt stress and plant regeneration was investigated. Although growth of sugarcane calli was observed at 200 mM NaCl, regeneration was inhibited even at 50 mM NaCl in the medium. The regenerants from gamma irradiated material are under field evaluation. Variation was detected in both irradiated and salt treated calli by DNA fingerprinting using random amplified polymorphic DNA (RAPD) markers. In potato, 6-7 weeks old in vitro grown plants with single shoots having 8-10 buds were irradiated with 20 Gy gamma rays. Shoot formation was successful only from 50% of the axillary buds. The cultures were subjected to four levels of salinity (50, 100, 150 and 200 mM Nacl). Shoot-tip necrosis was observed along with significant reduction in shoot height. (author). 13 refs, 3 figs, 4 tabs

Development of salt tolerant potato and sugarcane through in vitro techniques

Energy Technology Data Exchange (ETDEWEB)

Asad, S; Iqbal, M J; Saif-Ur-Rasheed, M; Zafar, Y; Malik, K A [Plant Biotechnology Div., National Inst. of Biotechnology and Genetic Engineering, Faisalabad (Pakistan)

1997-07-01

Improvement of sugarcane and potato in Pakistan is hampered by their intricate flowering behavior under natural day-length conditions. The improvement of these crops for their salt tolerance can be carried out by tissue culture mediated techniques. To induce variation in sugarcane, five-week old white yellow nodular embryogenic calli were irradiated with 5, 20, 40 and 60 Gy gamma rays. After one month, the calli were cultured on regeneration media, and plant hardening procedures were optimized. Irradiated and non-irradiated calli were subjected to various levels of salt stress and plant regeneration was investigated. Although growth of sugarcane calli was observed at 200 mM NaCl, regeneration was inhibited even at 50 mM NaCl in the medium. The regenerants from gamma irradiated material are under field evaluation. Variation was detected in both irradiated and salt treated calli by DNA fingerprinting using random amplified polymorphic DNA (RAPD) markers. In potato, 6-7 weeks old in vitro grown plants with single shoots having 8-10 buds were irradiated with 20 Gy gamma rays. Shoot formation was successful only from 50% of the axillary buds. The cultures were subjected to four levels of salinity (50, 100, 150 and 200 mM Nacl). Shoot-tip necrosis was observed along with significant reduction in shoot height. (author). 13 refs, 3 figs, 4 tabs.

In vitro embryo rescue and plant regeneration following self ...

African Journals Online (AJOL)

In vitro embryo rescue and plant regeneration following self-pollination with irradiated ... AFRICAN JOURNALS ONLINE (AJOL) · Journals · Advanced Search ... shows that pollen irradiation coupled with self-pollination and embryo rescue ...

In vitro regeneration and morphogenesis in Phyllanthus niruri L., an ...

African Journals Online (AJOL)

Kingsley

2012-10-04

Oct 4, 2012 ... The high morphogenetic response of Phyllanthus niruri in vitro can be used to propagate ... filled with a mixture of soil-cow dung-coconut husk in a ratio of 3:1:1 ... examined for the presence of shoots, roots or somatic embryos.

In vitro propagation of Homalomena aromatica Schott., an endangered aromatic medicinal herb of Northeast India.

Science.gov (United States)

Raomai, Shiveirou; Kumaria, Suman; Tandon, Pramod

2013-04-01

A successful report on the in vitro propagation of Homalomena aromatica via rhizome axillary bud multiplication is presented. Rhizome bud explants were cultured on Murashige and Skoog medium supplemented with various concentrations of cytokinins to induce multiple shoot formation for micropropagation. The highest number of shoots was achieved in MS medium supplemented with 2.0 mg l(-1) 6-benzylaminopurine. The regenerated shoots rooted most efficiently on half-strength MS medium supplemented with 0.5 mg l(-1) α-naphthalene acetic acid. The regenerated plantlets showed no morphological differences from the parent plant. This protocol takes approximately 6 months to reach the acclimatization stage from the initiation stage and facilitates commercial and rapid propagation of H. aromatica.

Influence of gamma ray treatment on in vitro regenerated plants of Atropa Belladonna L

International Nuclear Information System (INIS)

Toth, E.; Onisei, T.; Amariei, D.

1994-01-01

Regenerated plants were obtained through callus organogenesis after gamma ray treatment with 1 to 9 Krad doses. Vigorous shoots were regenerated on MS (1962) medium supplemented with 2 mg/l 6-benzylaminopurine. Shoot growth was inhibited with increasing irradiation doses. At 2 Krad dose root Primordia were observed after 18 days while 32 days were necessary to produce roots at 7-9 Krad. As compared to the control, the survival percent during acclimatization was 80 to 85% at 2-3 Krad, 65-70% at 4-6 Krad and 60-62% at 7-9 Krad. Approximately 330 plants were transferred to field conditions. Morphological and biochemical parameters were measured and the data were statistically processed. Plant height was negatively influenced by higher doses of irradiation. A chlorophyll deficient plant arose from the 6 Krad treatment which showed a different pattern of isoperoxidase and isoesterase as compared to the control. A yellow-flower mutant was obtained from the 3 Krad treatment and is assumed to be Atropa belladonna var. Flava Pater. (author)

Micropropagation of nucellar embryos and their histological comparative study for regeneration ability with other explants of kinnow mandarin (Citrus reticulata Blanco)

International Nuclear Information System (INIS)

Kazmi, S.K.; Khan, S.; Kabir, N.

2018-01-01

Polyembryony is the most beneficial and distinct character in citrus seeds. This characteristic can be beneficial in citrus improvement programs. Nucellar embryos developed from nucellar wall in citrus seeds along with zygotic embryo are found to have high plants regeneration ability in comparison to zygotic embryos. Under In vitro culture conditions, nucellar embryos were detected and multiplied on MT media with 0.5ml of coconut water along with 50g/L sucrose. These nucellar embryonic tissues were also regenerated on MT media upplemented with 0.5 mg/L BAP and 0.5 mg/L kinetin. Many shoots were regenerated from nucellar embryonic tissue; these shoots were rooted on MT media supplemented with IBA 0.5mg/L and NAA 0.1 mg/L in addition to activated charcoal 0.5gm/L. The resulting plantlets were acclimatized in the green house. After micropropagation, histological studies of nucellar embryonic tissues were carried out under fluorescence microscope to examine their high regeneration ability in comparison with usual plant parts, like seeds, shoots as well as with tissue culture stages including embryogenic and non-embryogenic callus. It was found that nucellar embryos have more regeneration ability as compared to usual plant parts and other tissue culture stages. (author)

Regeneração in vitro de louro-pardo (Cordia trichotoma (Vellozo Arrabida ex Steudel

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Nilton César Mantovani

2001-01-01

Full Text Available This study was carried out aiming at evaluating the in vitro regeneration of louro-pardo (Cordia trichotoma (Vellozo Arrabida ex Steudel. Nodal segments were desinfested and innoculated in two basal media: WPM (Wood Plant Médium, 1981 and MS (Murashige & Skoog, 1962 supplemented with BAP (6-Benzylaminopurine, TDZ (Thidiazuron, NAA (Naphthaleneacetic Acid, GA3 (Gibberellic Acid, IBA (Indole-3-Butyric Acid and A.C. (Activated Charcoal in different combinations and concentrations for each developmental step. The regeneration rate and the shoot elongation were higher in the WPM medium containing 0,1 mg.l-1 of BAP and GA3. The rooting of regenerated shoots was observed in IBA (0,5 mg.l-1 and Ativated Charcoal (1,5 g.l-1.

Physiological Response of In Vitro Cultured MAGNOLIA SP. to Nutrient Medium Composition

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S. Sokolov Rossen

2014-09-01

Full Text Available The objective of this study was to assess the regeneration response of in vitro cultured Magnolia × soulangeana ‘Alexandrina’ and Magnolia liliiflora ‘Nigra’ to nutrient medium composition. In the primary culture (initiated from dormant axillary buds combinations of Murashige and Skoog (MS basal salts with 6-benzylaminopurine and α-naphthaleneacetic acid were tested. The primary explants of cv. ‘Alexandrina’ expressed higher regeneration rate than cv. ‘Nigra’. For both species, the regen eration was most strongly potentiated at addition of 0.25 mg dm−3 of the cytokinin alone. The auxin exerted undesir–able effects. Several basal salts media were applied in proliferation stage and their physiological effects were evaluated in reference to traditionally used MS. At culturing on Chée & Pool C2d Vitis Medium (VM that is for the first time introduced to magnolia and on MS, M. liliiflora formed more but less elongated shoots than M. soulangeana. However, on VM, substantial increase (25-30% of the number of axillary shoots and leaves, shoot length and fresh and dry weights over MS was established for both species. This suggested VM as promising composition of nutrients in multiplication stage. Microshoots obtained on MS, VM, Rugini Olive Medium and DKW Juglans Medium were successfully rooted in vitro and subsequently established ex vitro. The findings expand the information on magnolia response to culture conditions and contribute to elaboration of innovative elements of protocols for establishing tissue cultures with high regeneration capacity.

Germination and In Vitro Regeneration Response of Local Nigerian ...

African Journals Online (AJOL)

Prof. Ogunji

the highest germination percentage of 57.33% and percentage contamination of. 11.11% ... development of in vitro regeneration of the crop from various explant sources. The advent of ..... Effect of variety and plant growth regulators on callus.

Reproducible in vitro regeneration system for purifying sugarcane ...

African Journals Online (AJOL)

This procedure may be considered as one of the best ever published report on regeneration from in vitro grown plants to purify clones without subjecting the plants to field conditions and harvesting the mature cane. This technique was used to purify transgenic sugarcane plants carrying Bacillus thuringiensis gene.

In vitro propagation and production of cardiotonic glycosides in shoot cultures of Digitalis purpurea L. by elicitation and precursor feeding.

Science.gov (United States)

Patil, Jitendra Gopichand; Ahire, Mahendra Laxman; Nitnaware, Kirti Manik; Panda, Sayantan; Bhatt, Vijay P; Kishor, Polavarapu B Kavi; Nikam, Tukaram Dayaram

2013-03-01

Digitalis purpurea L. (Scrophulariaceae; Foxglove) is a source of cardiotonic glycosides such as digitoxin and digoxin which are commercially applied in the treatment to strengthen cardiac diffusion and to regulate heart rhythm. This investigation deals with in vitro propagation and elicited production of cardiotonic glycosides digitoxin and digoxin in shoot cultures of D. purpurea L. In vitro germinated seedlings were used as a primary source of explants. Multiple shoot formation was achieved for three explant types (nodal, internodal, and leaf) cultured on Murashige and Skoog (MS) medium with several treatments of cytokinins (6-benzyladenine-BA; kinetin-Kin; and thidiazuron-TDZ) and auxins (indole-3-acetic acid-IAA; α-naphthaleneacetic acid-NAA; and 2,4-dichlorophenoxy acetic acid-2,4-D). Maximum multiple shoots (12.7 ± 0.6) were produced from nodal explants on MS + 7.5 μM BA. Shoots were rooted in vitro on MS containing 15 μM IAA. Rooted plantlets were successfully acclimatized. To further maintain the multiple shoot induction, mother tissue was cut into four equal parts and repeatedly sub-cultured on fresh shoot induction liquid medium after each harvest. On adaptation of this strategy, an average of 18 shoots per explant could be produced. This strategy was applied for the production of biomass and glycosides digitoxin and digoxin in shoot cultures on MS medium supplemented with 7.5 μM BA and several treatments with plant growth regulators, incubation period, abiotic (salicylic acid, mannitol, sorbitol, PEG-6000, NaCl, and KCl), biotic (Aspergillus niger, Helminthosporium sp., Alternaria sp., chitin, and yeast extract) elicitors, and precursors (progesterone, cholesterol, and squalene). The treatment of KCl, mycelial mass of Helminthosporium sp., and progesterone were highly effective for the production of cardenolides. In the presence of progesterone (200 to 300 mg/l), digitoxin and digoxin accumulation was enhanced by 9.1- and 11.9-folds

Establishment of an in vitro micropropagation protocol for Boscia senegalensis (Pers.) Lam. ex Poir.

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Khalafalla, Mutasim M; Daffalla, Hussien M; Abdellatef, Eltayb; Agabna, Elsadig; El-Shemy, Hany A

2011-04-01

This report describes in vitro micropropagation of Boscia senegalensis, so-called famine foods, that helped the people in Darfur and Kordofan, Sudan survive during the 1984-1985 famine. Four types of explants prepared from green mature zygotic embryos were cultured on Murashige and Skoog (MS) medium augmented with 1-5 mg/L 6-benzyladenine (BA). The highest number of shoots per explant (14.3±0.9) was achieved on MS medium supplemented with 3 mg/L BA, while the highest shoot length [(3.5±0.4) cm] was obtained with 1 mg/L BA. The shoot cluster, when subcultured to its same medium, significantly increased the rate of shoot multiplication by the end of the third subculture. The maximum mean number of shoots per explant (86.5±3.6) was produced after three multiplication cycles on 3 mg/L BA-supplemented medium. In vitro induced shoots were excised and rooted on half strength MS medium fortified with 0.25 mg/L indole-3-butyric acid (IBA) to obtain complete plantlets. B. senegalensis-regenerated plantlets obtained in vitro for the first time, were hardened and 95% survived under greenhouse conditions.

New potential markers of in vitro tomato morphogenesis identified by mRNA differential display.

Science.gov (United States)

Torelli, A; Soragni, E; Bolchi, A; Petrucco, S; Ottonello, S; Branca, C

1996-12-01

The identification of plant genes involved in early phases of in vitro morphogenesis can not only contribute to our understanding of the processes underlying growth regulator-controlled determination, but also provide novel markers for evaluating the outcome of in vitro regeneration experiments. To search for such genes and to monitor changes in gene expression accompanying in vitro regeneration, we have adapted the mRNA differential display technique to the comparative analysis of a model system of tomato cotyledons that can be driven selectively toward either shoot or callus formation by means of previously determined growth regulator supplementations. Hormone-independent transcriptional modulation (mainly down-regulation) has been found to be the most common event, indicating that a non-specific reprogramming of gene expression quantitatively predominates during the early phases of in vitro culture. However, cDNA fragments representative of genes that are either down-regulated or induced in a programme-specific manner could also be identified, and two of them (G35, G36) were further characterized. One of these cDNA fragments, G35, corresponds to an mRNA that is down-regulated much earlier in callus- (day 2) than in shoot-determined explants (day 6). The other, G36, identifies an mRNA that is transiently expressed in shoot-determined explants only, well before any macroscopic signs of differentiation become apparent, and thus exhibits typical features of a morphogenetic marker.

In vitro cultures of Salvia officinalis L. as a source of antioxidant compounds

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Izabela Grzegorczyk

2011-01-01

Full Text Available The concentrations of carnosic acid, carnosol and rosmarinic acid in different materials from differentiated (multiple shoot cultures and regenerated plants and undifferentiated (callus and cell suspension in vitro cultures of Salvia officinalis were determined by HPLC. The results suggested that diterpenoid (carnosic acid and carnosol production is closely related to shoot differentiation. The highest diterpenoid yield (11.4 mg g-1 for carnosic acid and 1.1 mg g-1 for carnosol was achieved in shoots of 10-week-old micropropagated plants. The levels were comparable to those found in shoots of naturally growing plants. Undifferentiated callus and cell suspension cultures produced only very low amounts of carnosol (ca. 0.05 mg g-1 of dry weight. In contrast, content of rosmarinic acid in callus and suspension cultures as well as shoots growing in vitro and in vivo was similar and ranged between 11.2 and 18.6 mg g-1 of dry weight.

In vitro regeneration of lychee (Litchi chinensis Sonn.)

African Journals Online (AJOL)

Administrator

In vitro plantlet regeneration in the main commercial variety of lychee (Litchi chinensis Sonn. cv Tai So) in Mauritius was achieved from callus cultures derived from young, tender leaf explants on Murashige and Skoog (MS) medium. Callogenesis was obtained in all media supplemented with auxin, but was most prominent ...

Development of efficient plant regeneration and transformation system for impatiens using Agrobacterium tumefaciens and multiple bud cultures as explants.

Science.gov (United States)

Dan, Yinghui; Baxter, Aaron; Zhang, Song; Pantazis, Christopher J; Veilleux, Richard E

2010-08-09

Impatiens (Impatiens walleriana) is a top selling floriculture crop. The potential for genetic transformation of Impatiens to introduce novel flower colors or virus resistance has been limited by its general recalcitrance to tissue culture and transformation manipulations. We have established a regeneration and transformation system for Impatiens that provides new alternatives to genetic improvement of this crop. In a first step towards the development of transgenic INSV-resistant Impatiens, we developed an efficient plant regeneration system using hypocotyl segments containing cotyledonary nodes as explants. With this regeneration system, 80% of explants produced an average of 32.3 elongated shoots per initial explant plated, with up to 167 elongated shoots produced per explant. Rooting efficiency was high, and 100% of shoots produced roots within 12 days under optimal conditions, allowing plant regeneration within approximately 8 weeks. Using this regeneration system, we developed an efficient Agrobacterium-mediated Impatiens transformation method using in vitro multiple bud cultures as explants and a binary plasmid (pHB2892) bearing gfp and nptII genes. Transgenic Impatiens plants, with a frequency up to 58.9%, were obtained within 12 to 16 weeks from inoculation to transfer of transgenic plants to soil. Transgenic plants were confirmed by Southern blot, phenotypic assays and T1 segregation analysis. Transgene expression was observed in leaves, stems, roots, flowers, and fruit. The transgenic plants were fertile and phenotypically normal. We report the development of a simple and efficient Agrobacterium-mediated transformation system for Impatiens. To the best of our knowledge, there have been no reports of Agrobacterium-mediated transformation of Impatiens with experimental evidence of stable integration of T-DNA and of Agrobacterium-mediated transformation method for plants using in vitro maintained multiple bud cultures as explants. This transformation system

Development of Efficient Plant Regeneration and Transformation System for Impatiens Using Agrobacterium tumefaciens and Multiple Bud Cultures as Explants

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Dan Yinghui

2010-08-01

Full Text Available Abstract Background Impatiens (Impatiens walleriana is a top selling floriculture crop. The potential for genetic transformation of Impatiens to introduce novel flower colors or virus resistance has been limited by its general recalcitrance to tissue culture and transformation manipulations. We have established a regeneration and transformation system for Impatiens that provides new alternatives to genetic improvement of this crop. Results In a first step towards the development of transgenic INSV-resistant Impatiens, we developed an efficient plant regeneration system using hypocotyl segments containing cotyledonary nodes as explants. With this regeneration system, 80% of explants produced an average of 32.3 elongated shoots per initial explant plated, with up to 167 elongated shoots produced per explant. Rooting efficiency was high, and 100% of shoots produced roots within 12 days under optimal conditions, allowing plant regeneration within approximately 8 weeks. Using this regeneration system, we developed an efficient Agrobacterium-mediated Impatiens transformation method using in vitro multiple bud cultures as explants and a binary plasmid (pHB2892 bearing gfp and nptII genes. Transgenic Impatiens plants, with a frequency up to 58.9%, were obtained within 12 to 16 weeks from inoculation to transfer of transgenic plants to soil. Transgenic plants were confirmed by Southern blot, phenotypic assays and T1 segregation analysis. Transgene expression was observed in leaves, stems, roots, flowers, and fruit. The transgenic plants were fertile and phenotypically normal. Conclusion We report the development of a simple and efficient Agrobacterium-mediated transformation system for Impatiens. To the best of our knowledge, there have been no reports of Agrobacterium-mediated transformation of Impatiens with experimental evidence of stable integration of T-DNA and of Agrobacterium-mediated transformation method for plants using in vitro maintained

Rapid Screening of In-Vitro Regenerated Plantlets of Four Nigerian ...

African Journals Online (AJOL)

Prof. Ogunji

Tissue culture technique provides a rapid means of studying plant-pathogen interaction in a controlled ... Keywords: cowpea, in-vitro regeneration, hormones, Fusarium wilt, protocols. .... solanacearum and evaluation of tomato in Nepal.

Researches concerning in vitro cultures optimization of the vulnerable species Dianthus nardiformis Janka

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Irina HOLOBIUC

2010-05-01

Full Text Available Our aim was to establish an optimized protocol for in vitro short-term conservation of the vulnerable species Dianthus nardiformis Janka belonging to Romanian Flora.For our purpose, different culture media compositions were tested: complex variants added with plant growth regulators and simplified media with low levels of macro- and microelements.This species had a good in vitro reaction, the regeneration occurs through direct morphogenesis as in all Dianthus species. Generally, the use of variants of MS medium supplemented with cytokinin/ auxin at 10/1 ratio proved to induce high regeneration response starting from the first month of culture.In our study, MS media complex variants with diminished levels of plant growth hormones at the same ratio of cytokinins/auxins conducted to acceptable levels of regeneration.By the other hand on complex regeneration variants, the rooting process was absent or underwent at low levels.Using simplified media with reduced levels of macro- and microelements, both rhizogenesis and shoots formation were induced. The in vitro response was characterized through the mean number of regenerated shoots/explants and the mean number of neoformed roots/initial inoculum. Despite of low micro- and macroelements levels (1/2 and 1/4 of MS salts, the modified MS basal medium composition proved to be beneficial for D. nardiformis, plants could be efficiently regenerated and rooted and regenerative aggregates can also be propagated as short-term cultures. The need of the supplementary transfer on particular rooting medium is also eliminated.Using simplified culture media, this species can be efficiently micropropagated and maintained as active tissue cultures, able tosupply viable plants whenever it is necessary.

Radiosensitivity of in vitro Cultured Banana Shoot-Tips

International Nuclear Information System (INIS)

Elagamawy, M.R.

2002-01-01

Longitudinally dissected shoot apices of Grand Nain, Williams and Maghrabi banana cultivars were exposed to gamma irradiation with Cobalt 60 source at the doses of 0, 20, 40 and 60 Gy and immediately placed into proliferation medium. A number of micropropagation cycles after irradiation were necessary up to M1 V2 to M1 V4 stage to let mutated sectors developing into non-chimeric shoots. Radiosensitivity was evaluated by the rate of shoot proliferation and by the shoot fresh weight increase. Increasing gamma doses caused reduction in survival rates and average number of shoots. Grand Nain exhibited the highest multiplication- rate (3.1). The lower dose (20 Gy) enhanced shoot multiplication ratio specially in Williams and Maghrabi, which however decreased with increased doses. The doses of 20-40 Gy yielded Ld50, with sensible degree of shoot multiplication, which occurred hardly ever beyond 40 Gy. The dose of 60 Gy resulted 80% lethal shoot growth. Linear decrease in fresh weight was observed in post-irradiation recovery, notably in the Maghrabi. In contradictory vulnerable damage was observed in Williams which showed the highest fresh weight value. Shoot proliferation appeared generally on the surface of the corm. Root formation was observed without additional hormone. The roots were dark colored and was decreased with the increased of dosage

Establishment of an efficient in vitro regeneration protocol for rapid and mass propagation of Dendrobium chrysotoxum Lindl. using seed culture.

Science.gov (United States)

Nongdam, Potshangbam; Tikendra, Leimapokpam

2014-01-01

An efficient in vitro regeneration protocol from seed culture has been established successfully for Dendrobium chrysotoxum, an epiphytic orchid having tremendous ornamental and medicinal values. Seed germination response was encouraging in Mitra (M) medium enriched with different combinations of auxins and cytokinins. Medium supplemented with 0.4% activated charcoal (AC), 2 mg/L 6-benzyl amino purine (BAP), and 2 mg/L indole-3-acetic acid (IAA) produced best seed germination percentage in 2 weeks of culture. Incorporation of higher concentration of kinetin (KN) or BAP in combination with low auxin in medium induced pronounced shooting and leaf formation. Reduction in leaf development was evident when cytokinins exist singly in medium indicating synergistic effect of auxin and cytokinin in leaf induction. Presence of elevated level of indole-3-butyric acid (IBA) or 1-naphthalene acetic acid (NAA) with low cytokinin content in medium generated more in vitro rooting, though IBA was found to be more effective in rooting induction as compared to NAA. The in vitro protocol for asymbiotic seed germination developed from the present investigation can be used for rapid mass propagation of this highly important Dendrobium orchid species.

Establishment of an Efficient In Vitro Regeneration Protocol for Rapid and Mass Propagation of Dendrobium chrysotoxum Lindl. Using Seed Culture

Science.gov (United States)

2014-01-01

An efficient in vitro regeneration protocol from seed culture has been established successfully for Dendrobium chrysotoxum, an epiphytic orchid having tremendous ornamental and medicinal values. Seed germination response was encouraging in Mitra (M) medium enriched with different combinations of auxins and cytokinins. Medium supplemented with 0.4% activated charcoal (AC), 2 mg/L 6-benzyl amino purine (BAP), and 2 mg/L indole-3-acetic acid (IAA) produced best seed germination percentage in 2 weeks of culture. Incorporation of higher concentration of kinetin (KN) or BAP in combination with low auxin in medium induced pronounced shooting and leaf formation. Reduction in leaf development was evident when cytokinins exist singly in medium indicating synergistic effect of auxin and cytokinin in leaf induction. Presence of elevated level of indole-3-butyric acid (IBA) or 1-naphthalene acetic acid (NAA) with low cytokinin content in medium generated more in vitro rooting, though IBA was found to be more effective in rooting induction as compared to NAA. The in vitro protocol for asymbiotic seed germination developed from the present investigation can be used for rapid mass propagation of this highly important Dendrobium orchid species. PMID:25401154

Improved regeneration and transformation protocols for three strawberry cultivars

Science.gov (United States)

Zakaria, Hossam; Hussein, Gihan M; Abdel-Hadi, Abdel-Hadi A; Abdallah, Naglaa A

2014-01-01

Strawberry (Fragaria × ananassa) is an economically important soft fruit crop with polyploid genome which makes the breeding of new cultivars difficult. Simple and efficient method for transformation and regeneration is required for cultivars improvement in strawberry. In the present study, adventitious shoot regeneration has been investigated in three cultivated strawberry plants, i.e., Festival, Sweet Charly and Florida via direct organogenesis using the in vitro juvenile leaves as explants. Explants were collected after sub-culturing on a propagation medium composed of MS supplemented with 0.5 mg/l BA; 0.1 mg/l GA3 and 0.1 mg/l IBA. To select the suitable organogenesis, the explants of the three cultivars were cultured on MS medium supplemented with different concentrations of TDZ (1, 2, 3, and 4 mg/l), then incubated at a temperature of 22 °C ± 2. Medium containing 2 mg/l TDZ revealed the best regeneration efficiency with the three cultivars (72% for Festival, and 73% for Sweet Charly and Florida). After 4 weeks, the produced shoots were cultured on MS medium with different concentrations of BA and Kin to enhance shoot elongation. Results showed that the medium containing 1.5 mg/l BA and 0.5 mg/l Kin revealed highest elongation efficiency (88% and 94%) for Festival and Sweet Charly, respectively. On the other hand, medium containing 1.5 mg/l BA and 0.1 mg/l Kin showed highest elongation efficiency (90%) in Florida. Elongated shoots were successfully rooted on MS medium containing 1.5 mg/l NAA. Furthermore, transformation of the two cultivars, Festival and Sweet Charly, has been established via Agrobacterium strain LBA44404 containing the plasmid pISV2678 with gus-intron and bar genes. Three days post co-cultivation, GUS activity was screening using the histochemical assay. The results showed 16% and 18% of the tested plant materials has changed into blue color for Festival and Sweet Charly, respectively. Out of 120 explants only 13 shoots were developed on

Plant regeneration protocol of Andrographis paniculata (Burm. f ...

African Journals Online (AJOL)

Plant regeneration protocol of Andrographis paniculata (Burm. f.) - an important medicinal plant. ... Inclusion of 1.0 mg/l 1-naphthalene acetic acid (NAA) in the culture medium along with BA + Ads promoted a higher rate of shoot bud regeneration. Maximum mean number of shoot bud per explant (28.6) was achieved on the ...

A high-frequency in vitro multiplication, micromorphological studies and ex vitro rooting of Cadaba fruticosa (L.) Druce (Bahuguni): a multipurpose endangered medicinal shrub.

Science.gov (United States)

Lodha, Deepika; Patel, Ashok Kumar; Shekhawat, N S

2015-07-01

An efficient and reproducible in vitro propagation protocol has been established for Cadaba fruticosa (L.) Druce. Surface-sterilized nodal stem segments of mature plant were used as explants for culture establishment. Multiple shoots were optimally differentiated from the nodal stem explants through bud breaking on Murashige and Skoog (1962) medium containing 3.0 mg l(-1) benzyladenine (BA). The effect of different plant growth regulators and minerals were studied on different stages of micropropagation procedure (i.e., explant establishment, shoot multiplication/growth and ex vitro rooting). Additionally, for enhancing shoot multiplication during subculture, MS medium was modified (MMS) with higher levels of magnesium, potassium and sulphate ions. Out of these, MMS3 medium containing 0.25 mg l(-1) each of BA and Kin (N6-furfuryladenine), with 0.1 mg l(-1) NAA (α-naphthalene acetic acid) was found the best for shoot multiplication (42.45 ± 3.82 per culture vessel). The in vitro regenerated shoots were rooted under ex vitro conditions on treating the shoot base with 500 mg l(-1) of IBA (indole-3 butyric acid) for 3 min on sterile Soilrite®. The ex vitro rooted plants were hardened in the greenhouse and transferred to the field with ≈85 % survival rate. There were not any visual differences between wild and micropropagated plants in the field, although the later underwent significant changes during acclimatization. Micromorphological changes on leaf surface characters from in vitro to acclimatized plantlets were studied in terms of development of glandular trichomes, changes in vein spacing and vein structure in order to understand the nature of plant responses towards environmental conditions. The method developed and defined can be applied for commercial cultivation, which may be important for extraction of bioactive compounds and may facilitate conservation of this multipurpose endangered medicinal shrub.

Indução de superbrotamento e regeneração de plantas in vitro, nas cultivares de algodão colorido Induction of multiple shoots and regeneration of plants in vitro, in coloured cotton cultivars

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Maria do S. Rocha

2008-10-01

Full Text Available A micropropagação in vitro tem-se apresentado como técnica que possibilita várias metodologias que, por sua vez, contribuem com a redução no tempo, para a obtenção de novas cultivares. Objetivou-se com esse trabalho avaliar o comportamento dos genótipos BRS-Verde, BRS-200-Marrom, 6M-Mocó-Branco e BRs(-187 8H Branco na indução do superbrotamento em diferentes combinações de reguladores de crescimento. As sementes, para obtenção do material de partida, foram desinfectadas em solução de hipoclorito de sódio a 1% de cloro ativo por 20 min. Os brotos foram induzidos, a partir de explante de nós cotiledonares em meio básico MS, suplementado com 6-benzylaminopurine (BAP, Cinetina ( KIN e Tiadiazuron (TDZ, isolados ou associados em diferentes concentrações. O material foi mantido por 40 dias em sala de crescimento, sob condições ambientais controladas. Utilizou-se 10 tubos de ensaio por tratamento com um explante por frasco, em um delineamento inteiramente casualizado, com arranjo fatorial de 4 x 17 (quatro genótipos x dezessete meios. Observou-se que o meio MS suplementado com BAP (2,0 mg L-1 isolado ou associado com KIN (1,0 mg L-1, promoveu maior capacidade de regeneração e altura de brotos; o meio MS suplementado com BAP (2,5 mg L-1 estimulou maior altura de brotos e o meio MS suplementado com TDZ (1,0, 0,50 e 0,25 mg L-1 afetou a capacidade de regeneração de brotos, observando-se maior formação de calos.The micropropagation in vitro has been presented as a technique that allows various methodologies which, for in turn, contribute to a reduction in the time to obtain new cultivares. The objective of this work was to evaluate the behavior of the genotypes BRS-Verde, BRS-200-Marrom, 6M-Mocó-white and BRs(-187-8H- white, in the induction of the multiple shoots in different combinations of growth regulators. The seeds, to obtain the initial material, were placed in a solution of 1% sodium hypochlorite of active chlorine

RAPD analysis of mutants obtained by ion beam irradiation to hinoki cypress shoot primordia

International Nuclear Information System (INIS)

Ishii, K.; Yamada, Y.; Hase, Y.; Shikazono, N.; Tanaka, A.

2003-01-01

Mutants were induced by irradiation of the shoot primordia of Hinoki cypress with 50 MeV 4 He 2+ heavy ion beam. Fresh shoot primordia on the CD medium in the plastic Petri dish (35 x 10 mm) were irradiated. Xanta mutants were induced from 38 to 266 Gy irradiation. Waxy mutants were induced from 76 to 266 Gy irradiation. Xanta, waxy and control type of regenerated Hinoki cypress in vitro were checked for their DNA level difference using RAPD analysis. Among 81 primers used, 23 primers produced the 68 bands. Among them stable 44 bands produced by 15 primers were compared between mutants and control plant. So far, there is no variation among the RAPD analysis band patterns of those mutants. Bigger test size may detect the gene variation specific for mutants

Interactions of phytochromes A, B1 and B2 in light-induced competence for adventitious shoot formation in hypocotyl of tomato (Solanum lycopersicum L.).

Science.gov (United States)

Lercari, B; Bertram, L

2004-02-01

The interactions of phytochrome A (phyA), phytochrome B1 (phyB1) and phytochrome B2 (phyB2) in light-dependent shoot regeneration from the hypocotyl of tomato was analysed using all eight possible homozygous allelic combinations of the null mutants. The donor plants were pre-grown either in the dark or under red or far-red light for 8 days after sowing; thereafter hypocotyl segments (apical, middle and basal portions) were transferred onto hormone-free medium for culture under different light qualities. Etiolated apical segments cultured in vitro under white light showed a very high frequency of regeneration for all of the genotypes tested besides phyB1phyB2, phyAphyB1 and phyAphyB1phyB2 mutants. Evidence is provided of a specific interference of phyB2 with phyA-mediated HIR to far-red and blue light in etiolated explants. Pre-treatment of donor plants by growth under red light enhanced the competence of phyB1phyB2, phyAphyB1 and phyAphyB1phyB2 mutants for shoot regeneration, whereas pre-irradiation with far-red light enhanced the frequency of regeneration only in the phyAphyB1 mutant. Multiple phytochromes are involved in red light- and far-red light-dependent acquisition of competence for shoot regeneration. The position of the segments along the hypocotyl influenced the role of the various phytochromes and the interactions between them. The culture of competent hypocotyl segments under red, far-red or blue light reduced the frequency of explants forming shoots compared to those cultured under white light, with different genotypes having different response patterns.

Somatic embryo-like structures of strawberry regenerated in vitro on media supplemented with 2,4-D and BAP.

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Omar, Genesia F; Mohamed, Fouad H; Haensch, Klaus-Thomas; Sarg, Sawsan H; Morsey, Mohamed M

2013-09-01

Somatic embryo-like structures (SELS) were produced in vitro from leaf disk and petiole explants of two cultivars of strawberry (Fragaria x ananassa Duch) on Murashige and Skoog medium with different concentrations and combinations of 2,4-dichlorophenoxyacetic acid (2,4-D), 6-benzylaminopurine (BAP) and sucrose to check the embryonic nature of these structures histologically. A large number of SELS could be regenerated in both cultivars on media with 2-4 mg L(-1) 2,4-D in combination with 0.5 -1 mg L(-1) BAP and 50 g x L(-1) sucrose. Histological examination of SELS revealed the absence of a root pole. Therefore these structures cannot be strictly classified as somatic embryos. The SELS formed under the tested culture conditions represent malformed shoot-like and leaf-like structures. The importance of these results for the propagation of strawberries via somatic embryogenesis is discussed.

In vitro micropropagation in Polygonatum verticillatum (L.) All. an important threatened medicinal herb of Northern India.

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Bisht, Shivani; Bisht, N S; Bhandari, Snehlata

2012-01-01

An ideal micropropagation method of Polygonatum verticillatum has been developed using stem disc explants. Multiple shoots were initiated from stem disc explants on Murashige and Skoog (MS) medium fortified with different concentrations (0.25-10.0 mgl(-1)) and combinations of cytokinins (BAP, Kn and TDZ) along with (0.5-1.0 mgl(-1)) auxins (NAA/IBA/IAA). 1.0 mgl(-1) BAP with 0.5 mgl(-1) NAA was found to be the most effective in producing maximum number of shoots. Regular subculturing of these in vitro multiple shoots induced profuse growth of lateral roots in the same medium. Individual shoots were excised and rooted in vitro on half strength MS medium with 1.0 mgl(-1) NAA. Regenerants were hardened in growth chamber with high humidity and showed a high rate of survival.

Banana and coconut in-vitro cultures for induced mutations studies

International Nuclear Information System (INIS)

Guzman, E.V. de; Ubalde, E.M.; Rosario, A.G. del

1976-01-01

Explants obtained from shoot tips of developing suckers of banana were found to be suitable materials for plantlet formation in vitro. Subculture of the shoots formed from the original explant can lead to regeneration of numerous shoots from a single sucker. The technique can be a practical method of propagation. Though a more rapid rate of multiplication is still to be desired the present rate is much faster than the maximum rate of sucker production obtained under natural conditions. The conditions which favor shoot formation were the addition of benzyladenine, the use of dextrose at 2% instead of 4%, and the use of either Murashige and Skoog or of Vacin and Went salt solution. Viable explants can be obtained from suckers irradiated up to 10 Kr. Survival and shoot formation are sometimes not adversely affected by irradiation. Chlorosis and reduction in plantlet size and unusual seedling characters were among the features observed in plantlets regenerated from irradiated tissues. Proliferations derived from the cotyledonary sheath in coconut embryos could be successfully subcultured. The initial and subsequent proliferations may appear as nodular growths. In continuous subculture a nodular growth may change over to a large mass with smooth surface. Cellular characteristics of the two types of growth are markedly different. (author)

Plants regeneration of a papaya hybrid (IBP 42-99 from callus obtained from apexes of in vitro plants.

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Jorge Gallardo Colina

2004-07-01

Full Text Available In Cuba there realizes innumerable efforts to increase the food production and especially the fruit trees Inside which the papaya has great importance. In this sense studies are realized to obtain plants resistant to virus that they need of tools that they support and increase the indexes of obtaining transgenic line in the events of transformation specifically in a papaya hybrid. As main objective was to develop a protocol for the regeneration of plants of papaya hybrid from callus obtained of in vitro plant apexes.plan to develop a tool that supports and increases the indexes of obtaining line transgenic in the events of transformation in a hybrid of papaya.In vitro plants of the hybrid IBP 42-99 were used as plant material. The culture medium Nitsh and Nitsh was used and the growth regulators that permitted the obtaining of the best callus with embryogenic structures were studied, and also, the concentrations in which they were more efficient were adjusted. The capacity of callus formation from different parts of the stem of the in vitro plants was studied. Different culture medium for the regeneration of papaya plants from the obtained callus was studied. It was possible to obtain callus by combining 6-BAP with ANA and AIA. The best results are obtained when segments from in vitro plants shoot, 1 cm length from the meristem, were used. Also, by eliminating the meristem in the apexes, an increase in the callus formation capacity of the explant was achieved. Plants were obtained from callus using the culture medium MS supplement with 0.09mg.l-1 of AIA, 0.01mg.l-1 of AG3 and 2mg.l-1 of Zeatin and the best percentage was achieved with the callus coming from the treatments with less concentration of 6-BAP and AIA. Key words: culture medium, micropropagation, organogenesis, segments

Effects of fungicides and bactericides on orchid seed germination and shoot tip cultures in vitro

OpenAIRE

Brown, DM; Groom, CL; Cvitanik, M; Brown, M; Cooper, JL; Arditti, J

1981-01-01

Amphotericin B, benomyl, gentamycin, nystatin, quintozene penicillin G, sodium omadine, and vancomycin singly and in several combinations have no deleterious effects on the germination of orchid seeds, but inhibit the growth in vitro of shoot tip explants. © 1981 Martinus Nijhoff/Dr W. Junk Publishers.

Development of In Vitro Systems for Switchgrass (Panicum virgatum) - Final Report for 1992 to 2002

Energy Technology Data Exchange (ETDEWEB)

Conger, B.V.

2003-01-16

Our project began on July 1, 1992, with the objective of developing systems that could be used in biotechnological approaches to switchgrass improvement. Within six months after initiation of the project, we had worked out protocols in which plants could be regenerated from callus cultures through both organogenesis and somatic embryogenesis. Documentation for both modes of regeneration was provided in our progress reports and in publications. One thousand regenerated plants were established in the field during the first year. We found that Alamo (lowland type) was much more amenable to in vitro culture, and plants could be regenerated much more easily than from Cave-in-Rock (upland type). During the first three years of the project, we studied the influence of genotype, culture medium components, explant type, etc., on regeneration. As mentioned, we found that the lowland cultivars Alamo and Kanlow were much easier to regenerate than upland cultivars, such as Trailblazer, Blackwell, and Cave-in-Rock. For callus induction, we initially used mature caryopses, young leaf tissue, and portions of seedlings. We were successful in inducing callus and regenerating plants from all explants. Two other systems developed during the 4th to 6th year period of the project included multiple shoot formation initiated from germinated seedlings and regenerable suspension cultures. The latter were initiated from embryogenic calluses produced from in vitro developed inflorescences. An important factor for producing multiple shoots was the presence of thidiazuron in the medium. The shoots could be easily rooted and numerous plantlets produced. The last 3 to 4 years of the project focused on anther and microspore culture experiments to produce haploid plants and on genetic transformation. Although thousands of putative haploid plants were produced from a few anthers, they were very weak and difficult to keep alive. Chromosome counts revealed the gametic number in cells where it was

Optimization of Regeneration Conditions and In Vitro Propagation of Sideritis Stricta Boiss & Heldr.

Science.gov (United States)

Yavuz, Dudu Özkum

2016-09-01

In this study the micropropagation of endemic species Sideritis stricta was investigated. Leaf segments and shoot explants (hypocotyl, single node and shoot tips) taken from in vitro growing plantlets and cultured on MS and B5 media containing different growth regulators combinations BAP (0.0, 1.0, 2.0 and 3.0mg/l) and NAA (0.0, 0.1 and 0.5mg/l). MS and B5 media supplemented with BAP (1.0, 2.0 and 3.0mg/l) and NAA (0.1mg/l) combinations or only BAP and kinetin (2.0 and 3.0mg/l) were used at the subculture experiments of shoots and MS and B5 media supplemented with different concentrations of IBA (0.0, 1.5, 3.0, 4.5 and 10.mg/l) were used at the rooting experiments. S. stricta seeds germinated at the rate of 100% when the seed coat was removed and endoperm with embryo part cultured on B5 medium. The single node explants taken from in vitro germinated and grown 30-40 days plantlets on B5 medium have been determined as the most successful explant at all used hormone combinations. B5 medium supplemented with 1.0mg/l BAP+0.1mg/l NAA and 2.0mg/l BAP+0.5mg/l NAA was determined as the most effective medium on shoot formation. At the first and second subculture, the highest shoot formation was maintained on medium supplemented with 1.0mg/l BAP+0.1mg/l NAA and the number of shoots per explant were 4 and 2.11, respectively. The highest multiplication rate has been determined as 33.76 at the end of second subculture. The best rooting was achieved on B5 medium supplemented with 4.5mg/l IBA. The rooted shoots were successfully acclimatized to outdoor conditions and survival rate was determined as 90%. Copyright © 2015 Elsevier B.V. All rights reserved.

Stable transformation via particle bombardment in two different soybean regeneration systems.

Science.gov (United States)

Sato, S; Newell, C; Kolacz, K; Tredo, L; Finer, J; Hinchee, M

1993-05-01

The Biolistics(®) particle delivery system for the transformation of soybean (Glycine max L. Merr.) was evaluated in two different regeneration systems. The first system was multiple shoot proliferation from shoot tips obtained from immature zygotic embryos of the cultivar Williams 82, and the second was somatic embryogenesis from a long term proliferative suspension culture of the cultivar Fayette. Bombardment of shoot tips with tungsten particles, coated with precipitated DNA containing the gene for β-glucuronidase (GUS), produced GUS-positive sectors in 30% of the regenerated shoots. However, none of the regenerants which developed into plants continued to produce GUS positive tissue. Bombardment of embryogenic suspension cultures produced GUS positive globular somatic embryos which proliferated into GUS positive somatic embryos and plants. An average of 4 independent transgenic lines were generated per bombarded flask of an embryogenic suspension. Particle bombardment delivered particles into the first two cell layers of either shoot tips or somatic embryos. Histological analysis indicated that shoot organogenesis appeared to involve more than the first two superficial cell layers of a shoot tip, while somatic embryo proliferation occurred from the first cell layer of existing somatic embryos. The different transformation results obtained with these two systems appeared to be directly related to differences in the cell types which were responsible for regeneration and their accessibility to particle penetration.

Regeneration of three sweet potato (Ipomea batatas (L.)) accessions via meristem, Nodal and callus induction

International Nuclear Information System (INIS)

Addae-Frimpomaah, F.

2012-11-01

In vitro regeneration of three sweet potato accessions UE007, UK-BNARI and SA-BNARI using meristem, nodal cuttings or callus induction was studied. Meristematic explants cultured on Murashige and Skoog (1962) basal medium supplemented with low concentration of benzylaminopurine (BAP) or kinetin resulted in callus with or without shoot development which delayed shoot emergence. The degree of callus development increased as the concentration of the cytokinin in the culture medium increased. Although, callus development was comparatively lower on kinetin amended medium than BAP amended medium, Murashige and Skoog medium supplemented with 0.25mg/1BAP had the highest shoot induction (80%). For further differentiation of callus or shoots into distinct stem and leaves, the culture were transferred into fresh MS medium supplemented with 0.25mg/1 BAP, 0.1 mg/1 NAA and 0.1 mg/1 Gibberellic acid (GA 3 . To overcome the delay in shoot initiation using meristem culture, nodal cuttings of sweet potato were used as explants and cultured on MS medium amended with 0.3 - 0.9mg/1 BAP. All explants cultured on 0.3 or 0.6mg/1 BAP developed shoots. Furthermore, liquid MS medium amended with 0.25mg/1 BAP, 0.1mg/I NAA, and 0.1mg/1 GA 3 also enhanced early shoot development from nodal cutting explants compared to solid culture. Post flask acclimatisation of meristem or nodal cutting-derived plantlets showed that meristem derived plantlets were better acclimatised than nodal cutting plants due to vigorous root development leading to higher percentage survival in pots and subsequent tuber production. Callusogenesis was achieved when leaf lobe explants were cultured on CLC/ Ipomoea medium supplemented with 1.0 - 4.0mg/1 2,4-D with 4.0mg/1 2,4-D being the optimal concentration. However, the calli were non-embryogenic and therefore could not produce embryos when transferred to 0.1mg/1 BAP amended medium but rather produced either single or multiple shoots. The highest percentage shoot (83

Optimization of soybean (glycine max L.) regeneration for korean cultivars

International Nuclear Information System (INIS)

Phat, P.; Rehman, S. U.; Ju, H. J.; Jung, H. I.

2015-01-01

Tissue culture could provide key insights into the development of transgenic plants, production of good cultivars and secondary metabolites, conservation of endangered plants, and safeguarding of germplasms. In this study, the effects of shoot induction media, explants, cultivars, and phytohormone concentrations on the regeneration efficiency of Korean soybean cultivars were evaluated. Restricted dormancy and poor germination may affect regeneration, depending on the type of germination medium or initiation of phytohormone treatment. Therefore, we analyzed the effects of different germination media containing plant growth regulators, i.e. 6-benzyladenine (BAP), gibberellic acid 3 (GA /sub 3/), and naphthalene acetic acid (NAA), prior to investigating the influences of explant types, media with or without vitamins, cultivars, and different phytohormones (BAP and GA3). A high frequency of germination was observed in Murashige and Skooge (MS) medium with vitamins supplemented with 1 mg L /sup -1/ BAP and 0.25 mg L /sup -1/ GA /sub 3/. Cotyledonary node explants and Gamborg B5 with vitamins supplemented with 1 mg L /sup -1/ BAP and 0.17 mg L /sup -1/ GA /sub 3/ in callus induction medium (CIM) and 1 mg L /sup -1/ BAP in shoot induction medium (SIM) were found to be the most efficient conditions for induction of soybean regeneration, both in callus development and shoot regeneration. Two Korean soybean cultivars, cv. Daepung and Nampung, showed similar development of shoot regeneration efficiency, but significantly different shoot induction times. Therefore, the protocol reported here may be used for further development of regeneration efficiency and can be employed for efficient transformation in soybeans. (author)

In Vitro Mimetic Models for the Bone-Cartilage Interface Regeneration.

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Bicho, Diana; Pina, Sandra; Oliveira, J Miguel; Reis, Rui L

2018-01-01

In embryonic development, pure cartilage structures are in the basis of bone-cartilage interfaces. Despite this fact, the mature bone and cartilage structures can vary greatly in composition and function. Nevertheless, they collaborate in the osteochondral region to create a smooth transition zone that supports the movements and forces resulting from the daily activities. In this sense, all the hierarchical organization is involved in the maintenance and reestablishment of the equilibrium in case of damage. Therefore, this interface has attracted a great deal of interest in order to understand the mechanisms of regeneration or disease progression in osteoarthritis. With that purpose, in vitro tissue models (either static or dynamic) have been studied. Static in vitro tissue models include monocultures, co-cultures, 3D cultures, and ex vivo cultures, mostly cultivated in flat surfaces, while dynamic models involve the use of bioreactors and microfluidic systems. The latter have emerged as alternatives to study the cellular interactions in a more authentic manner over some disadvantages of the static models. The current alternatives of in vitro mimetic models for bone-cartilage interface regeneration are overviewed and discussed herein.

Efficient in vitro regeneration protocol of Centella asiatica (L.) Urban ...

African Journals Online (AJOL)

The present communication reports an efficient in vitro plantlet regeneration protocol for endemic umbellifer Centella asiatica (L.) urban via callus mediated organogenesis from leaf and stem explants. The plant is pharmacologically very important and its consumption as underutilized green leafy vegetable affluent in ...

Production of mutants by irradiation of in vitro-cultured tissues of coconut and banana and their mass propagation by the tissue culture technique

International Nuclear Information System (INIS)

Guzman, E.V. de; Rosario, A.G. del; Pagcaliwagan, P.C.

1982-01-01

Regeneration of buds/shoots as well as plantlets was induced from banana shoot tip explants cultured in highly modified Murashige and Skoog's medium supplemented with coconut water and benzyladenine. Initially shoot regeneration was sparse, but on further subculture became profuse. Gamma irradiation at low dosage (1.0 kR) was stimulating to explant growth and bud formation with the two types of explants used. With Bungulan stimulation was observed even at 2.5 kR. Several morphological aberrations were exhibited by shoots of 'irradiated' in vitro plants growing in potted soil. A highly and continuously proliferating tissue strain has been isolated from a subculture which was ultimately derived from an irradiated explant. Its continued proliferation is dependent on an external supply of coconut water and benzyladenine. In vitro-produced plants have been established under field conditions. The 'irradiated' plants are comparable with, and some seem to be better than, the unirradiated controls with respect to height, girth, sucker production and number of hands and fingers per bunch. Higher doses of irradiation are required to produce an adverse effect on growth of coconut embryos during the liquid culture than when growing in solid medium. (author)

An evaluation of a new approach to the regeneration of Helichrysum italicum (Roth) G. Don, and the molecular characterization of the variation among sets of differently derived regenerants.

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Perrini, Rosaria; Alba, Vittorio; Ruta, Claudia; Morone-Fortunato, Irene; Blanco, Antonio; Montemurro, Cinzia

2009-01-01

A protocol for the induction of regeneration from leaves of Helichrysum italicum was established. Calli were found to form on the basal medium only when it was supplemented with thidiazuron (TDZ) alone or in combination with naphthalene acetic acid (NAA), with a percentage ranking of at least 80%. The hormone-free medium showed the highest percentage of shoot regeneration (62%) even though no callus formed. AFLP markers were employed to verify tissue culture-induced variation in the regenerated plantlets obtained by direct shoot regeneration or the indirect shoot regeneration process (callus formation). Seven out of the eleven AFLP primer pairs yielded polymorphic patterns. The average number of fragments per primer pair was 64.1. Singletons were represented by 12 (2.7%) fragments. Student's T-test was performed both on the average number of shared fragments and on the nucleotide diversity, and no significant statistical difference was observed between the two regeneration treatments.

In vitro propagation of peanut (Arachis hypogaea L.) by shoot tip culture.

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Ozudogru, Elif Aylin; Kaya, Ergun; Lambardi, Maurizio

2013-01-01

Peanut (Arachis hypogaea L.), also known as groundnut, is the most important species of Arachis genus, originating from Brazil and Peru. Peanut seeds contain high seed oil, proteins, amino acids, and vitamin E, and are consumed worldwide as edible nut, peanut butter, or candy, and peanut oil extracted from the seeds. The meal remaining after oil extraction is also used for animal feed. However, its narrow germplasm base, together with susceptibility to diseases, pathogens, and weeds, decreases yield and seed quality and causes great economic losses annually. Hence, the optimization of efficient in vitro propagation procedures would be highly effective for peanut propagation, as it would raise yield and improve seed quality and flavor. Earlier reports on traditional micropropagation methods, based on axillary bud proliferation which guarantees the multiplication of true-to-type plants, are still limited. This chapter describes a micropropagation protocol to improve multiple shoot formation from shoot-tip explants by using AgNO(3) in combination with plant growth regulators.

An Improved Micropropagation Protocol by Ex Vitro Rooting of Passiflora edulis Sims. f. flavicarpa Deg. through Nodal Segment Culture.

Science.gov (United States)

Shekhawat, Mahipal S; Manokari, M; Ravindran, C P

2015-01-01

A procedure for rapid clonal propagation of Passiflora edulis Sims. f. flavicarpa Deg. (Passifloraceae) has been developed in this study. Nodal explants were sterilized with 0.1% HgCl2 and inoculated on Murashige and Skoog (MS) basal medium. The addition of 2.0 mgL(-1) 6-benzylaminopurine (BAP) to MS medium caused an extensive proliferation of multiple shoots (8.21 ± 1.13) primordial from the nodal meristems. Subculturing of these multiple shoots on the MS medium augmented with 1.0 mgL(-1) of each BAP and Kinetin (Kin) was successful for the multiplication of the shoots in vitro with maximum numbers of shoots (25.73 ± 0.06) within four weeks of incubation. Shoots were rooted best (7.13 ± 0.56 roots/shoots) on half strength MS medium supplemented with 2.0 mgL(-1) indole-3 butyric acid (IBA). All in vitro regenerated shoots were rooted by ex vitro method, and this has achieved 6-7 roots per shoot by pulsing of cut ends of the shoots using 200 as well as 300 mgL(-1) IBA. The plantlets were hardened in the greenhouse for 4-5 weeks. The hardened plantlets were shifted to manure containing nursery polybags after five weeks and then transferred to a sand bed for another four weeks for acclimatization before field planting with 88% survival rate.

High taxonomic diversity of cultivation-recalcitrant endophytic bacteria in grapevine field shoots, their in vitro introduction, and unsuspected persistence.

Science.gov (United States)

Thomas, Pious; Sekhar, Aparna C; Shaik, Sadiq Pasha

2017-11-01

Molecular and microscopic analyses reveal enormous non-cultivable endophytic bacteria in grapevine field shoots with functional significance. Diverse bacteria enter tissue cultures through surface-sterilized tissues and survive surreptitiously with varying taxonomic realignments. The study was envisaged to assess the extent of endophytic bacterial association with field shoot tissues of grapevine and the likelihood of introduction of such internally colonizing bacteria in vitro adopting molecular techniques targeting the non-cultivable bacterial community. PowerFood ® -kit derived DNA from surface-sterilized field shoot tips of grapevine Flame Seedless was employed in a preliminary bacterial class-specific PCR screening proving positive for major prokaryotic taxa including Archaea. Taxonomic and functional diversity were analyzed through whole metagenome profiling (WMG) which revealed predominantly phylum Actinobacteria, Proteobacteria, and minor shares of Firmicutes, Bacteroidetes, and Deinococcus-Thermus with varying functional roles ascribable to the whole bacterial community. Field shoot tip tissues and callus derived from stem segments were further employed in 16S rRNA V3-V4 amplicon taxonomic profiling. This revealed elevated taxonomic diversity in field shoots over WMG, predominantly Proteobacteria succeeded by Actinobacteria, Firmicutes, Bacteroidetes, and 15 other phyla including several candidate phyla (135 families, 179 genera). Callus stocks also displayed broad bacterial diversity (16 phyla; 96 families; 141 genera) bearing resemblance to field tissues with Proteobacterial dominance but a reduction in its share, enrichment of Actinobacteria and Firmicutes, disappearance of some field-associated phyla and detection of a few additional taxonomic groups over field community. Similar results were documented during 16S V3-V4 amplicon taxonomic profiling on Thompson Seedless field shoot tip and callus tissues. Video microscopy on tissue homogenates

In vitro propagation and whole plant regeneration from callus in ...

African Journals Online (AJOL)

user

2011-01-17

Jan 17, 2011 ... Leaf explants and mature embryos of Datura were separately cultured in MS basal medium ... al., 2001). It was found that in vitro culture can enable plants to produce secondary metabolites under controlled culture conditions. Furthermore, the establishment of ..... formation and plant regeneration in tomato.

Shoot differentiation from protocorm callus cultures of Vanilla planifolia (Orchidaceae): proteomic and metabolic responses at early stage.

Science.gov (United States)

Palama, Tony L; Menard, Patrice; Fock, Isabelle; Choi, Young H; Bourdon, Emmanuel; Govinden-Soulange, Joyce; Bahut, Muriel; Payet, Bertrand; Verpoorte, Robert; Kodja, Hippolyte

2010-05-05

Vanilla planifolia is an important Orchid commercially cultivated for the production of natural vanilla flavour. Vanilla plants are conventionally propagated by stem cuttings and thus causing injury to the mother plants. Regeneration and in vitro mass multiplication are proposed as an alternative to minimize damage to mother plants. Because mass production of V. planifolia through indirect shoot differentiation from callus culture is rare and may be a successful use of in vitro techniques for producing somaclonal variants, we have established a novel protocol for the regeneration of vanilla plants and investigated the initial biochemical and molecular mechanisms that trigger shoot organogenesis from embryogenic/organogenic callus. For embryogenic callus induction, seeds obtained from 7-month-old green pods of V. planifolia were inoculated on MS basal medium (BM) containing TDZ (0.5 mg l(-1)). Germination of unorganized mass callus such as protocorm -like structure (PLS) arising from each seed has been observed. The primary embryogenic calli have been formed after transferring on BM containing IAA (0.5 mg l(-1)) and TDZ (0.5 mg l(-1)). These calli were maintained by subculturing on BM containing IAA (0.5 mg l(-1)) and TDZ (0.3 mg l(-1)) during 6 months and formed embryogenic/organogenic calli. Histological analysis showed that shoot organogenesis was induced between 15 and 20 days after embryogenic/organogenic calli were transferred onto MS basal medium with NAA (0.5 mg l(-1)). By associating proteomics and metabolomics analyses, the biochemical and molecular markers responsible for shoot induction have been studied in 15-day-old calli at the stage where no differentiating part was visible on organogenic calli. Two-dimensional electrophoresis followed by matrix-assisted laser desorption ionization time-of-flight-tandem mass spectrometry (MALDI-TOF-TOF-MS) analysis revealed that 15 protein spots are significantly expressed (P tissue culture, phenolic compounds such

An efficient and reproducible method for in vitro clonal multiplication of Rauvolfia tetraphylla L. and evaluation of genetic stability using DNA-based markers.

Science.gov (United States)

Faisal, Mohammad; Alatar, Abdulrahman A; Ahmad, Naseem; Anis, Mohammad; Hegazy, Ahmad K

2012-12-01

An efficient protocol is described for the rapid in vitro clonal propagation of an endangered medicinal plant, Rauvolfia tetraphylla L., through high frequency shoot induction from nodal explants collected from young shoots of a field grown plant. Effects of growth regulators [6-benzyladenine (BA), kinetin (Kin) 2iP, or α-naphthalene acetic acid (NAA)], carbohydrates, different medium [Murashige and Skoog (MS), Woody Plant Medium (WPM), Gamborg medium (B5), Linsmier and Skoog medium (LS)], and various pH levels on in vitro morphogenesis were investigated. The highest frequency of shoot regeneration (90 %) and maximum number of shoot (35.4 ± 2.3) per explant were observed on WPM medium supplemented with 7.5 μM BA, 2.5 μM NAA, and 30 g/l sucrose at pH 5.8. Well-developed shoots, 4-5 cm in length, were successfully rooted ex vitro at 90 % by a 30-min pulse treatment with 150 μM IBA prior to their transfer in planting substrates. The survival rate of transplantation reached 90 % when transferred to field condition. Genetic stability of micropropagated plantlets was assessed and compared with mother plant using Random Amplified Polymorphic DNA and Inter Simple Sequence Repeats markers. No variation was observed in DNA fingerprinting patterns among the micropropagated plants, which were similar to that of the donor plant illustrating their genetic uniformity and clonal fidelity. This confirms that clonal propagation of this plant using axillary shoot buds can be used for commercial exploitation of the selected genotype where a high degree of fidelity is an essential prerequisite. The work contributed to a better in vitro regeneration and clonal mass multiplication of R. tetraphylla and to develop a strategy for the germplasm conservation of this endangered medicinal plant.

Micropropagation and validation of genetic and biochemical fidelity amongst regenerants of Cassia angustifolia Vahl employing RAPD marker and HPLC.

Science.gov (United States)

Chetri, Siva K; Sardar, Pratima Rani; Agrawal, Veena

2014-10-01

In vitro protocol has been established for clonal propagation of Cassia angustifolia Vahl which is an important source of anticancerous bioactive compounds, sennoside A and B. Nodal explants excised from field raised elite plant (showing optimum level of sennoside A and B) of C. angustifolia when reared on Murashige and Skoog's medium augmented with different cytokinins, viz. N(6)-benzyladenine (BA), N(6)-(2-isopentenyl) adenine (2iP) and 6-furfuryl aminopurine (Kn) differentiated multiple shoots in their axils. Of the three cytokinins, BA at 5 μM proved optimum for differentiating multiple shoots in 95 % cultures with an average of 9.14 shoots per explant within 8 weeks of culture. Nearly, 95 % of the excised in vitro shoots rooted on half strength MS medium supplemented with 10 μM indole-3-butyric acid (IBA). The phenotypically similar micropropagated plants were evaluated for their genetic fidelity employing random amplified polymorphic DNA (RAPD) markers. Eleven individuals, randomly chosen amongst a population of 120 regenerants were compared with the donor plant. A total of 36 scorable bands, ranging in size from 100 to 1,000 bp were generated amongst them by the RAPD primers. All banding profiles from micropropagated plants were monomorphic and similar to those of mother plant proving their true to the type nature. Besides, high performance liquid chromatography evaluation of the sennoside A and B content amongst leaves of the mature regenerants and the elite mother plant too revealed consistency in their content.

Human skeletal muscle fibroblasts stimulate in vitro myogenesis and in vivo muscle regeneration

DEFF Research Database (Denmark)

Mackey, Abigail L; Magnan, Mélanie; Chazaud, Bénédicte

2017-01-01

immediately surrounding regenerating muscle fibres. These novel findings indicate an important role for fibroblasts in supporting the regeneration of muscle fibres, potentially through direct stimulation of satellite cell differentiation and fusion, and contribute to understanding of cell-cell cross......-talk during physiological and pathological muscle remodelling. ABSTRACT: Accumulation of skeletal muscle extracellular matrix is an unfavourable characteristic of many muscle diseases, muscle injury and sarcopenia. In addition to the indispensable role satellite cells play in muscle regeneration......, there is emerging evidence in rodents for a regulatory influence on fibroblast activity. However, the influence of fibroblasts on satellite cells and muscle regeneration in humans is unknown. The purpose of this study was to investigate this in vitro and during in vivo regeneration in humans. Following a muscle...

Organogenesis from in vitro-derived leaf and internode explants of Hoya wightii ssp. palniensis -a vulnerable species of Western Ghats

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Subbaiah Revathi Lakshmi

2013-06-01

Full Text Available An efficient system was developed for indirect plant regeneration from in vitro-derived leaf and internode explants of Hoya wightii ssp. palniensis. Maximum percentage of the organogenic callus was obtained on MS medium supplemented with NAA (1.0 mg/l and 2,4-D (2.0 mg/l. The best shoot bud induction was observed on MS medium with BA (1.0 mg/l +IBA (0.5 mg/l. The coconut water (15% was better, resulting in a differentiation of the shoot initials in to well-developed shoots. The elongated shoots (› 3cm long were rooted on a full strength MS basal medium, supplemented with 0.2 mg/l of IBA. Finally, the rooted plants were transferred to the soil with 80% success rate. This protocol was utilized for the in vitro propagation of this endangered plant species.

A valued Indian medicinal plant – Begonia malabarica Lam. : Successful plant regeneration through various explants and field performance

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Sevanan Rajeshkumar

2009-05-01

Full Text Available A cost-effective and efficient protocol has been described in the present work for large-scale and rapid in vitro propagation of a valuable medicinal herb Begonia malabarica Lam. (Begoniaceae by shoot auxillary-bud proliferation and organogenesis on MS medium supplemented with 6-benzylaminopurine (BA; 0.0-8.8 mg/l and indole-3-acetic acid (IAA; 0.0-2.88 mg/l at different concentrations, either alone or in combinations. Initiation of callus formation from the base of the leaf lamina was observed on MS supplemented with BA, IAA and adenine sulphate. Root induction on shoots was achieved on full strength MS with IAA/ indole-3-butyric acid (IBA at different concentrations. MS medium with 4.4 mg/l BA and 1.4 mg/l IAA elicited the maximum number of shoots (10 multiple shoots from nodal explants. Leaf-based callus differentiated into more than 28 shoots on MS with 150 mg/l adenine sulphate. The regenerated shoots were rooted on MS with 1.2 mg/l IBA within ten days. Almost 95% of the rooted shoots survived hardening when transferred to the field. The regenerated plants did not show any morphological change and variation in levels of secondary metabolites when compared with the mother stock. Thus, a reproduction of B. malabarica was established through nodal and leaf explants. This protocol can be exploited for conservation and commercial propagation of this medical plant in the Indian subcontinent and might be useful for genetic improvement programs.

In Vitro Proliferation and Cryoconservation of Banana and Plantain Elite Clones

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Reyes Guillermo

2017-12-01

Full Text Available Agriculture and modern biotechnology are increasingly becoming interdependent, and many new techniques have brought new opportunities for enhancing production and marketing. Germplasm storage is an alternative for the conservation of plant genetic diversity, contributing to the improvement and maintenance of propagation programs for species of interest. In this work, banana corms were collected as plant material from relatively young commercial plantations of three different cultivars: ‘Williams’, Valery (AAA genome; Cavendish subgroup, and ‘Barraganete’ (AAB genome; Plantain subgroup. Their shoot tips were introduced into in vitro conditions, and subcultured monthly to obtain the required number of shoots. The shoots were subsequently rooted and stimulated to invigoration in order to extract apical meristems (0.8–1.0 mm, which were prepared for cryopreservion in liquid nitrogen (−196 °C following pre-conditioning in PVS2 vitrification solution. Thereafter, the explants were rapidly thawed and then recovered and regenerated using two different methods – by Panis (2009 and Korneva et al. (2009 – consisting of two different sets of recovery and subsequent regeneration media. Statistical analysis of the results showed that the banana cultivar ‘Williams’ demonstrated higher survival and regeneration rates after cry-opreservation using the Korneva method, whereas in cultivars ‘Valery’ and ‘Barraganete’, there were no significant differences between the tested methods. The ‘Barraganete’ cultivar had the lowest survival and regeneration rates, regardless of the applied method.

In vitro propagation and withaferin A production in Withania ashwagandha, a rare medicinal plant of India.

Science.gov (United States)

Mir, Bilal Ahmad; Mir, Shabir Ahmad; Koul, Sushma

2014-07-01

Withania ashwagandha, belonging to the family Solanaceae, is an important medicinal herb of India with restricted geographic distribution. It is a rich source of withaferin A (WA) and other bioactive withanolides. In the present study a rapid in vitro mass propagation protocol of W. ashwagandha was developed from nodal explants. Nodal explants were cultured on MS medium supplemented with various concentrations and combinations of plant growth regulators (PGRs). The highest number of regenerated shoots per ex-plant (33 ± 2.7) and highest WA (13.4 ± 1.15 mg/g of DW) production was obtained on MS medium supplemented with 5.0 μM 6-benzyladenine (BA) and 1.0 μM Kinetin (Kn). In vitro raised shoots were further rooted on half-strength MS medium containing 2.0 μM Indole-3-butyric acid (IBA) and analyzed for WA production. The rooted plantlets when transferred to poly bags in the greenhouse showed 90 % survival frequency. Levels of WA were higher in the in vitro and ex vitro derived shoot and root tissues as compared to field grown mother plants. In an attempt to further maximize WA production, shoot cultures were further grown in liquid MS medium supplemented with 5.0 μM 6-benzyladenine (BA) and 1.0 μM Kinetin (Kn). Root cultures were grown on half strength MS liquid medium fortified with 2.0 μM of IBA. WA production in the liquid cultures was significantly higher compared to the static composition of the same media. This protocol, first of its kind in this plant, can be successfully employed for conservation, proliferation and large-scale production of WA. The regenerated plants can also be used in traditional medicine as an alternative to naturally collected plants.

Improvement of efficient in vitro regeneration potential of mature callus induced from Malaysian upland rice seed (Oryza sativa cv. Panderas).

Science.gov (United States)

Mohd Din, Abd Rahman Jabir; Iliyas Ahmad, Fauziah; Wagiran, Alina; Abd Samad, Azman; Rahmat, Zaidah; Sarmidi, Mohamad Roji

2016-01-01

A new and rapid protocol for optimum callus production and complete plant regeneration has been assessed in Malaysian upland rice (Oryza sativa) cv. Panderas. The effect of plant growth regulator (PGR) on the regeneration frequency of Malaysian upland rice (cv. Panderas) was investigated. Mature seeds were used as a starting material for callus induction experiment using various concentrations of 2,4-D and NAA. Optimal callus induction frequency at 90% was obtained on MS media containing 2,4-D (3 mg L(-1)) and NAA (2 mg L(-1)) after 6 weeks while no significant difference was seen on tryptophan and glutamine parameters. Embryogenic callus was recorded as compact, globular and light yellowish in color. The embryogenic callus morphology was further confirmed with scanning electron microscopy (SEM) analysis. For regeneration, induced calli were treated with various concentrations of Kin (0.5-1.5 mg L(-1)), BAP, NAA and 0.5 mg L(-1) of TDZ. The result showed that the maximum regeneration frequency (100%) was achieved on MS medium containing BAP (0.5 mg L(-1)), Kin (1.5 mg L(-1)), NAA (0.5 mg L(-1)) and TDZ (0.5 mg L(-1)) within four weeks. Developed shoots were successfully rooted on half strength MS free hormone medium and later transferred into a pot containing soil for acclimatization. This cutting-edge finding is unique over the other existing publishable data due to the good regeneration response by producing a large number of shoots.

Effect of nickel on regeneration in Jatropha curcas L. and assessment of genotoxicity using RAPD markers

KAUST Repository

Sarkar, Tanmoy

2010-07-08

The aim of the present study was to determine the effect of nickel on shoot regeneration in tissue culture as well as to identify polymorphisms induced in leaf explants exposed to nickel through random amplified polymorphic DNA (RAPD). In vitro leaf explants of Jatropha curcas were grown in nickel amended Murashige and Skoog (MS) medium at four different concentrations (0, 0.01, 0.1, 1 mM) for 3 weeks. Percent regeneration, number of shoots produced and genotoxic effects were evaluated by RAPD using leaf explants obtained from the first three treatments following 5 weeks of their subsequent subculture in metal free MS medium. Percent regeneration decreased with increase in addition of nickel to the medium up to 14 days from 42.31% in control to zero in 1.0 mM. The number of shoot buds scored after 5 weeks was higher in control as compared to all other treatments except in one of the metal free subculture medium wherein the shoot number was higher in 0.01 mM treatment (mean = 7.80) than control (mean = 7.60). RAPD analysis produced only 5 polymorphic bands (3.225%) out of a total of 155 bands from 18 selected primers. Only three primers OPK-19, OPP-2, OPN-08 produced polymorphic bands. The dendrogram showed three groups A, B, and C. Group A samples showed 100% genetic similarity within them. Samples between groups B and C were more genetically distant from each other as compared to samples between groups A and B as well as groups A and C. Cluster analysis based on RAPD data correlated with treatments. © 2010 Springer Science+Business Media, LLC.

An efficient protocol for regeneration and transformation of Symphyotrichum novi-belgii

DEFF Research Database (Denmark)

Mørk, Eline Kirk; Henriksen, Karin; Brinch-Pedersen, Henrik

2012-01-01

) fusion gene, and an enhanced green fluorescent protein gene. Pre-cultured hypocotyl explants were transformed in the presence of 100 μM acetosyringone using 90 s sonication plus 10 min vacuum-infiltration. Kanamycin at 20 mg l−1 was used for selecting transformed cells. Adventitious shoots regenerated......II genes in GUS-positive shoots were confirmed by PCR and copy number of the nptII gene in PCR-positive shoots was determined by Southern blotting. Three transgenic plantlets were acclimatized to the greenhouse. This transformation and regeneration system using hypocotyls provides a foundation...

Development of Sirococcus shoot blight following thinning in western hemlock regeneration.

Science.gov (United States)

Charles G. Shaw; Thomas H. Laurent; Spencer. Israelson

1981-01-01

Shoot mortality from Sirococcus strobilinus Preuss. and other causes was recorded by crown position from April 1978 through October 1979 in younggrowth western hemlock (Tsuga heterophylla (Raf.) Sarg.) crop trees released in a 1977 thinning at Thomas Bay, Alaska. All study trees contained some infected shoots, but no terminal...

In vitro propagation of the Garden Heliotrope, Valeriana officinalis L.: influence of pre-chilling and light on seed germination.

Science.gov (United States)

Bhat, B; Sharma, V D

2015-03-01

Valeriana officinalis is an important medicinal herb commonly found in Kashmir valley. This study forms an important preliminary step for in-vitro micro propagation of V. officinalis from breaking the seed dormancy, inducing rapid seed germination and its subsequent micro propagation. We investigated the influence of pretreatment of V. officinalis seeds with reduced temperature and light on seed germination and in-vitro propagation. Culture of explants from cultivated seeds have demonstrated its potential for in vitro propagation and plantlet regeneration. Individual as well as combinations of treatments such as temperature and light availability influenced the germination of seeds variedly. Unchilled seeds of V. officinalis were given dip in GA3 (200 ppm) for 24, 48 and 120 h. Seeds treated with GA3 for 24 h and kept in darkness showed the best results, i.e. 48%. Seeds pretreated with GA3 for 120 h and incubated in dark showed 40% germination. Pre-chilling up to 72 h and kept in light showed maximum germination of 60% followed by 40% kept in darkness. Pre-chilling for 48 h resulted in 40 and 25% seed germination in light and darkness, respectively. GA3 pre-treatment for 72 h and 24 h pre chilling were most effective in inducing seed germination. Maximum shoot response was obtained on MS enriched with BAP (1 mg/L) + IAA (0.1 mg/L) combinations using shoot tips as explants. Multiple shoot regeneration from shoot apices was recorded on BAP (1 mg/L) and BAP (1 mg/L) + IAA (0.1 mg/L).

Improvement of Seed Germination and In Vitro Propagation of a Multipurpose Plateau Shrub Species Sophora Moorcroftiana

International Nuclear Information System (INIS)

Fu, Y.; Yao, W.; Li, S.; Li, H.

2016-01-01

Sophora moorcroftiana (Benth.) Baker (Fabaceae) with blue flowers is a shrub species in Qinghai-Tibet Plateau. This species is an important ornamental, ecological and economical species mainly distributed in the wide valleys of Yarlung Tsangpo and Lhasa River. It contains strong drought, solar radiation and sand burial resistance. However, it is on the verge of extinction because of over-exploitation in recent years, seedling production is a bottleneck in this plant's development, and information is lacking on an In vitro propagation system for S. moorcroftiana. Therefore, an effective propagation technique is urgently needed. In the present study, S. moorcroftiana seed germinations both In vitro and on an open moistened filter paper were significantly improved. The best germination percentage (88.67+-0.67) and percentage of cotyledons presented (69.67+-0.33) were obtained when the seeds were placed in moistened filter paper for 30 d after treatment with 98 percent concentration sulphuric acid for 70 min, and then treated with either Gibberellic acid (GA3) 100 mg L/sup-1/ or 500 mg L/sup -1/. Besides, an In vitro propagation system was developed for this species, the impacts of various media combined with different plant growth regulators (PGR) on multiple shoots proliferation were evaluated. The result showed Woody Plant Medium (WPM) supplemented with 1.0 mg L/sup -1/ N/sup 6/ -Benzyladenine (BA) and 0.2 mg L/sup -1/ alpha-Naphthalene acetic acid (NAA) was found to be optimum for production of multiple shoots (89.33 percent shoot regeneration frequency and 18.67 shoots per explant) using cotyledonary node. The best rooted medium for regenerated shoots (89.72 percent, 4.68 roots per shoot) was half-strength Murashige and Skoog (1/2 MS) supplemented with 0.5 mg L/sup -1/ indole-3-butyric acid (IBA) and 0.02 mg L/sup -1/ NAA. The rooted plantlets were successfully acclimated, and survival of plantlets was 75 percent after transfer to soil. These results could be

In vitro organogenesis optimization and plantlet regeneration in Citrus sinensis and C. limonia

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Almeida Weliton Antonio Bastos de

2002-01-01

Full Text Available Exogenous genes can be introduced in plants by genetic transformation techniques. However, an efficient tissue culture system with high rates of plant recovery is necessary for gene introduction. This work aimed to define organogenesis and plant regeneration protocols for sweet orange varieties Natal, Valencia and Hamlin (Citrus sinensis L. Osbeck and Rangpur lime (Citrus limonia L. Osbeck which can be used in plant transformation experiments. Seeds of which teguments were removed, were germinated in vitro and maintained in the dark for three weeks, followed by one week at 16-h photoperiod (40 µmol m-2 s-1 and 27 ± 2°C. Organogenesis induction was done by introducing epicotyl segments in MT medium with 25 g L-1 sucrose and different BAP concentrations. After adventitious bud growth, the shoots were transferred to MT medium with either NAA or IBA (1 mg L-1, or absence of auxin, for rooting. The best results were obtained with 1 mg L-1 BAP for bud induction and 1 mg L-1 IBA for rooting for all three sweet orange cultivars. The use of 0.5-2.5 mg L-1 BAP, followed by 1 mg L-1 IBA were the best growth regulator combinations for bud induction and rooting, respectively, for 'Rangpur' lime. The protocols presented in this work are suitable for associations with genetic transformation experiments for these cultivars.

Regeneration efficiency based on genotype, culture condition and growth regulators of eggplant (Solanum melongena L.

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Md Abdul Muktadir

2016-01-01

Full Text Available Several experiments were carried out to establish an efficient regenerating protocol for cultivated eggplant varieties. Among the five varieties cultured on Murashige and Skoog (MS medium with free plant growth regulator (PGR, Nayantara performed better considering the number of shoots/explant (2.48. Considering explant types and culture conditions, better performance was observed (3.68 shoots/explant when seed germination in the dark was proceeded by bottom hypocotyl segments cultured under dark conditions. A higher rate of shoot regeneration was observed in Nayantara when cultured in Zeatin Riboside (ZR and Thidizuron (TDZ supplemented MS medium. The highest number of shoots per explant was produced on MS medium supplemented with 2.0 mg/L ZR and 0.1 mg/L indole acetic acid (6.65 shoots/explant. Proliferation and elongation of the regenerated shoots were obtained in the MS medium with free PGR. The best rooting performance was observed in MS medium supplemented with 2.0 mg/L indole butyric acid. Plantlets with well developed roots and shoots were successfully transferred to soil.

Effects of donor plant age and explants on in vitro culture of Cedrela montana Moritz ex Turcz

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Sofía Basto

2012-12-01

Full Text Available To evaluate the organogenic potential of Cedrela montana Moritz ex Turcz. Explants from mature (10-20 year-old and juvenile (7-18 month-old trees were collected. The first grouping included buds, leaves, and nodes derived from juvenile basal offshoots and rejuvenated shoots from cuttings. The second, included leaves, petioles, nodes, internodes and nodes of in vitro elongated shoots. The highest organogenic potential was observed in nodes from juvenile trees: 45.8% of explants presented axillary bud elongation, while 56.2% presented rooting in a growth regulator free culture medium. Fifty-one percent of elongated shoots produced adventitious shoots with 0.5 μM NAA and 0.5 μM BA; 30% with 0.5 μM NAA and 1 μM BA; and 30% with 1 μM BA. Twenty percent presented roots with 0.5 μM NAA. Root formation was stimulated in a medium supplemented with activated charcoal (5 gL-1. The acclimatization of eighty percent of plantlets regenerated from nodes, and of 72.5% in vitro generated shoots was successful. On the contrary, mature trees material presented low organogenic response. Axillary bud elongation was recorded just in 10.7% of explants from juvenile shoots and in 6.7% of explants from rejuvenated shoots. The age of donor plant and type of explant affect the organogenic potential of C. montana. This study contributes to the understanding of this species’ response under in vitro conditions.

The Effect of Plant Growth Regulators on Shoot Proliferation and Rooting of Crataegus Pseudohetrophylla Pojark. Via in Vitro Culture

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F. Ahmadloo

2015-12-01

Full Text Available Crataegus is a tree species from Rosaceae family with medicinal, ornamental and commercial utilizations. Effect of different concentrations of plant growth regulators on shoot proliferation of Crataegus pseudohetrophylla Pojark. via in vitro culture was studied using single node explants. The grown shoots were transferred to MS medium supplemented with different concentrations of cytokinins (BAP, KIN, 2iP and Zt in combination with NAA in 30 treatments, with 3 replications and each replication included 5 explants. Data were recorded after 3 subcultures. For rooting, basal end of shoots were dipped into 300mg/l of IBA at different durations (0, 5, 10, 20, 30, 40, 50 and 60 minutes, then the dipped shoots were cultured on hormone free 1/2 VS medium. The results of analysis of variance showed that there were significant differences in all of the traits. Results indicated that the highest number of shoots (39.33 was obtained on medium containing 8 mg / l BAP plus 2 mg / l NAA. The highest shoot length (4.67 cm and leaf size (3.73 cm was achieved on 7 mg / l BAP plus 2 mg / l NAA and 2 mg / l Zt plus 1 mg / l NAA, respectively. The highest rate of rooting (33.33% and root number (4n were induced on shoots dipped for 40 minutes in 300 mg / l IBA.

Perbanyakan In Vitro Sansevieria trifasciata ‘Lorentii’: Regenerasi Tunas, Pengakaran, dan Aklimatisasi Planlet

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, Yusnita

2013-10-01

Full Text Available This research aimed to study effects of benzyladenine (BA on in vitro shoot formation and effects of indolebutyric acid(IBA and acclimatization media on ex vitro rooting and acclimatization of Sansevieria trifasciata ‘Lorentii’. Leaf segmentswere taken from young fully-expanded leaves, surface sterilized and cultured on Murashige and Skoog (MS basal mediumcontaining 0.25 mg L-1 2,4-dichlorophenoxyacetic acid (2,4-D for 2 weeks, transferred to medium without growth regulatorfor 2 weeks, and then subcultured on MS medium containing BA (0, 0.5, 1 and 2 and 5 mg L-1. The results showed thatadventitious shoot regeneration occured after callus formation. The best BA concentration was 2 mg L-1, producing 4.5 shootsper explants in 3 months and 11.1 shoots per explant in 4 months. Application of 2000 ppm IBA and the use of acclimatization  medium consisting rice husk charcoal and compost (1:1 produced the highest number of primary roots, length of roots and root fresh weight. However, the ex vitro rooting did not influence the success of plantlet acclimatization, the survival rate being 96% and there were no significant difference in plant growth.Keywords: benzyladenine, ex vitro, indolebutyric acid, 2,4-dichlorophenoxyacetic acid, micropropagation, Sansevieriatrifasciata

Development of an Efficient Regeneration and Transformation Method for Nicothiana tabacum L. through the Optimization of Growth Regulators and Sucrose Concentration

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maria Beihaghi

2018-03-01

Full Text Available Introduction: Plant tissue culture is a collection of techniques used to maintain or grow plant cells, tissues or organs under sterile conditions on a nutrient culture medium of known composition and widely used to produce clones of a plant in a method known as micropropagation. Plant research often involves growing new plants in a controlled environment. These may be plants that we have genetically altered in some way or may be plants of which we need many copies all exactly alike. These things can be accomplished through tissue culture of small tissue pieces from the plant of interest. These small pieces may come from a single mother plant or they may be the result of genetic transformation of single plant cells which are then encouraged to grow and to ultimately develop into a whole plant. Tissue culture techniques are often used for commercial production of plants as well as for plant research. Tobacco (Nicotiana tabacum L. is one of the most important model plants used in the physiologic, genetic and tissue culture studies. The manipulation of tobacco genetic structure requires an efficient technique of gene transferring and regeneration. Whereas, the tobacco plant is a very effective bioreactor in the production of recombinant proteins, in this research we optimized the best tissue culture system and also, genetic transformation process of this plant. Materials and Methods: Our plant tissue culture protocols, Include helpful information for Murashige and Skoog media, plant growth regulators, plant growth hormones, plant transformation systems, and other products for plant tissue culture. For this purpose, different concentrations of sucrose and 4 combinations of growth regulators (BAP and NAA on callus induction, direct shoot regeneration and rooting were examined in a factorial experiment based on completely randomized design with 3 replications. The sensitivity of tobacco explants to kanamycin was examined through the cultivation of them

Plant Regeneration Through Tissue Culture Of Pear Millet ...

African Journals Online (AJOL)

1. 1. 2,5), MS(5) and N6(1.100.25) culture media, calli embryogenic potential and fertile plants regeneration were conserved for more than 12 months. Characteristics of regenerated plants were similar to control. It appears that dissected shoot ...

Guiding tissue regeneration with ultrasound in vitro and in vivo

Science.gov (United States)

Dalecki, Diane; Comeau, Eric S.; Raeman, Carol H.; Child, Sally Z.; Hobbs, Laura; Hocking, Denise C.

2015-05-01

Developing new technologies that enable the repair or replacement of injured or diseased tissues is a major focus of regenerative medicine. This paper will discuss three ultrasound technologies under development in our laboratories to guide tissue regeneration both in vitro and in vivo. A critical obstacle in tissue engineering is the need for rapid and effective tissue vascularization strategies. To address this challenge, we are developing acoustic patterning techniques for microvascular tissue engineering. Acoustic radiation forces associated with ultrasound standing wave fields provide a rapid, non-invasive approach to spatially pattern cells in three dimensions without affecting cell viability. Acoustic patterning of endothelial cells leads to the rapid formation of microvascular networks throughout the volumes of three-dimensional hydrogels, and the morphology of the resultant microvessel networks can be controlled by design of the ultrasound field. A second technology under development uses ultrasound to noninvasively control the microstructure of collagen fibers within engineered tissues. The microstructure of extracellular matrix proteins provides signals that direct cell functions critical to tissue regeneration. Thus, controlling collagen microfiber structure with ultrasound provides a noninvasive approach to regulate the mechanical properties of biomaterials and control cellular responses. The third technology employs therapeutic ultrasound to enhance the healing of chronic wounds. Recent studies demonstrate increased granulation tissue thickness and collagen deposition in murine dermal wounds exposed to pulsed ultrasound. In summary, ultrasound technologies offer noninvasive approaches to control cell behaviors and extracellular matrix organization and thus hold great promise to advance tissue regeneration in vitro and in vivo.

In vitro mass propagation of Salvia canariensis by axillary shoots

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Sebastiana Mederos Molina

2014-01-01

Full Text Available During the establishment of shoots of Salvia canariensis L., five environmental factor treatments were applied. For each axillary node two shoots grew well when explants were incubated at continued ligth for 15 days followed by 16 hrs photoperiod by 30 days. Shoots multiplication was improved on a modified Murashige and Skoog (MS (1962 medium - MS + 825 mg/l NH4NO3 - supplemented with 10-7 M BA and 10-7 M NAA. The shoots produced well developed root systems within three weeks after transfer to the same culture medium supplemented with 5x 10-7 M NAA.

Shoot differentiation from protocorm callus cultures of Vanilla planifolia (Orchidaceae: proteomic and metabolic responses at early stage

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Payet Bertrand

2010-05-01

Full Text Available Abstract Background Vanilla planifolia is an important Orchid commercially cultivated for the production of natural vanilla flavour. Vanilla plants are conventionally propagated by stem cuttings and thus causing injury to the mother plants. Regeneration and in vitro mass multiplication are proposed as an alternative to minimize damage to mother plants. Because mass production of V. planifolia through indirect shoot differentiation from callus culture is rare and may be a successful use of in vitro techniques for producing somaclonal variants, we have established a novel protocol for the regeneration of vanilla plants and investigated the initial biochemical and molecular mechanisms that trigger shoot organogenesis from embryogenic/organogenic callus. Results For embryogenic callus induction, seeds obtained from 7-month-old green pods of V. planifolia were inoculated on MS basal medium (BM containing TDZ (0.5 mg l-1. Germination of unorganized mass callus such as protocorm -like structure (PLS arising from each seed has been observed. The primary embryogenic calli have been formed after transferring on BM containing IAA (0.5 mg l-1 and TDZ (0.5 mg l-1. These calli were maintained by subculturing on BM containing IAA (0.5 mg l-1 and TDZ (0.3 mg l-1 during 6 months and formed embryogenic/organogenic calli. Histological analysis showed that shoot organogenesis was induced between 15 and 20 days after embryogenic/organogenic calli were transferred onto MS basal medium with NAA (0.5 mg l-1. By associating proteomics and metabolomics analyses, the biochemical and molecular markers responsible for shoot induction have been studied in 15-day-old calli at the stage where no differentiating part was visible on organogenic calli. Two-dimensional electrophoresis followed by matrix-assisted laser desorption ionization time-of-flight-tandem mass spectrometry (MALDI-TOF-TOF-MS analysis revealed that 15 protein spots are significantly expressed (P Conclusion The

[Induction and in vitro culture of hairy roots of Dianthus caryophyllus and its plant regeneration].

Science.gov (United States)

Shi, Heping; Zhu, Yuanfeng; Wang, Bei; Sun, Jiangbing; Huang, Shengqin

2014-11-01

To use Agrobacterium rhizogenes-induced hairy roots to create new germplasm of Dianthus caryophyllus, we transformed D. caryophyllus with A. rhizogenes by leaf disc for plant regeneration from hairy roots. The white hairy roots could be induced from the basal surface of leaf explants of D. caryophyllus 12 days after inoculation with A. rhizogenes ATCC15834. The percentage of the rooting leaf explants was about 90% 21 days after inoculation. The hairy roots could grow rapidly and autonomously in liquid or solid phytohormone-free MS medium. The transformation was confirmed by PCR amplification of rol gene of Ri plasmid and silica gel thin-layer chromatography of opines from D. caryophyllus hairy roots. Hairy roots could form light green callus after cultured on MS+6-BA 1.0-3.0 mg/L + NAA 0.1-0.2 mg/L for 15 days. The optimum medium for adventitious shoots formation was MS + 6-BA 2.0 mg/L + NAA 0.02 mg/L, where the rate of adventitious shoot induction was 100% after cultured for 6 weeks. The mean number of adventitious shoot per callus was 30-40. The adventitious shoots can form roots when cultured on phytohormone-free 1/2 MS or 1/2 MS +0.5 mg/L NAA for 10 days. When the rooted plantlets transplanted in the substrate mixed with perlite sand and peat (volume ratio of 1:2), the survival rate was above 95%.

In vitro propagation, micromorphological studies and ex vitro rooting of cannon ball tree (Couroupita guianensis aubl.): a multipurpose threatened species.

Science.gov (United States)

Shekhawat, Mahipal S; Manokari, M

2016-01-01

In vitro propagation methods using seeds and nodal segments of a 21-year old Couroupita guianensis - a medicinally important but threatened tree have been developed. Hundred percent of the seeds germinated on half strength Murashige and Skoog (MS) medium with 2.0 mg l(-1) indole-3 butyric acid (IBA). Nodal segments were found most suitable for the establishment of cultures. About 90 % explants responded and 4.1 ± 0.23 shoots per node were induced after five weeks of inoculation on MS medium +4.0 mg l(-1) 6-benzylaminopurine (BAP). Further shoot multiplication was achieved by repeated transfer of mother explants and subculturing of in vitro produced shoots on fresh medium. Maximum number (8.2 ± 0.17) of shoots were regenerated on MS medium with 1.0 mg l(-1) each of BAP and Kinetin (Kin) + 0.5 mg l(-1) α-naphthalene acetic acid (NAA) with additives (50 mg l(-1) of ascorbic acid and 25 mg l(-1) each of adenine sulphate, L-arginine and citric acid). The multiplied shoots rooted (4.3 ± 0.26 roots/shoot) on half strength MS medium with 2.5 mg l(-1) IBA. All the shoots were rooted ex vitro when pulse treated with 400 mg l(-1) of IBA for five min with an average of 7.3 ± 0.23 roots per shoot. Nearly 86 % of these plantlets were acclimatized within 7-8 weeks and successfully transferred in the field. Biologically significant developmental changes were observed during acclimation particularly in leaf micromorphology in terms of changes in stomata, veins and vein-islets, and trichomes. This study helps in understanding the response by the plants towards outer environmental conditions during acclimatization. This is the first report on micropropagation of C. guianensis, which could be used for the large-scale multiplication, restoration and conservation of germplasm of this threatened and medicinally important tree.

Efficient genetic transformation of Lotus corniculatus L. using a direct shoot regeneration protocol, stepwise hygromycin B selection, and a super-binary Agrobacterium tumefaciens vector

Directory of Open Access Journals (Sweden)

Nikolić Radomirka

2007-01-01

Full Text Available Cotyledons from 6-day-old Lotus corniculatus cv. Bokor seedlings, transversally cut into two halves, were capa­ble of regenerating buds without intervening callus formation. The explants were co-cultivated with the Agrobacterium tumefaciens LBA4404/pTOK233 superbinary vector carrying the uidA-intron gene and the genes hpt and nptII. They were cultured for 14 days on a regeneration medium, then subjected to a stepwise hygromycin B selection procedure consisting of gradually increasing antibiotic concentrations (5-15 mg L-1 over 21 weeks. Transformed shoots were obtained within 5 months after co-cultivation. Out of 124 initially co-cultivated explants, 52 (42% plants survived hygromycin B selection. The presence of transgenes in regenerated plants was verified by β-glucuronidase histochemical assays and PCR analysis for the presence of uidA gene sequences. Hygromycin B-resistant and PCR-positive T0 plants were cultured in the greenhouse to produce flowers and seeds. The obtained data demonstrate that the reported transformation protocol could be useful for introducing agriculturally important genes into the new L. corniculatus cultivar Bokor.

High frequency plant regeneration from shoot tip explants of ...

African Journals Online (AJOL)

USER

2010-08-02

Aug 2, 2010 ... 16/8-h (light/dark) photoperiod provided by cool white fluorescent light. Multiple .... formation from shoot tip explant of C. colocynthis on MS-medium. S. No. .... micropropagation of Musa sapientum L. (Cavendish Dwarf). Afr. J.

The antioxidant effect of Asparagus cochinchinensis (Lour. Merr. shoot in d-galactose induced mice aging model and in vitro

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Linghua Lei

2016-04-01

Conclusion: The aqueous extract of A. cochinchinensis (Lour. Merr. shoot had a strong radical scavenging capability in vivo and in vitro, and might be used to diminish radicals in the body and consequently prevent aging.

Application of in vitro flowering technique on evaluating of mutation capacity and colour selection of Torenia fournieri L. following irradiation

International Nuclear Information System (INIS)

Le Van Thuc; Le Thi Thuy Linh; Hoang Hung Tien; Dang Thi Dien; Le Thi Bich Thy; Han Huynh Dien

2013-01-01

The transformation from vegetative to reproductive stage (flowering stage) depends on a lot of elements: physical and chemical elements, age of explant source, and plant growth regulators. In this study, we examined some elements that affect to the differentiating ability of in vitro Torenia fournieri L. flower shoot. Results showed that the older the shoots were, the higher the percentage of flowering shoot formation would be 60-day-old shoots gave the highest percentage of flowering shoot formation (65%) after 30-day culture. Moreover, the amount of macronutrients and micronutrients, sucrose concentration, growth regulators, activated charcoal and lighting conditions affect significantly the flowering ability. Shoots cultured on plant growth regulator free 1/4MS media supplemented with 60 g.l -1 sucrose and 1 g.l -1 activated charcoal in vessels covered with either plastic wrap with millipore filter or unabsorbable cotton-wool plug under a 10 h light (45 µmol.m -2 .s -1 )/14 h dark photoperiod resulted in the best flowering shoot formation of Torenia shoots cultured in vitro (89.18%, 2.80 flower buds/explant). In vitro flowers and the ex vitro ones have no significant difference in their morphology and color. Gamma-ray irradiation is one of the most effective techniques to produce mutants in plant breeding programs. In this research, the effect of Co 60 gamma-rays to produce flower color mutants in combination with in vitro flowering technique for morphological change isolation in Torenia was also investigated. After treated with gamma-rays, shoot regeneration was obtained when leaf explants were cultured on MS medium containing 0.5 mg/l BAP and 0.5 mg/l NAA. Shoots were grown on MS medium supplemented with 0.2 mg/l BA for shoot proliferation. The mutation efficiencies of the gamma-ray irradiation for flower color was examined owing to in vitro flowering technique. LG 50 at 30 - 40 Gy and 0.098 Gy/sec for leaf blades as well as plantlets was found to be suitable

In vitro root induction of faba bean (Vicia faba L.).

Science.gov (United States)

Ismail, Roba M; Elazab, Heba E M; Hussein, Gihan M H; Metry, Emad A

2011-01-01

A major challenge for regeneration of faba bean (Vicia faba L.) plants is the difficulty of in vitro root induction. In the present study, in vitro rooting and its architecture have been studied. Adventitious root formation was successfully induced from regenerated faba bean shoots of four Egyptian cultivars, i.e., Giza 461, Giza 40, Giza 834 and Giza 716 on hormone free MS medium supplemented with 5 mg/l silver nitrate. Among the four cultivars, Giza 461 and Giza 40 were recorded as the highest root formation response (75 % and 65) followed by cultivars Giza716 and Giza843 (20%, and 10%). Anatomical study proved that the produced roots are initiated as the adventitious lateral root (LR) with tri-arch xylem strands as compared with the penta-arch of the primary roots of the intact faba bean seedling. The obtained results overcome the root induction problem in faba bean.

Effects of calcium gluconate and ascorbic acid on controlling shoot ...

African Journals Online (AJOL)

In vitro shoot necrosis is a quite widespread disorder affecting raspberry micropropagation. This study was conducted to investigate effects of calcium gluconate and ascorbic acid on shoot necrosis and dieback of raspberry shoots during micropropagation. Nodal segments of primocane-fruiting raspberry cultivars 'Allgold', ...

In vitro propagation of critically endangered species Scilla autumnalis L. – biochemical analyses of the regenerants

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Cristian BANCIU

2010-11-01

Full Text Available The present study belongs to the international efforts for plant conservation from the areas threatened by human activities. The saline soils areas are restricting for agriculture and in some cases for fishery facilities and the plant species are extinct from those areas. Scilla autumnalis L. is one of the threatened plants (rare on the national red list of vascular plants from Romania that grows in the Natural Park Comana, Giurgiu County, South Romania. Seeds from plants grown in the natural habitat have been used for in vitro plant regeneration and multiplication. After successfully rooting and acclimatization of the regenerated plantlets from germinated seeds, biochemical studies have been performed in order to compare the regenerants from in vitro cultures with native plants from genetically point of view. Peroxydase and esterase’s spectra were the biochemical markers used.The results indicated that this plant species can be multiplicated, rooted and acclimatized on synthetic medium (MS supplemented with NAA, IBA, IAA, kinetin and BAP with a good efficiency and the regenerants had no genetic alterations determinated by culture conditions.

PLETHORA genes control regeneration by a two-step mechanism

NARCIS (Netherlands)

Kareem, Abdul; Durgaprasad, Kavya; Sugimoto, Kaoru; Du, Yujuan; Pulianmackal, Ajai J.; Trivedi, Zankhana B.; Abhayadev, Pazhoor V.; Pinon, Violaine; Meyerowitz, Elliot M.; Scheres, Ben; Prasad, Kalika

2015-01-01

Summary Regeneration, a remarkable example of developmental plasticity displayed by both plants and animals, involves successive developmental events driven in response to environmental cues. Despite decades of study on the ability of the plant tissues to regenerate a complete fertile shoot

In vitro techniques for mutation breeding of tropical root and tuber crops

International Nuclear Information System (INIS)

Novak, F.

1987-01-01

Full text: To assist IAEA Technical Co-operation projects, the Agricultural Section of the IAEA Laboratory in Seibersdorf is developing techniques for in vitro mutation breeding of cassava (Manihot esculenta) and yam (Dioscorea alata, D. rotundata). The first aim was to induce morphogenesis (plant regeneration) in tissue culture and establish techniques for in vitro propagation. Subsequently, the in vitro mutation breeding technology is being developed. (i) Cassava is one of the important staple food crops of tropical countries. Pest and disease resistance as well as low toxic cyanide content are among the objectives for genetic improvement. For in vitro mutation induction we use shoot-tip and node culture. Shoot apices (1 and 2 mm long) are aseptically dissected from cassava buds and cultured on MS medium with 1 mg/l thiamine, -naphthalene acetic acid, 6-benzyladenine and gibberellic acid. Elevated concentration of 6-benzyladenine is used for multiple shoot formation. The rapid multiplication was induced in liquid medium, when flasks were placed on a gyratory shaker with 60 rpm at 28 deg. C during 16/8 light/dark photoperiod. Nodes with axillary buds from in vitro growing plantlets were irradiated with gamma rays. Doses of 30 to 45 Gy allowed the survival of approx. 50 percent of explants and subsequent shoot proliferation from axillary buds. Radiosensitivity of cassava genotypes may be different and this will be investigated in future experiments. (ii) Yams are likewise important tuber crops, particularly in West Africa, South-East Asia and the Caribbean. The main breeding objectives are improved yield, shortened growth period, improved storability (resistance of tubers to fungal attack), shoot tip cultures have been utilized for clonal propagation, and germplasm preservation and exchange. At present, the IAEA Laboratory at Seibersdorf is trying this technique for mutation induction. Somatic embryogenesis in cell and tissue culture is worked on to develop a

Plant regeneration from organ culture in white Guinea Yam

Energy Technology Data Exchange (ETDEWEB)

Nwachukwu, E C; Mbanaso, E N.A. [National Root Crops Research Inst., Umudike, Umuahia, Abia State (Nigeria); Sonnino, A [Centro Recerche Energia, ENEA, Rome (Italy)

1997-07-01

Explants from leaves, leaf segments, petioles and internodal stem of in vitro grown seedlings of white guinea yam, Dioscorea rotundata Poir, cv. `Obiaoturugo` were cultured on defined media. NAA at concentrations of 0.5-1.0 mg/1 induced shoot regeneration from petiolar and inter-nodal stem pieces, and rooting occurred with little or no callusing from whole leaves or leaf segments. With concentration of 3.0-10.0 m/1 NAA, explants from petioles, inter-nodal stem, whole leaves and leaf segments formed callus which produced roots. These explants developed plantlets when subcultured on MS medium supplemented with 2.0 mg/1 BAP and 0.1 mg/1 NAA. (author). 11 refs, 1 tab.

Plant regeneration from organ culture in white Guinea Yam

International Nuclear Information System (INIS)

Nwachukwu, E.C.; Mbanaso, E.N.A.; Sonnino, A.

1997-01-01

Explants from leaves, leaf segments, petioles and internodal stem of in vitro grown seedlings of white guinea yam, Dioscorea rotundata Poir, cv. 'Obiaoturugo' were cultured on defined media. NAA at concentrations of 0.5-1.0 mg/1 induced shoot regeneration from petiolar and inter-nodal stem pieces, and rooting occurred with little or no callusing from whole leaves or leaf segments. With concentration of 3.0-10.0 m/1 NAA, explants from petioles, inter-nodal stem, whole leaves and leaf segments formed callus which produced roots. These explants developed plantlets when subcultured on MS medium supplemented with 2.0 mg/1 BAP and 0.1 mg/1 NAA. (author). 11 refs, 1 tab

IN VITRO ORGANOGENESIS OF TWO SANSEVIERIA CULTIVARS ON DIFFERENT CONCENTRATIONS OF BENZYLADENINE (BA

Directory of Open Access Journals (Sweden)

Yusnita

2011-06-01

Full Text Available In vitro regeneration of buds and shoots via organogenesis in two genotypes of Sansevieria trifasciata was established. Leaf segments (1cm x 1cm of cv. Hahnii and cv. Lorentii were cultured on Murashige and Skoog (MS basal medium containing 2.4-dichloro-phenoxyacetic acid (2.4-D for 1 week, trans-ferred into MS medium without plant growth regulator (MS0 for 1 week, and then cultured to MS medium containing different concentrations of benzyladenine (BA:0; 0.1; 0.25; 0.5; 1 and 2 mg/l for 3 consecutive passages with 4 week intervals. The results showed that BA proved to be an effective cytokinin to induce the formation of adventitious buds and shoots in two cultivars of Sansevieria trifasciata. The maximum of 12 propagules per explant of Lorentii and 9.3 propagules of Hahnii were obtained in medium with 2 mg/l BA after 14 weeks. Furthermore, regenerative capacity to form shoot buds and propagules was genotype dependent. The popagules number formed by cv. Lorentii was significantly higher than those formed by cv. Hahnii. The average shoot length formed by cv. Lorentii was also higher than those of cv. Hahnii. Shoots of both cultivars were successfully ex vitro rooted and acclimatized to the greenhouse with high survival rate (95.9 -100%.

Regeneration and acclimatization of salt-tolerant arachis hypogaea plants through tissue culture

International Nuclear Information System (INIS)

Ghauri, E.G.

2006-01-01

Excised embryos of Arachis hypogaea were cultured on Murashige and Skoog's medium (MS medium) supplemented with different combinations of growth hormones. The highest frequency of callus proliferation (80%) was recorded on MS medium mixed with 1.0 mg/1 of 2,4-D and 0.5 mg/1 of BAP. These cultures were treated with 0.65 mg/l of trans-4-hydroxy-L-proline (HyP) a:1d various concentrations (0.1-0.5%) of NaCl. In all cases the presence of salt reduced the fresh mass of callus. Shoot regeneration in the cultures took place when transferred to MS medium supplemented with 1.0 mg/1 of kinetin (Kin) and 0.5 mg/1 of 6-benzyl aminopurine (BAP). Percentage of shoot regeneration decreased with the increase of NaCl (0.1- 0.5%) in the shoot regeneration medium. Root formation in these cultures took place when the cultures were nurtured on MS medium free of growth hormones. Regeneration, hardening and acclimatization of the salt tolerant plants was conducted. (author)

Regeneration of Centella asiatica plants from non-embryogenic cell lines and evaluation of antibacterial and antifungal properties of regenerated calli and plants

Directory of Open Access Journals (Sweden)

Habib Darima

2011-10-01

Full Text Available Abstract Background The threatened plant Centella asiatica L. is traditionallyused for a number of remedies. In vitro plant propagation and enhanced metabolite production of active metabolites through biotechnological approaches has gained attention in recent years. Results Present study reveals that 6-benzyladenine (BA either alone or in combination with 1-naphthalene acetic acid (NAA supplemented in Murashige and Skoog (MS medium at different concentrations produced good quality callus from leaf explants of C. asiatica. The calli produced on different plant growth regulators at different concentrations were mostly embryogenic and green. Highest shoot regeneration efficiency; 10 shoots per callus explant, from non-embryogenic callus was observed on 4.42 μM BA with 5.37 μM NAA. Best rooting response was observed at 5.37 and 10.74 μM NAA with 20 average number of roots per explant. Calli and regenerated plants extracts inhibited bacterial growth with mean zone of inhibition 9-13 mm diameter when tested against six bacterial strains using agar well diffusion method. Agar tube dilution method for antifungal assay showed 3.2-76% growth inhibition of Mucor species, Aspergillus fumigatus and Fusarium moliniformes. Conclusions The present investigation reveals that non-embryogenic callus can be turned into embryos and plantlets if cultured on appropriate medium. Furthermore, callus from leaf explant of C. asiatica can be a good source for production of antimicrobial compounds through bioreactor.

Sterilization of African Violet in the in Vitro Culture Using Synthesized Silver Nanoparticles by Two Plant Extracts

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M. Solgi

2015-12-01

Full Text Available One of the major advantages of in vitro culture of African violet (Saintpaulha ionantha is production of new cultivars and propagation of their chimera which might not be propagated by the other methods. In this study, we tested the effects of silver nanoparticles on the sterilization rate (antifungal and antibacterial activity, regeneration and shoot formation of African violet "Pink Amiss" explants. These nanoparticles were synthesized from pomegranate peels and Damask rose petals extracts. We used a completely randomized design test with factorial arrangement to investigate various volumetric ratios of plant extracts to silver nitrate (1:20, 1:10, 1:5 and 1:1 on the culture contaminations. Using silver nanoparticles synthesized by the plant extracts, especially Damask rose petals extract resulted in no fungal and bacterial contamination in the African violet explants after 1 and 3 weeks as compared to the control, and silver nitrate (1mM. All tested concentrations of the silver nanoparticles significantly (P &le 0.05 controlled both bacterial and fungal contaminations. The 1:20 ratio of plant extracts to silver nitrate showed the best control. In addition, the highest regeneration (%52 and shoot regeneration (%38 was observed in this treatment. In conclusion, we suggest using silver nanoparticles synthesized by plant extracts for sterilization of in Vitro Culture for African Violet rather than using other chemicals such as silver nitrate.

Genetic stability among date palm plantlets regenerated from petiole ...

African Journals Online (AJOL)

The regeneration capacity between the shoot tip and petiole explants that excised from two date palm (Phoenix dactylifera L.) cutivar namely Unknown and Ferhi was compared. It was noted that the shoot tip explants started to swill after several subculture when placed on Murashige and Skoog (MS) medium supplemented ...

Increase in the rate of recombinants in tomato (Lycopersicon esculentum L.) after in vitro regeneration.

Science.gov (United States)

Sibi, M; Biglary, M; Demarly, Y

1984-07-01

Modification to the cross-over (C. O.) rate of tomato (Lycopersicon esculentum) was attempted by using in vitro plant regeneration. F1 hybrids with the same genetical homozygous background were compared at two loci: "bs-ms32" on chromosome I, and "aa-d" on chromosome II. For each, the genetic distance separating the two markers was about 20 to 30 map units. One cotyledon of each F2 hybrid seedling was used as in vitro tissue culture material, while the rest of the plantlet was grown as a control. Recombination rates of the selfed progenies from each regenerated and matched control couple were compared. For the first set of markers 59,000 seeds were analysed (5 controls' and 7 regenerated progenies), and for the second, 11,000 (5 controls' and 8 regenerated progenies). There were significant increases in the genetic distance between markers in about half the regenerated individuals. For the first set the increases ranged from 6.07 to 6.91 units out of a control distance of the 19.84 to 25.65, corresponding to lengthenings of 30.59 to 35.29%. For the second set they ranged from 4.92 to 6.04 out of a control distance of 25.05 to 26.57, representing increases of 19.64 to 22.75%. Such a phenomenon can be important either from a fundamental or practical viewpoint, regarding selection efficiency in plants, and potential for gene reassortment.

Effect of in vitro mutagenesis on plant regeneration in Citrus aurantifolia S

International Nuclear Information System (INIS)

Srivastava, R.K.; Sandhu, A.S.; Gosal, S.S.

2001-01-01

Callus was induced from different explants excised from in vitro raised seedlings on MS medium enriched with naphthalene acetic acid (NAA) (10 mg/l) and kinetin (0.2 mg/l). The cultures were maintained on the same media for 30 days. Part of the 30-day-old calli were exposed to gamma radiation (5 and 10 Gy) and the rest were treated with ethyl methanesulphonate (EMS) (0.1 to 0.4%) for 8 hours. All the treated calli were immediately transferred to regeneration medium [1/2 MS+Benzyl Amino Purine (BAP) (5 mg/l)] along with the untreated control. The cultures were maintained under conditions of 25±2 deg. C, 16/8 hours day and night regime and 2500-3000 lux light intensity. The results indicated a significant effect of mutagenic agent on callus regeneration and regenerants' morphological features. The same phenomenon was observed in Triticum aestivum and Zea mays. Regenerated mutants showed variation in morphological traits like, plant height, leaf length and breadth. Moreover, the mutants are being screened for resistance against citrus canker. However, the genetic origin of the mutants has not been determined

Biotechnological applications in in vitro plant regeneration studies of broccoli (Brassica oleracea L. var. italica), an important vegetable crop.

Science.gov (United States)

Kumar, Pankaj; Srivastava, Dinesh Kumar

2016-04-01

Biotechnology holds promise for genetic improvement of important vegetable crops. Broccoli (Brassica oleracea L. var. italica) is an important vegetable crop of the family Brassicaceae. However, various biotic and abiotic stresses cause enormous crop yield losses during commercial cultivation of broccoli. Establishment of a reliable, reproducible and efficient in vitro plant regeneration system with cell and tissue culture is a vital prerequisite for biotechnological application of crop improvement programme. An in vitro plant regeneration technique refers to culturing, cell division, cell multiplication, de-differentiation and differentiation of cells, protoplasts, tissues and organs on defined liquid/solid medium under aseptic and controlled environment. Recent progress in the field of plant tissue culture has made this area one of the most dynamic and promising in experimental biology. There are many published reports on in vitro plant regeneration studies in broccoli including direct organogenesis, indirect organogenesis and somatic embryogenesis. This review summarizes those plant regeneration studies in broccoli that could be helpful in drawing the attention of the researchers and scientists to work on it to produce healthy, biotic and abiotic stress resistant plant material and to carry out genetic transformation studies for the production of transgenic plants.

In vitro and in vivo investigations on bone regeneration potential of laminated hydroxyapatite/gelatin nanocomposite scaffold along with DBM

Energy Technology Data Exchange (ETDEWEB)

Tavakol, Shima [School of Advanced Technologies in Medicine, Tehran University of Medical Sciences, Department of Medical Nanotechnology (Iran, Islamic Republic of); Ragerdi Kashani, Iraj [School of Medicine, Tehran University of Medical Sciences, Department of Anatomy (Iran, Islamic Republic of); Azami, Mahmood [School of Advanced Technologies in Medicine, Tehran University of Medical Sciences, Department of Tissue Engineering (Iran, Islamic Republic of); Khoshzaban, Ahad [Tehran University of Medical Sciences, Iranian Tissue Bank Research and Preparation Center (Iran, Islamic Republic of); Tavakol, Behnaz [Kashan University of Medical Sciences, Department of Medicine (Iran, Islamic Republic of); Kharrazi, Sharmin [School of Advanced Technologies in Medicine, Tehran University of Medical Sciences, Department of Medical Nanotechnology (Iran, Islamic Republic of); Ebrahimi, Somayeh [University of Tarbiat Moallem, Department of Biology, Faculty of Sciences (Iran, Islamic Republic of); Rezayat Sorkhabadi, Seyed Mahdi, E-mail: sh_tavakol@razi.tums.ac.ir [School of Advanced Technologies in Medicine, Tehran University of Medical Sciences, Department of Medical Nanotechnology (Iran, Islamic Republic of)

2012-12-15

Bone regeneration ability of a scaffold strongly depends on its structure and the size of its components. In this study, a nanostructured scaffold was designed for bone repair using nano hydroxyapatite (nHA) (8-16 nm Multiplication-Sign 50-80 nm) and gelatin (GEL) as main components. In vitro investigations of calcium matrix deposition and gene expression of the seeded cells for this scaffold, demineralized bone matrix (DBM), scaffold plus DBM, and the control group were carried out. Bone regeneration in rat calvarium with critical defect size after 1, 4, and 8 weeks post implantation was investigated. The calcium matrix depositions by the osteoblast and RUNX2, ALP, osteonectin, and osteocalcin gene expression in scaffold were more significant than in other groups. Histomorphometry analysis confirmed in vitro results. In vitro and in vivo bone regeneration were least in scaffold plus DBM group. Enhanced effects in scaffold could be attributed to the shape and size of nHA particles and good architecture of the scaffold. Reduction of bone regeneration might be due to tight bonding of BMPs and nHA particles in the third group. Results obtained from this study confirmed that nano-scale size of the main components and the scaffold architecture (pore diameter, interconnectivity pores, etc.) have significant effects on bone regeneration ability of the scaffold and are important parameters in designing a temporary bone substitute.

In vitro and in vivo investigations on bone regeneration potential of laminated hydroxyapatite/gelatin nanocomposite scaffold along with DBM

International Nuclear Information System (INIS)

Tavakol, Shima; Ragerdi Kashani, Iraj; Azami, Mahmood; Khoshzaban, Ahad; Tavakol, Behnaz; Kharrazi, Sharmin; Ebrahimi, Somayeh; Rezayat Sorkhabadi, Seyed Mahdi

2012-01-01

Bone regeneration ability of a scaffold strongly depends on its structure and the size of its components. In this study, a nanostructured scaffold was designed for bone repair using nano hydroxyapatite (nHA) (8–16 nm × 50–80 nm) and gelatin (GEL) as main components. In vitro investigations of calcium matrix deposition and gene expression of the seeded cells for this scaffold, demineralized bone matrix (DBM), scaffold plus DBM, and the control group were carried out. Bone regeneration in rat calvarium with critical defect size after 1, 4, and 8 weeks post implantation was investigated. The calcium matrix depositions by the osteoblast and RUNX2, ALP, osteonectin, and osteocalcin gene expression in scaffold were more significant than in other groups. Histomorphometry analysis confirmed in vitro results. In vitro and in vivo bone regeneration were least in scaffold plus DBM group. Enhanced effects in scaffold could be attributed to the shape and size of nHA particles and good architecture of the scaffold. Reduction of bone regeneration might be due to tight bonding of BMPs and nHA particles in the third group. Results obtained from this study confirmed that nano-scale size of the main components and the scaffold architecture (pore diameter, interconnectivity pores, etc.) have significant effects on bone regeneration ability of the scaffold and are important parameters in designing a temporary bone substitute.

In-vitro neoformation of woody plants (apple, pear, Weigela)

International Nuclear Information System (INIS)

Chevreau, E.; Dufour, M.; Duron, M.

1990-01-01

Full text: Adventitious bud neoformation may induce variability in two ways: somaclonal variation, if the explant has undergone undifferentiated growth (callus); induced mutations, if a physical or chemical mutagen has been applied to the explant prior to the regeneration. Three apple scion cultivars ('Gala', 'Granny Smith', 'Golden Delicious') and four apple rootstock cultivars ('Mark', 'Novole', 'Lancep', 'Cepiland') were grown on basal apple multiplication medium. Leaves, internodes and roots were used for callogenesis and/or regeneration experiments. Scion varieties were more responsive than rootstocks. 1000 'Gala' neoformations will be planted in the field to investigate somaclonal variation. In pear, shoot regeneration was obtained from in-vitro leaves of 3 varieties of Pyrus communis ('Seckel', 'Louise Bonne', 'Comice') and one variety of Pyrus bretschneideri ('Crystal Pear'). It was observed that regeneration is still possible from leaves irradiated with up to 40 Gy of γ rays, or treated with 5mM ENU for one hour. In Weigela, stem segments (0.5 cm) from rooted in vitro plants of 5 cultivars were used in experiments to induce adventitious buds. On the cultivar 'Eva Rathke' the adventitious buds were detected as early as 10 to 15 days after the beginning of the culture. A mutagenic treatment was carried out by immersing explants of 'Bristol Ruby' in a suspension of ethyl methane sulfonate (EMS). Among 400 plants regenerated from treated explants, 5 mutants were detected after 5 years of field observation. Two of them seem to be solid mutants. Apparently in Weigela both homogeneous mutants and chimeras are produced by adventitious regeneration from mutagenised explants. (author)

Rapid plant regeneration of chrysanthemum (Chrysanthemum ...

African Journals Online (AJOL)

STORAGESEVER

2009-05-04

May 4, 2009 ... demand for chrysanthemum, it has become one of the first commercial targets for ... frequency of multiple shoot regeneration response was. 95 and 91%, for nodal ..... Dordrecht, The Netherlands, pp. 91-101. Prasad RN ...

Regeneration and growth rates of allofragments in four common stream plants

DEFF Research Database (Denmark)

Riis, Tenna; Madsen, Tom Vindbæk; Sennels, R. S. H.

2009-01-01

perfoliatus L. and Ranunculus baudotii x pseudofluitans. The objectives of this study were to determine (1) if shoots with an apical tip have higher regeneration (growth of new shoots and rhizomes from allofragments) and colonisation (root attachment in sediment) abilities and higher relative growth rates...

In vitro shoot multiplication of Ziziphus spina-christi by shoot tip culture

African Journals Online (AJOL)

USER

2010-02-08

Feb 8, 2010 ... Key words: Clonal propagation, cidir, shoot tip culture, Ziziphus spina-christi (L.) Desf. INTRODUCTION. Ziziphus spina-christi (L.) Desf., locally known as cidir, is a multipurpose tree species belonging to the botanical family Rhamnaceae. It is an important cultivated tree and one of the few truly native tree ...

[Influence of genotype, explant type and component of culture medium on in vitro callus induction and shoot organogenesis of tomato (Solanum lycopersicum L.)].

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Khaliluev, M R; Bogoutdinova, L R; Baranova, G B; Baranova, E N; Kharchenko, P N; Dolgov, S V

2014-01-01

The influence of explant type as well as of the type of growth regulators and concentration on callus induction processes and somatic organogenesis of shoots was studied in vitro on four tomato genotypes of Russian breeding. Cytological study of callus tissue was conducted. It was established that tomato varieties possess a substantially greater ability to indirect shoot organogenesis compared with the F1 hybrid. The highest frequency of somatic organogenesis of shoots, as well as their number per explant, was observed for most of the genotypes studied during the cultivation of cotyledons on Murashige-Skoog culture medium containing 2 mg/l of zeatin in combination with 0.1 mg/l of 3-indoleacetic acid. An effective protocol of indirect somatic organogenesis of shoots from different explants of tomato varieties with a frequency of more than 80% was developed.

In vitro aging of mineralized collagen-based composite as guided tissue regeneration membrane

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Pan, S.X. [Department of Prothodontics, School of Stomatology, Peking University, Beijing 100875 (China)]. E-mail: sx_pan@sina.com; Li, Y. [Department of Materials Science and Engineering, Tsinghua University, Beijing 100084 (China); Feng, H.L. [Department of Prothodontics, School of Stomatology, Peking University, Beijing 100875 (China); Bai, W. [Department of Prothodontics, School of Stomatology, Peking University, Beijing 100875 (China); Gu, Y.Y. [Department of Materials Science and Engineering, Tsinghua University, Beijing 100084 (China)

2006-05-15

The technique of guided tissue regeneration (GTR) has been developed for the regeneration of periodontal tissues, bone around natural teeth and dental implants. The aim of this study is to investigate the biodegradability and mechanic behavior of a novel mineralized nano-hydroxyapatite/collagen/poly (lactic acid) (nHAC/PLA) composite as GTR membrane in vitro. The elastic modulus and maximum tensile strength of GTR film samples with different nHAC/PLA ratio were measured to get an optimal nHAC/PLA ratio. Thermogravimetric analysis was conducted to evaluate the change of the inorganic component in the samples during the process of in vitro aging. Morphology of samples was checked by using scanning electron microscopy. On the basis of the above results, it can be concluded that the GTR membranes maintained integrity and the original appearance throughout the 1-month in vitro aging. There is an active dissolution and deposition process of crystals which is propitious to the bone formation on the surface of the composite membrane. The optimal nHAC/PLA ratio of the novel membrane is 0.4:1. For a longer period of bone repair, PLA with higher molecular weight should be chosen as the scaffold for the GTR membrane.

Evaluation of the tolerance to Finale® in the germination and regeneration of Cuban rice varieties (IACuba-17 and IACuba-19

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Daymí Abreu

2005-01-01

Full Text Available Selection agent used during the shoot selection has an important role on the transgenic plant generation efficiency. In this work, the tolerance to the herbicide Finale® in two Cuban rice cultivars, IACuba17 and IACuba-19 was evaluated, and determined that 10 days exposure to 5 and 10 mg.l-1 of Finale® were enough to avoid seedlings of IAC-17 and IAC-19, respectively. Cultivated calluses (0, 2, 4 and 6 days in the absence of Finale®in the regeneration medium were used to evaluate the minimal concentration of Finale®that totally inhibits shoot regeneration. Pre-induced calluses cultured during two days and 3 mg.l-1 of Finale® in the regeneration medium was the most efficient combination to select shoots during the generation of transgenic plants resistant to the herbicide. Our shoot selection procedure reduces to 3 weeks the time to obtain shoots during the generation of transgenic rice plants. Key words: germination, mature seeds, Oryza, phosphinothricin, regeneration, selection markers

Genotype dependent callus induction and shoot regeneration in ...

African Journals Online (AJOL)

This study aims to observe the effect of genotype, hormone and culture conditions on sunflower (Helianthus annuus L.) callus induction and indirect plant regeneration. Calli were obtained from hypocotyl and cotyledon explants of five different sunflower genotypes; Trakya 80, Trakya 129, Trakya 259, Trakya 2098 and ...

In vitro propagation and assessment of the genetic fidelity of Musa acuminata (AAA) cv. Vaibalhla derived from immature male flowers.

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Hrahsel, Lalremsiami; Basu, Adreeja; Sahoo, Lingaraj; Thangjam, Robert

2014-02-01

An efficient in vitro propagation method has been developed for the first time for Musa acuminata (AAA) cv. Vaibalhla, an economically important banana cultivar of Mizoram, India. Immature male flowers were used as explants. Murashige and Skoog's (MS) medium supplemented with plant growth regulators (PGRs) were used for the regeneration process. Out of different PGR combinations, MS medium supplemented with 2 mg L(-1) 6-benzylaminopurine (BAP) + 0.5 mg L(-1) α-naphthalene acetic acid (NAA) was optimal for production of white bud-like structures (WBLS). On this medium, explants produced the highest number of buds per explant (4.30). The highest percentage (77.77) and number (3.51) of shoot formation from each explants was observed in MS medium supplemented with 2 mg L(-1) kinetin + 0.5 mg L(-1) NAA. While MS medium supplemented with a combination of 2 mg L(-1) BAP + 0.5 mg L(-1) NAA showed the maximum shoot length (14.44 cm). Rooting efficiency of the shoots was highest in the MS basal medium without any PGRs. The plantlets were hardened successfully in the greenhouse with 96% survival rate. Random amplified polymorphic DNA (RAPD) and inter-simple sequence repeat (ISSR) markers were employed to assess the genetic stability of in vitro regenerated plantlets of M. acuminata (AAA) cv. Vaibalhla. Eight RAPD and 8 ISSR primers were successfully used for the analysis from the 40 RAPD and 30 ISSR primers screened initially. The amplified products were monomorphic across all the regenerated plants and were similar to the mother plant. The present standardised protocol will find application in mass production, conservation and genetic transformation studies of this commercially important banana.

Prolific plant regeneration through organogenesis from scalps of ...

African Journals Online (AJOL)

Four types of potting media comprising of sand, peat, sand + top soil + goat dung (3:2:1 v/v) and top soil + sand (1:1 v/v) were evaluated during acclimatization of the plantlets. Prolific shoot regeneration from scalps was obtained on MS medium containing 2.5 mM BAP, at 9.61 and 40.6 shoots per explant after 4 and 8 weeks ...

In Vitro and Cryopreservation Techniques for Conservation of Snow Mountain Garlic.

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Mahajan, Ritu

2016-01-01

Garlic is an important medicinal herb of culinary value by imparting its flavors and odors to the food. Allicin, a notable flavonoid in garlic, is a powerful antibiotic and antifungal compound. Due to poor bioavailability, garlic is of limited use for oral human consumption. Being sexually sterile, propagation of garlic is done by individual cloves from a bulb which increases the chances of transfer of viral diseases. In this chapter, an efficient and improved regeneration protocol for explant establishment and shoot multiplication under in vitro conditions is described. A high rate of shoot multiplication is obtained on MS medium supplemented with 0.5 mg/l BAP, 1.0 mg/l KN, and 2.0 mg/l GA3. Addition of 1.0 mg/l NAA to MS medium resulted in rooting at the shoot bases. A detailed method for encapsulation of explant in sodium alginate beads and their cryopreservation using encapsulation-dehydration is also described.

Human skeletal muscle fibroblasts stimulate in vitro myogenesis and in vivo muscle regeneration.

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Mackey, Abigail L; Magnan, Mélanie; Chazaud, Bénédicte; Kjaer, Michael

2017-08-01

Accumulation of skeletal muscle extracellular matrix is an unfavourable characteristic of many muscle diseases, muscle injury and sarcopenia. The extent of cross-talk between fibroblasts, as the source of matrix protein, and satellite cells in humans is unknown. We studied this in human muscle biopsies and cell-culture studies. We observed a strong stimulation of myogenesis by human fibroblasts in cell culture. In biopsies collected 30 days after a muscle injury protocol, fibroblast number increased to four times control levels, where fibroblasts were found to be preferentially located immediately surrounding regenerating muscle fibres. These novel findings indicate an important role for fibroblasts in supporting the regeneration of muscle fibres, potentially through direct stimulation of satellite cell differentiation and fusion, and contribute to understanding of cell-cell cross-talk during physiological and pathological muscle remodelling. Accumulation of skeletal muscle extracellular matrix is an unfavourable characteristic of many muscle diseases, muscle injury and sarcopenia. In addition to the indispensable role satellite cells play in muscle regeneration, there is emerging evidence in rodents for a regulatory influence on fibroblast activity. However, the influence of fibroblasts on satellite cells and muscle regeneration in humans is unknown. The purpose of this study was to investigate this in vitro and during in vivo regeneration in humans. Following a muscle injury protocol in young healthy men (n = 7), the number of fibroblasts (TCF7L2+), satellite cells (Pax7+), differentiating myogenic cells (myogenin+) and regenerating fibres (neonatal/embryonic myosin+) was determined from biopsy cross-sections. Fibroblasts and myogenic precursor cells (MPCs) were also isolated from human skeletal muscle (n = 4) and co-cultured using different cell ratios, with the two cell populations either in direct contact with each other or separated by a permeable

Combination of Bioactive Polymeric Membranes and Stem Cells for Periodontal Regeneration: In Vitro and In Vivo Analyses.

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Gonçalves, Flávia; de Moraes, Míriam Santos; Ferreira, Lorraine Braga; Carreira, Ana Cláudia Oliveira; Kossugue, Patrícia Mayumi; Boaro, Letícia Cristina Cidreira; Bentini, Ricardo; Garcia, Célia Regina da Silva; Sogayar, Mari Cleide; Arana-Chavez, Victor Elias; Catalani, Luiz Henrique

2016-01-01

Regeneration of periodontal tissues requires a concerted effort to obtain consistent and predictable results in vivo. The aim of the present study was to test a new family of bioactive polymeric membranes in combination with stem cell therapy for periodontal regeneration. In particular, the novel polyester poly(isosorbide succinate-co-L-lactide) (PisPLLA) was compared with poly(L-lactide) (PLLA). Both polymers were combined with collagen (COL), hydroxyapatite (HA) and the growth factor bone morphogenetic protein-7 (BMP7), and their osteoinductive capacity was evaluated via in vitro and in vivo experiments. Membranes composed of PLLA/COL/HA or PisPLLA/COL/HA were able to promote periodontal regeneration and new bone formation in fenestration defects in rat jaws. According to quantitative real-time polymerase chain reaction (qRT-PCR) and Alizarin Red assays, better osteoconductive capacity and increased extracellular mineralization were observed for PLLA/COL/HA, whereas better osteoinductive properties were associated with PisPLLA/COL/HA. We concluded that membranes composed of either PisPLLA/COL/HA or PLLA/COL/HA present promising results in vitro as well as in vivo and that these materials could be potentially applied in periodontal regeneration.

In vitro mutagenesis of roses

International Nuclear Information System (INIS)

Salahbiah Abdul Majid; Rusli Ibrahim

2006-01-01

In roses, numerous in vivo mutation induction experiments have been described, but only a few commercial mutants were published. The reason for this restriction may be that it sometimes takes a few years before mutants can be isolated and propagated by conventional methods. Roses mutate readily and most selected mutants concern flower colour, shape and plant type. A major problem for improvement of roses by means of mutation breeding is chimera formation, particularly when it aims to induce changes in quantitative characters. In vitro propagation could probably accelerate the isolation of periclinal chimera. Studies were conducted to investigate the potential of using gamma rays in orderto get mutations. Dormant axillary bud explants subjected to increasing doses of gamma rays showed a decrease in regeneration capacity, which was completely suppressed at 100 Gy. The lethal dose for 50 % of the regenerating explants (LD50) for both cut and miniature roses were observed between 20-40 Gy. For the main experiment, doses between 20 and 40 Gy were found to be most suitable for the induction of high mutation rate. A few new flower mutants, with new colour and shape were selected for further testing in order to produce stable mutants and this had to be micro propagated for a few generations. Thus, using axillary bud explants for the induction of mutation through in vitro shoots regeneration, several potential stable mutants of horticultural value were isolated. (Author)

Elimination of browning exudate and in vitro development of shoots in Pistacia vera L. cv. mateur and Pistacia atlantica Desf. Culture

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M. I. Trujillo

2014-01-01

Full Text Available We report diminution and/or elimination of browning exudate followed by in vitro establishment of in Pistacia vera cv. mateur and Pistacia atlantica explants. Soaking P. vera cv. mateur explants prior to culture in L-cysteine HCl for 15 min (100 µM inhibits blackening of the modified Murashige and Skoog medium - MS + 400 mg/l NH4NO3 - and of the explants; while shoot formation was increased. The browning in P. vera cv. mateur and P. atlantica explants dissolved when modified MS and Quoirin and Lepoivre - QL.4 - media were supplemented with activated charcoal (from 1 to 3 g l-1 and with 4 and 8 days of darkness. These treatments were enough to eliminate browning from the explants and to improve the shoots elongation, but symptoms of chlorosis were detected. On the other hand, AgNO3 (from 15 to 40 µ1V1 showed a very strong antibrowning effect on the medium and explants of P. atlantica. Thus shoot organogenesis was best achieved and the developing sturdy shoots had large and green leaves.

Morphogenetic Potential of Tomato (Lycopersicon esculentum cv. ‘Arka Ahuti’ to Plant Growth Regulators

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Kanakapura K. NAMITHA

2013-05-01

Full Text Available A highly reproducible in vitro regeneration method for tomato (Lycopersicon esculentum Mill. cultivar ‘Arka Ahuti’ was established by using hypocotyl, leaf and cotyledon explants from in vitro raised seedlings on Murashige and Skoog medium supplemented with different concentrations and combinations of hormones 6-Benzylamino purine (2 to 4 mg/L and Indole-3-acetic acid (0.1 to 1 mg/L. The medium supplemented with 2 mg/L 6-benzylamino purine and 0.1 mg/L indole-3-acetic acid was found to be the best for inducing direct shoot regeneration and multiple shoots per explant from hypocotyl explants. Callus induction was observed in all the explants and regeneration of shoots was also promoted by all these combinations. Shoots were transferred to the elongation medium which also induced 100% rooting. After hardening, plants were transferred to soil. Thus, a tissue culture base line was established for ‘Arka Ahuti’ cultivar of tomato for obtaining direct regeneration using hypocotyl, leaf and cotyledon as explants.

Enzyme-Linked Immunosorbent Assay Testing of Shoots Grown In Vitro and the Use of Immunocapture-Reverse Transcription-Polymerase Chain Reaction Improve the Detection of Prunus necrotic ringspot virus in Rose.

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Moury, B; Cardin, L; Onesto, J P; Candresse, T; Poupet, A

2000-05-01

We developed and evaluated two different methods to improve the detection of the most prevalent virus of rose in Europe, Prunus necrotic ring-spot virus (PNRSV). Immunocapture-reverse transcription-polymerase chain reaction was estimated to be about 100 times more sensitive than double-antibody sandwich-enzyme-linked immunosorbent assay (DAS-ELISA) and showed an equivalent specificity. Based on the observation that PNRSV multiplies actively in young growing tissues (axillary shoots and cuttings), an in vitro culture method allowing rapid (about 15 days) and homogeneous development of dormant axillary buds with high virus titers was standardized. ELISA tests of these young shoots showed, in some cases, a 10(4) to 10(5) increase in sensitivity in comparison to adjacent leaf tissues from the rose mother plants. Between 21 and 98% (depending on the season) more samples were identified as positive by using ELISA on samples from shoot tips grown in vitro rather than on leaves collected directly from the PNRSV-infected mother plants. This simple method of growing shoot tips in vitro improved the confidence in the detection of PNRSV and eliminated problems in sampling appropriate tissues.

Rapid in vitro production of cloned plants of Uraria picta (Jacq.) DC-A rare medicinal herb in long-term culture.

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Rai, Santosh Kumar; Sharma, Meena; Jain, Madhu; Awasthi, Abhishek; Purshottam, Dharmendra Kumar; Nair, Narayanan Kuttanpillai; Sharma, Ashok Kumar

2010-11-01

An efficient in vitro process for rapid production of cloned plants of Uraria picta has been developed employing nodal stem segments taken from field-grown plants. Explants showed bud-break followed by regeneration of shoots with restricted growth within 12 days on modified Murashige and Skoog's medium supplemented with 0.25 mg l(-1) each of 6-benzylaminopurine and indole-3-acetic acid and 25 mg l(-1) adenine sulfate. Normal growth of shoots with good proliferation rate was achieved by reducing the concentrations of 6-benzylaminopurine and indole-3-acetic acid to 0.1 mg l(-1) each and incorporating 0.5 mg l(-1) gibberellic acid in the medium in which, on an average, 19.6 shoots per explant were produced. Further, during successive subcultures, increased concentrations of adenine sulfate (50 mg l(-l)) and gibberellic acid (2 mg l(-l)) along with the addition of 20 mg l(-l)  DL: -tryptophan were found conducive to control the problem of necrosis of shoots. In this treatment, several "crops" of shoots were obtained from single culture by repeated subculturing of basal portion of stalk in long-term. Isolated shoots rooted 100% in 0.25 mg l(-1) indole-3-butyric acid. In vitro-raised plants after hardening in inorganic salt solution grew normally in soil and came to flowering. Genetic fidelity of in vitro-raised plants was ascertained by rapid amplified polymorphic DNA (RAPD) markers. Also, quantitative estimation of two isoflavonones in their root extracts further confirmed true-to-type nature of plantlets.

Genetic stability and phytochemical analysis of the in vitro regenerated plants of Dendrobium nobile Lindl., an endangered medicinal orchid

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Bhattacharyya, Paromik; Kumaria, Suman; Diengdoh, Reemavareen; Tandon, Pramod

2014-01-01

An efficient genetically stable regeneration protocol with increased phytochemical production has been established for Dendrobium nobile, a highly prized orchid for its economic and medicinal importance. Protocorm like bodies (PLBs) were induced from the pseudostem segments using thidiazuron (TDZ; 1.5 mg/l), by-passing the conventional auxin–cytokinin complement approach for plant regeneration. Although, PLB induction was observed at higher concentrations of TDZ, plantlet regeneration from those PLBs was affected adversely. The best rooting (5.41 roots/shoot) was achieved in MS medium with 1.5 mg/l TDZ and 0.25% activated charcoal. Plantlets were successfully transferred to a greenhouse with a survival rate of 84.3%, exhibiting normal development. Genetic stability of the regenerated plants was investigated using randomly amplified polymorphic DNA (RAPD) and start codon targeted (SCoT) polymorphism markers which detected 97% of genetic fidelity among the regenerants. The PIC values of RAPD and SCoT primers were recorded to be 0.92 and 0.76 and their Rp values ranged between 3.66 and 10, and 4 and 12 respectively. The amplification products of the regenerated plants showed similar banding patterns to that of the mother plant thus demonstrating the homogeneity of the micropropagated plants. A comparative phytochemical analysis among the mother and the micropropagated plants showed a higher yield of secondary metabolites. The regeneration protocol developed in this study provides a basis for ex-situ germplasm conservation and also harnesses the various secondary metabolite compounds of medicinal importance present in D. nobile. PMID:25606433

Influence of UV-B radiation on developmental changes, ethylene, CO2 flux and polyamines in cv. Doyenne d'Hiver pear shoots grown in vitro

International Nuclear Information System (INIS)

Predieri, S.; Krizek, D.T.; Wang, C.Y.; Mirecki, R.M.; Zimmerman, R.H.

1993-01-01

In vitro shoots of cv. Doyenne d'Hiver pear (Pyrus communis L.) were irradiated under controlled environments for 6 h per day at 5 different levels of biologically effective UV-B radiation (UV-B BE ). UV-B exposure caused a progressive increase in apical necrosis above background levels and stimulated leaf abscission. Shoots grown for 2 weeks at 7. 8 mol m −2 day −1 of photosynthetic photon flux (PPF) and treated with 8. 4 or 12. 0 kJ m −2 day −1 UV-B BE produced up to 4 times more ethylene than those given 2. 2 or 5. 1 kJ m −2 day −1 UV-B BE or untreated controls. Exposure of shoots to 12 kJ m −2 day −1 of UV-BBE caused an increase in free putreseine content after 4 to 14 days of irradiation. Shoots showed a decrease in CO2 uptake after 3 days of UV-B: thereafter, they appeared to recover their photosynthetic capacity. Under typical PPF conditions used in micropropagation (90 μmol m −2 s −1 ). 8. 4 kJ m −2 day −1 of UV-B radiation was injurious to realatively tender tissues of in vitro pear shoots: increasing the level of UV-B BE to 12 kJ m −2 day −1 produced even more adverse effects. (author)

''In vitro'' mutation breeding methodology for Fusarium wilt resistance in banana

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Tulmann Neto, A; Domingues, E T; Mendes, B M.J.; Ando, A [Centre for Nuclear Energy in Agriculture (CENA), Piracicaba, SP. (Brazil)

1990-07-01

Full text: Besides ''in vivo'' methods, the Radiation Genetics Section of CENA/USP is also using ''in vitro'' methods for mutation breeding to obtain resistance to Panama disease caused by Fusarium oxysporum f.sp. cubense in the banana cultivar ''Maca''. A protocol has been established for the ''in vitro'' development of shoot tips, obtained from plants in the field or already cultivated under {sup i}n vitro'' conditions. For both cases, only one culture medium was used during all steps of ''in vitro'' cultivation. New buds were formed and these buds grew and developed to form roots. The medium was composed of macro and micro nutrients, with added Morel vitamines, BAP (5 mg/l), saccharose (30 g/l) and agar (6.5 g/l), at pH 5.7. Cultures were allowed to grow in a controlled environment at 27 deg. C and 16 h illumination. Shoot tips which originated from ''in vitro'' plantlets, were cut longitudinally down the middle. This was done to avoid a tendency of regeneration of the original tissue instead of the formation of new lateral buds. To resolve the chimerism resulting when mutagenic treatment is applied to shoot tips, there is a need of vegetative propagation of new lateral buds. Selection can then be done at M{sub 1}V{sub 4} generation. Once the protocol was established, the gamma ray sensitivity was determined. The dose that produced a 50% decrease in the number of new lateral buds was around 40 Gy and this dose will be utilised. The methodology was completed by soil inoculation with Fusarium of young plants 15 cm in height, obtained from ''in vitro'' cultures. After 3 weeks all inoculated plants showed symptoms of wilt, demonstrating the possibility of screening. The method is now being utilised on a large scale in an attempt to induce a resistant mutant. (author)

''In vitro'' mutation breeding methodology for Fusarium wilt resistance in banana

International Nuclear Information System (INIS)

Tulmann Neto, A.; Domingues, E.T.; Mendes, B.M.J.; Ando, A.

1990-01-01

Full text: Besides ''in vivo'' methods, the Radiation Genetics Section of CENA/USP is also using ''in vitro'' methods for mutation breeding to obtain resistance to Panama disease caused by Fusarium oxysporum f.sp. cubense in the banana cultivar ''Maca''. A protocol has been established for the ''in vitro'' development of shoot tips, obtained from plants in the field or already cultivated under i n vitro'' conditions. For both cases, only one culture medium was used during all steps of ''in vitro'' cultivation. New buds were formed and these buds grew and developed to form roots. The medium was composed of macro and micro nutrients, with added Morel vitamines, BAP (5 mg/l), saccharose (30 g/l) and agar (6.5 g/l), at pH 5.7. Cultures were allowed to grow in a controlled environment at 27 deg. C and 16 h illumination. Shoot tips which originated from ''in vitro'' plantlets, were cut longitudinally down the middle. This was done to avoid a tendency of regeneration of the original tissue instead of the formation of new lateral buds. To resolve the chimerism resulting when mutagenic treatment is applied to shoot tips, there is a need of vegetative propagation of new lateral buds. Selection can then be done at M 1 V 4 generation. Once the protocol was established, the gamma ray sensitivity was determined. The dose that produced a 50% decrease in the number of new lateral buds was around 40 Gy and this dose will be utilised. The methodology was completed by soil inoculation with Fusarium of young plants 15 cm in height, obtained from ''in vitro'' cultures. After 3 weeks all inoculated plants showed symptoms of wilt, demonstrating the possibility of screening. The method is now being utilised on a large scale in an attempt to induce a resistant mutant. (author)

Insulin and IGF-II, but not IGF-I, stimulate the in vitro regeneration of adult frog sciatic sensory axons

DEFF Research Database (Denmark)

Edbladh, M; Svenningsen, Åsa Fex; Ekström, P A

1994-01-01

We used the in vitro regenerating frog sciatic nerve to look for effects of insulin and insulin-like growth factors I and II (IGF-I, IGF-II) on regeneration of sensory axons and on injury induced support cell proliferation in the outgrowth region. In nerves cultured for 11 days, a physiological...

Transformation and regeneration of the holoparasitic plant Phelipanche aegyptiaca

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Fernández-Aparicio Mónica

2011-11-01

Full Text Available Abstract Background Transformation and subsequent regeneration of holoparasitic plants has never been reported, in part due to challenges in developing transformation protocols, but also because regeneration of obligate parasites is difficult since their survival depends completely on successful haustorium penetration of a host and the formation of vascular connections. The recent completion of a massive transcriptome sequencing project (the Parasitic Plant Genome Project will fuel the use of genomic tools for studies on parasitic plants. A reliable system for holoparasite transformation is needed to realize the full value of this resource for reverse genetics and functional genomics studies. Results Here we demonstrate that transformation of Phelipanche aegyptiaca is achieved by infection of 3 month-old in vitro grown P. aegyptiaca calli with Agrobacterium rhizogenes harboring the yellow fluorescent protein (YFP. Four months later, YFP-positive regenerated calli were inoculated onto tomato plants growing in a minirhizotron system. Eight days after inoculation, transgenic parasite tissue formed lateral haustoria that penetrated the host and could be visualized under UV illumination through intact host root tissue. YFP-positive shoot buds were observed one month after inoculation. Conclusions This work constitutes a breakthrough in holoparasitic plant research methods. The method described here is a robust system for transformation and regeneration of a holoparasitic plant and will facilitate research on unique parasitic plant capabilities such as host plant recognition, haustorial formation, penetration and vascular connection.

Micropropagação de cebola a partir de bulbinhos induzidos in vitro Micropropagation of onion from in vitro induced bulblet

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Benedita Maria Rodrigues

1996-01-01

Full Text Available Visando determinar um método para micropropagação a partir de bulbinhos in vitro, de cultivares de cebola (Allium cepa L., foram realizados três experimentos, avaliando-se a influência do 6-benzilaminopurina (BAP x ácido naftalenoacético (ANA e posição do explante no meio de cultura, a influência de bulbinhos desenvolvidos em diversas concentrações de sacarose e o efeito do diâmetro dos bulbinhos sobre a taxa de multiplicação in vitro. Pelos resultados, a posição vertical dos explantes mostrou maior tendência para porcentagem de regeneração nos cultivares Pira Ouro e Pirana Precoce. 0 efeito das concentrações de fitorreguladores na porcentagem de regeneração não foi verificado para o 'Pira Ouro', enquanto para o 'Pirana Precoce', porcentagem mais elevada foi obtida em 4,0 mg/L de BAP x 0,5 mg/L de ANA. Maior taxa de multiplicação foi observada para os dois cultivares em BAP (2,0 mg/L x ANA (0,25 mg/L. Bulbinhos do cultivar Pira Ouro e híbrido entre Pira Ouro e Pirana Precoce, desenvolvidos em 120 g/L de sacarose, mostraram-se mais adequados à micropropagação in vitro no material utilizado. Com relação ao diâmetro inicial do explante, o 'Pira Ouro' não mostrou diferença estatística para os parâmetros avaliados, enquanto no 'Pirana Precoce' maior porcentagem de regeneração foi obtida nos diâmetros M (5 a 10 mm e P (Series of three different experiments were carried out, in order to study the optimization of the use of bulblets obtained in vitro, in the micropropagation of the onion (Allium cepa L.. Initially, it was evaluated the effects of the interaction between two doais of 6-benzylaminopurine (BAP and naphthaleneacetic acid (NAA, and the position of the explant in the medium, for the cultivara Pira Ouro and Pirana Precoce. For 'Pira Ouro' the phytohormone concentrations did not affect the percentage of regeneration; however, for 'Pirana Precoce', the rates of 4.0 mg/L of BAP and 0.5 mg/L of NAA gave the

In-vitro induction of mutations in Chrysanthemum using x- and gamma-radiation

International Nuclear Information System (INIS)

Jerzy, M.

1990-01-01

Full text: Spontaneous and induced mutations occur frequently in the hexaploid and very heterozygous Chrysanthemum. Alterations in flower colours can thus be obtained for an outstanding cultivar produced by cross-breeding. Whole families of mutants have been reported for cultivars like 'Westland', 'Indianapolis', 'Horim' and 'Bravo'. The mutation-breeding programme in Poland started in 1986 with the use of in vitro techniques to obtain early cultivars flowering in unheated plastic greenhouses. One of the successful examples was the irradiation of cv. 'Richmond'. Leaves from plants regenerated through shoot tip culture were treated with 15 Gy x- and gamma-rays. Each treatment comprised 400 leaf explants on MS medium with IAA 2 mg/l and BA 0,6 mg/l. After 6 weeks, regenerated adventitious shoots were transferred to the rooting medium with HAA 0,02 mg/l. Rooted plantlets were used as stock plants for the production of shoot cuttings to be subjected to mutant selection. Many plants with drastically changed flower colours were obtained. The violet-pink colour, which is typical for 'Richmond' flowers changed to 13 different colours: white, yellow, golden, pink, salmon, lilac, violet, purple, orange, apricot, amber, bronze and red. Eleven colours occurred as a result of x-ray treatment, eight colours after gamma irradiation. Chimeras were not observed. Among irradiated plants there were also inflorescences with all-ligulate florets scrolled up into tubes. (author)

Phytochemical investigation of natural and in vitro raised Vṛddhadāruka plants.

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Bharati, Asha Jyoti; Bansal, Yogendra Kumar

2014-01-01

Argyreia nervosa commonly known as elephant creeper (English) and Vṛddhadāruka (Sanskrit) is a woody climber that belongs to the family Convolvulaceae. Seeds of this plant contain hallucinogens including ergot alkaloids and a naturally occurring lysergic acid amide. Traditionally the plant is used in the treatment of gonorrhea, strangury, chronic ulcers, diabetes, anemia and cerebral disorders. The plant is also used as appetitiser, brain tonic, cardiotonic, aphrodisiac. It possesses anti-inflammatory, immunomodulatory, antibacterial, antiviral and antifungal activities. To give an account of information on in vitro regeneration and phytochemical analysis of the plant. Nodal explants were selected for in vitro regeneration. Different aerial parts viz., seeds, natural and in vitro leaf, stem and callus were dried and extracted with different solvents and were subjected to various phytochemical analyses. Different concentrations of 6-benzylaminopurine showed shoot and root initiation. The study of phytochemical screening of different extracts showed the presence of bioactive substances like flavonoids, alkaloids, terpenoids, etc. The study will provide an efficient in vitro protocol for micropropagation as an alternative method to conserve the plant and shows the presence of some important secondary metabolites in the nature grown and in vitro raised plants which can be useful for treatment of various diseases.

Phytochemical investigation of natural and in vitro raised Vṛddhadāruka plants

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Asha Jyoti Bharati

2014-01-01

Full Text Available Background: Argyreia nervosa commonly known as elephant creeper (English and Vṛddhadāruka (Sanskrit is a woody climber that belongs to the family Convolvulaceae. Seeds of this plant contain hallucinogens including ergot alkaloids and a naturally occurring lysergic acid amide. Traditionally the plant is used in the treatment of gonorrhea, strangury, chronic ulcers, diabetes, anemia and cerebral disorders. The plant is also used as appetitiser, brain tonic, cardiotonic, aphrodisiac. It possesses anti-inflammatory, immunomodulatory, antibacterial, antiviral and antifungal activities. Objective: To give an account of information on in vitro regeneration and phytochemical analysis of the plant. Materials and Methods: Nodal explants were selected for in vitro regeneration. Different aerial parts viz., seeds, natural and in vitro leaf, stem and callus were dried and extracted with different solvents and were subjected to various phytochemical analyses. Results: Different concentrations of 6-benzylaminopurine showed shoot and root initiation. The study of phytochemical screening of different extracts showed the presence of bioactive substances like flavonoids, alkaloids, terpenoids, etc. Conclusion: The study will provide an efficient in vitro protocol for micropropagation as an alternative method to conserve the plant and shows the presence of some important secondary metabolites in the nature grown and in vitro raised plants which can be useful for treatment of various diseases.

IN VITRO SHOOT REGENERATION OF MOMORDICA BALSAMINA ...

African Journals Online (AJOL)

user

2011-11-09

Nov 9, 2011 ... good opportunity to use tissue culture as a complementary tool for breeding and genetics for other applications. ... contributes towards combating the problem of ... was maintained by covering with white plastic bags. After 15 ...

IN VITRO SHOOT REGENERATION OF MOMORDICA BALSAMINA ...

African Journals Online (AJOL)

user

2011-11-09

Nov 9, 2011 ... nutritional plant. Gulab Singh Thakur1,4*, Rohit Sharma1, Bhagwan S. Sanodiya1,4, Mukeshwar Pandey3,. Rakesh Baghel2, Astha Gupta1, GBKS Prasad4 and P. S. Bisen1,2,4. 1Plant Biotechnology Laboratory, R&D Division, Tropilite Foods Pvt. Ltd., Davar Campus, Tansen Road, Gwalior-474002. (M.P.) ...

Influence of UV-B radiation on developmental changes, ethylene, CO[sub 2] flux and polyamines in cv. Doyenne d'Hiver pear shoots grown in vitro

Energy Technology Data Exchange (ETDEWEB)

Predieri, S. (Consiglio Nazionale delle Ricerche, Centro Studi di Technica Frutticola, Bologna (Italy)); Krizek, D.T. (Climate Stress Laboratory, USDA, Beltsville, MD (United States)); Chien, Y. Wang (Horticultural Crops quality Laboratory, USDA, Beltsville, MD (United States)); Mirecki, R.M. (Climate Stress Laboratory, USDA, Beltsville, MD (United States)); Zimmerman, R.H. (Fruit Laboratory, USDA, Beltsville, MD (United States))

1993-01-01

In vitro shoots of cv. Doyenne d'Hiver pear (Pyrus communis L.) were irradiated under controlled environments for 6 h per day at 5 different levels of biologically effective UV-B radiation (UV-B[sub BE]). UV-B exposure caused a progressive increase in apical necrosis above background levels and stimulated leaf abscission. Shoots grown for 2 weeks at 7.8 mol m[sup -2] day[sup -1] of photosynthetic photon flux (PPF) and treated with 8.4 or 12.0 kJ m[sup -1] day[sup -1] UV-B[sub BE] produced up to 4 times more ethylene than those given 2.2 or 5.1 kJ m[sup -2] day[sup -1] UV-B[sub BE] or untreated controls. Exposure of shoots to 12 kJ m[sup -2] day[sup -1] of UV-B[sub BE] caused an increase in free putrescine content after 4 to 14 days of irradiation. Shoots showed a decrease in CO[sub 2] uptake after 3 days of UV-B; thereafter, they appeared to recover their photosynthetic capacity. Under typical PPF conditions used in micropropagation (90 [mu]mol m[sup -2] s[sup -1]), 8.4 kJ m[sup -2] day[sup -1] of UV-B radiation was injurious to relatively tender tissues of in vitro pear shoots; increasing the level of UV-B[sub BE] to 12 kJ m[sup -2] day[sup -1] produced even more adverse effects. (au).

In Vitro Propagation, Phytochemical Analysis, and Evaluation of Free Radical Scavenging Property of Scrophularia kakudensis Franch Tissue Extracts.

Science.gov (United States)

Manivannan, Abinaya; Soundararajan, Prabhakaran; Park, Yoo Gyeong; Jeong, Byoung Ryong

2015-01-01

The current study deals with in vitro propagation, antioxidant property estimation, and assessment of acacetin content in Scrophularia kakudensis Franch. Adventitious shoot induction was achieved from the nodal explant with the highest number of adventitious shoots per explant (17.4) on Murashige and Skoog's (MS) medium fortified with 2.0 mg·L(-1) 6-benzyladenine (BA) and 0.5 mg L(-1) indole-3-acetic acid (IAA). Maximum number of roots per plant (16.5) was noted in half strength MS medium supplemented with 0.5 mg·L(-1) IAA. The regenerated plants displayed successful survival ratio (95%) in the greenhouse. The highest content of acacetin, a pharmaceutically important flavonoid, was observed in the shoot extracts (in vitro: 32.83 µg·g(-1) FW; in vivo: 30.05 µg·g(-1) FW) followed by root extracts. Total phenol and flavonoid contents along with free radical scavenging assays revealed the occurrence of larger amount of antioxidants in shoot extract in comparison with callus and root extracts of S. kakudensis. Thus, the outcome of the present study can be highly beneficial for the germplasm conservation and commercial cultivation of S. kakudensis for therapeutic purposes.

TDZ-Induced Plant Regeneration in Jatropha curcas: A Promising Biofuel Plant

KAUST Repository

Kumar, Nitish

2018-03-23

In recent years, Jatropha curcas has pronounced attention due to its capacity of production of biodiesel. Uniform large-scale propagation of J. curcas is one of the significant keys that will eventually decide victory. Direct regeneration is one of the methods which help in the production of uniform and homogenous plant, and TDZ plays an important role in the production of plantlets by direct organogenesis in several number of plant species including J. curcas. Measuring the economical importance of J. curcas and the role of TDZ in shoot regeneration, the present book chapter briefly reviews the impact of TDZ on shoot bud induction from various explants of J. curcas.

TDZ-Induced Plant Regeneration in Jatropha curcas: A Promising Biofuel Plant

KAUST Repository

Kumar, Nitish; Bhatt, Vacha D.; Mastan, Shaik G.; Reddy, Muppala P.

2018-01-01

In recent years, Jatropha curcas has pronounced attention due to its capacity of production of biodiesel. Uniform large-scale propagation of J. curcas is one of the significant keys that will eventually decide victory. Direct regeneration is one of the methods which help in the production of uniform and homogenous plant, and TDZ plays an important role in the production of plantlets by direct organogenesis in several number of plant species including J. curcas. Measuring the economical importance of J. curcas and the role of TDZ in shoot regeneration, the present book chapter briefly reviews the impact of TDZ on shoot bud induction from various explants of J. curcas.

Differentiation of shoot elements from the rachis of Secale cereale L.

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Jan J. Rybczyński

2014-01-01

Full Text Available In vitro culture of young Secale cereale spikes is described with special attention given to changes in development of the rachis. 7 mm explants were cultivated on a modified M u r a s h i g e and S k o o g (1962 medium (MS supplemented with 2,4-D (dichlorophenoxyacetic acid, NAA (α-naph-thaleneacetic acid, IAA (β-indoleacetic acid, 2,4,5-T (trichlorophenoxyacetic acid, and ZEA (zeatin, KIN (kinetin, BAP (6-benzylaminopurine, IPA (izopentenyladenine or 2 iP in numerous combinations and concentrations. Rachises differentiated branches with node construction in the presence of synthetic auxin. Rhizogenesis of explants was stimulated by 2,4-D and 2,4,5-T. 2,4,5-T + IPA appeared to be the best combination for callus regeneration. Many meristematic centres were found previously and combinations of NAA + KIN stimulated direct, whereas 2,4-D + ZEA indirect, differentiation of shoot elements. Development was observed from plantlets to flowering plants.

Fox grape cv. Bordô (Vitis labrusca L. and grapevine cv. Chardonnay (Vitis vinifera L. cultivated in vitro under different carbohydrates, amino acids and 6-Benzylaminopurine levels

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Dayse Cristina de Carvalho

2013-04-01

Full Text Available The aim of this work was to study the influence of sucrose and glucose, amino acids and BAP (6-Benzylaminopurine levels on in vitro shoot regeneration of fox grape cv. Bordô and grapevine cv. Chardonnay. The nodal segments from micropropagated material were used as explants and half-strength MS medium as the basal medium. Sucrose and glucose at 15, 30 and 45 g.L-1 were tested as a carbon source and the supplementation of adenine, asparagine, alanine, glycine, cysteine, glutamine, arginine was tested at 40 g.L-1. The BAP levels (1 and 5 μM in solid and double-phase media were evaluated and compared with a control medium without BAP. Bordô had best in vitro growth than Chardonnay. Sucrose was a better carbohydrate source than glucose for both the cultivars. Bordô and Chardonnay had different amino acid preferences for some parameters. In conclusion, for in vitro shoot regeneration from the nodal segments, culture on solid medium with 5 μM BAP, 15 g.L-1 sucrose for Bordô and 45 g.L-1 sucrose for Chardonnay showed better results. Similarly, the supplementation of 40 g.L-1 arginine for Bordô and 40 g.L-1 arginine or glycine for Chardonnay showed better results.

Effect of antioxidants and associate changes in antioxidant enzymes in controlling browning and necrosis of proliferating shoots of elite Jatropha curcas L

International Nuclear Information System (INIS)

Misra, Pratibha; Toppo, Dibya D.; Gupta, Neha; Chakrabarty, Debasis; Tuli, Rakesh

2010-01-01

A high yielding elite plant of Jatropha curcas was established under aseptic condition from field-grown nodal explants. Shoots were proliferated in MS medium supplemented with 0.5 mg dm -3 benzyladenine and 0.1 mg dm -3 indolebutyric acid along with 10 mg dm -3 adenine sulphate and a combination of 15 mg dm -3 each of L-glutamine and L-arginine. However, within 15-20 d of culture incubation, tissue browning/necrosis leading to poor plant regeneration in vitro was observed. A set of different antioxidants, namely, reduced glutathione, ascorbic acid, tocopherol and cysteine were used in the medium individually and in combination to solve the problem of tissue browning and necrosis. The addition of antioxidants proved beneficial for the growth of the shoots. The optimum medium comprised of 25 mg dm -3 reduced glutathione and 10 mg dm -3 ascorbic acid, where proliferating shoots having highest leaf canopy area, remained fresh, green and regenerative up to 40 d of culture incubation without any subculture. The activities of antioxidant enzymes, such as superoxide dismutase was higher in control shoots, indicating that tissue browning/necrosis was associated with oxidative stress which was further supported by higher contents of H 2 O 2 and phenolics in control shoots compared to the other treatments. Similarly glutathione reductase, ascorbate peroxidase and guiacol peroxidase was higher in treated shoots than control indicating that these shoots have developed antioxidant enzymatic protective system which determine the ability to survive in oxidative stress and up regulation of these enzymes would help to reduce the built up of reactive oxygen species.

Radiation and chemical mutagen induced somaclonal variations through in vitro organogenesis of cotton (Gossypium hirsutum L.).

Science.gov (United States)

Muthusamy, Annamalai; Jayabalan, Narayanasamy

2014-12-01

The purpose of the investigation was to induce somaclonal variations by gamma rays (GR), ethylmethane sulphonate (EMS) and sodium azide (SA) during in vitro organogenesis of cotton. The shoot tip explants were irradiated with 5-50 Gray (Gy) GR (Cobalt 60), 0.5-5.0 mM EMS and SA separately, and inoculated on Murashige and Skoog (MS) medium fortified with plant growth regulator (PGR) for organogenesis. The plantlets with well-developed root systems were acclimatized and transferred into the experimental field to screen the somaclonal variations during growth and development. The number of somaclonal variations was observed in growth of irradiated/treated shoot tips, multiplication, plantlet regeneration and growth in vitro and ex vitro. The lower doses/concentrations of mutagenic treatments showed significant enhancement in selected agronomical characters and they showed decreased trends with increasing doses/concentrations of mutagenic agents. The results of the present study revealed the influence of lower doses/concentrations of mutagenic treatments on in vitro and ex vitro growth of cotton plantlets and their significant improvement in agronomical characters which needs further imperative stability analysis. The present observations showed the platform to use lower doses/concentrations of mutagenic agents to induce variability for enhanced agronomical characters, resistant and tolerant cotton varieties.

Studies on the reaction in tissue culture of tomato genotypes under biotic stress

Directory of Open Access Journals (Sweden)

Ewa Hanus-Fajerska

2014-01-01

Full Text Available Plant regeneration in vitro from virus-infected somatic tomato (Lycopersicon sp. tissue was performed. Regeneration experiments were started after the determination of virus presence, using enzyme-linked immunosorbent assay, in leaves used as a source of explants. Leaf explants infected with selected strains of tomato mosaic Tobamovirus or cucumber mosaic Cucumovirus respectively, were cultured on a standarised MS agar medium to induce adventitious shoots, which were afterwards excised, rooted in vitro and cultured to plants. Explants were also screened for their ability to produce callus. Diverse effects of viral infection, ranging from stimulation to inhibition of callus formation and of morphogenesis rate, were observed. The health condition of the tissue proved to affect regeneration potential of Lycopersicon esculentum, whereas wild accesions did not react in that case so distinctly. In cultivated tomato was encountered the decline in competence to reproduce shoots adventitiously in infected tissue. There was also relationship between donor plant health condition and adventitious root formation in regenerated shoots. Experiments with short-term cultures of L. esculenum reveled also that a certain number of shoots regenerated from diseased tissue can be virus-free.

In Vitro Propagation and Conservation of Withania somnifera (Dunal) L.

Science.gov (United States)

Fatima, Nigar; Ahmad, Naseem; Anis, Mohammad

2016-01-01

Plant tissue culture offers several techniques for rapid clonal propagation, germplasm conservation, regeneration of genetically manipulated superior clones, production of phyto-constituents, and ex vitro conservation of valuable phytodiversity. An improved and efficient micropropagation protocol for Withania somnifera (L.), a drug-producing medicinal plant, using juvenile explants (nodal explants) has been developed. Highest multiplication and subsequent elongation of shoots is observed on MS medium containing BA and NAA. The regenerated microshoots roots best on ½ MS medium containing NAA, established in earthen pots containing garden soil and are maintained in the greenhouse with 95 % survival rate. Genetic uniformity of micropropagated plants is confirmed by PCR-based DNA fingerprinting techniques, viz., RAPD and ISSR. No variation is observed in DNA fingerprinting patterns among the micropropagated plants, which are similar to that of the donor plant illustrating their genetic uniformity.

In vitro rescue of interspecific embryos from Elaeis guineensis x E. oleifera (Arecaceae).

Science.gov (United States)

Angelo, Paula Cristina da Silva; Moraes, Larissa Alexandra Cardoso; Lopes, Ricardo; Sousa, Nelcimar Reis; da Cunha, Raimundo Nonato Vieira; Quisen, Regina Caetano

2011-09-01

The African oil palm (Elaeis guineensis) is the most effective oil producer in tons per hectare. Nevertheless, its increasing cultivation in Latin America is harmed by the "lethal yellowing". Genetic resistance to this anomaly can be found in the germplasm of American oil palm or caiaué (E. oleifera), a native species from the Amazon rainforest. However, the procedures adopted to induce seeds of E. guineensis to germination frequently result mild for interespecific hybrids. Embryo in vitro cultivation can be a viable option. This work was aimed initially to test liquid MS medium supplemented with different glucose or sucrose concentrations for the in vitro cultivation of zygotic embryos from E. guineensis x E. oleifera controlled pollinations. Additionally we investigated different compost mixtures to acclimatize the regenerated hybrid plantlets. Concentrations of 10, 20 and 30g/L of both sugars were tested on flasks containing five mature zygotic embryos, with 15 repetitions per treatment in a total of 450 explants. The number of embryos displaying shoots and radicles at least 2mm in length per experimental unit was evaluated during phase one of in vitro cultivation. Plantlets displaying shoots and radicles were transferred to phase two of in vitro cultivation and subsequently to acclimatization, under 70% shading with manual water supply. The experiments of acclimatization were conducted with 130 plantlets randomly distributed in pure horticultural compost, 3:1 or 1:1 compost:sand mixtures and each plantlet was defined as an experimental unit. Data were submitted to ANOVA, t test and analyzes of correlation (p < or = 0.05). Highest emergence rates were 97% for shoots and 73% for radicles, observed in MS medium supplemented with 20g/L (110mM) of glucose. This sugar in concentrations of 20 or 30g/L provided balanced shoot/root development, and this was considered one of the reasons for the higher frequency of plantlet establishment. The survival percentage was 55

Effect of 60Co γ-rays irradiation on plant regeneration from callus of sea dallisgrass (paspalum vaginatum Sw.)

International Nuclear Information System (INIS)

Ye Xiaoqing; She Jianming; Wang Songfeng; Zhang Xu; Liang Liufang; Dong Mingqiang; Wu Yingying

2010-01-01

The pellet embryonic calli of Paspalum vagiantum Sw. cv. Adalay were used for 60 Co γ-rays irradiation. In the callus subculture medium with 2, 4-D 2.0 mg /L and BAP 0.05 mg /L, calli were irradiated at the dose rate 1 Gy /min of 60 Co γ-rays. Results showed that the rate of shoot regeneration was from 94% to 85% when calli irradiated with 20 ∼ 50 Gy; the rate of shoot regeneration was from 76% to 30%, and the rate of plant regeneration was from 44.5% to 8.7% between 60 ∼ 80 Gy. By 60 Gy of 60 Co γ-rays irradiation, the rate of shoot regeneration was about 76% in the differentiation medium with BAP 2.0 mg /L, and the relative rate of plant regeneration was 44.5% in the rooting medium with NAA 0.5 mg /L. The result of sequence-related amplified polymorphism (SRAP) markers analysis showed that the specific SRAP markers were associated with the somatic mutants. The mutation technique of 60 Co γ-rays has been established in the somatic cell of Paspalum vaginatum Sw. (authors)

Thidiazuron enhanced regeneration and silymarin content in silybum marianum

International Nuclear Information System (INIS)

Khan, M.A.; Abbasi, B.H.; Shinwari, Z.K.

2014-01-01

Silybum marianum, of family Asteraceae is renowned for production of biologically important silymarin, which has shown multi-dimensional medicinal properties. It has a high protective role against jaundice and hepatitis C worldwide. We hereby established a feasible and efficient method for indirect regeneration of S. marianum for production of consistent plantlets. Calli were induced from leaf explants of seed-derived plantlets on Murashige and Skoog (MS) medium supplemented with several concentrations of different plant growth regulators (PGRs). Highest callogenic response (89%) was recorded for 4.4 meu M Thidiazuron (TDZ) in combination with 6.6 meu M Kinetin (Kn). Subsequent sub-culturing of callus after 4 weeks of culture, on medium with similar compositions of PGRs induced shoot organogenesis. Highest shoot induction frequency (86%) with maximum mean multiple shoots (26 shoots per explant) were recorded for 11meu M TDZ after 4 weeks of transfer. Longest shoots (4.1 cm) were recorded for MS medium augmented with 6.6 meu M TDZ and 4.4 meu M naphthalene acetic acid (NAA). Furthermore, rooted plantlets were developed on MS medium containing different concentrations of indole acetic acid (IAA). Silymarin was determined by High performance liquid chromatography (HPLC) and 8.47 mg/g DW silymarin was detected in the regenerated plantlets. This study contributes to a better understanding of the different mechanisms involved in morphogenesis and production of biologically active principle in Silybum marianum. (author)

In Vitro Organogenesis of Two Sansevieria Trifasciata Cultivars on Different Concentrations of Benzyladenine (BA)

OpenAIRE

Yusnita, Yusnita; Pungkastiani, Wiry; Hapsoro, Dwi

2011-01-01

In vitro regeneration of buds and shoots via organogenesis in two genotypes of snake plants (Sansevieria trifasciata) were established. Leaf segments (1 x 1cm) of 'Hahnii' and 'Lorentii' were cultured on Murashige and Skoog (MS) basal medium containing 2,4- dichlorophenoxyacetic acid (2,4-D) for 1 week, transferred into MS medium without plant growth regulator (MS0) for 1 week, and then cultured on MS medium containing different concentrations of N6-benzyladenine (BA) at 0, 0.1, 0.25, 0.5, 1....

Callus induction and plant regeneration of Ulex europaeus

OpenAIRE

Ramírez,Ingrid; Dorta,Fernando; Cuadros-Inostroza,Álvaro; Peña-Cortés,Hugo

2012-01-01

A callus induction and plant regeneration protocol was developed from leaf and thorn explants for the plant Ulex europaeus. Explants were incubated on 2% sucrose half-strength Murashige and Skoog Medium (MS) with various combinations of plant growth regulators and antioxidants. The best frequency of callus and shoot formation was obtained with 2,4-dichlorophenoxyacetic acid (2,4-D) 1 mg/l x kinetin (Kin) 0.2 mg/l (DK Medium; callus induction) and zeatin (Z) 1 mg/l (DK medium; shoot induction)...

Induced mutation in rice by in-vitro radiation

International Nuclear Information System (INIS)

Qi Xiufang; Zhao Chengzhang; Zhen Kangle

1989-01-01

The experiment indicated the adequate dose and growth stage for in-vitro radiation treatment. The results were: 1) Effect of 137 Cs-γ rays on redifferentiation and rate of green plantlets of somatic cells at different growth stage is different. The sequence of effect is callus>callus with green spots>callus with green shoot>mature embryo of dry seed. 2) The fertility of regenerated rice plants decreased with the radiation dose from 0 to 2.58 C/kg, and the rate of exserted stigma increased. 3) The occurence with early maturity increased up to 3.8%, among with about 0.5% of the plant lines matured 15 days earlier. 4) Tow male sterile lines with extruded stigmas were obtained, whose plant heights were reduced. Its frequency of occurance by in-vitro radiation is more than by coventional radiation and somatic cell culture

In Vitro Analysis of the Role of Schwann Cells on Axonal Degeneration and Regeneration Using Sensory Neurons from Dorsal Root Ganglia.

Science.gov (United States)

López-Leal, Rodrigo; Diaz, Paula; Court, Felipe A

2018-01-01

Sensory neurons from dorsal root ganglion efficiently regenerate after peripheral nerve injuries. These neurons are widely used as a model system to study degenerative mechanisms of the soma and axons, as well as regenerative axonal growth in the peripheral nervous system. This chapter describes techniques associated to the study of axonal degeneration and regeneration using explant cultures of dorsal root ganglion sensory neurons in vitro in the presence or absence of Schwann cells. Schwann cells are extremely important due to their involvement in tissue clearance during axonal degeneration as well as their known pro-regenerative effect during regeneration in the peripheral nervous system. We describe methods to induce and study axonal degeneration triggered by axotomy (mechanical separation of the axon from its soma) and treatment with vinblastine (which blocks axonal transport), which constitute clinically relevant mechanical and toxic models of axonal degeneration. In addition, we describe three different methods to evaluate axonal regeneration using quantitative methods. These protocols constitute a valuable tool to analyze in vitro mechanisms associated to axonal degeneration and regeneration of sensory neurons and the role of Schwann cells in these processes.

Molecular and biochemical characterization in Rauvolfia tetraphylla plantlets grown from synthetic seeds following in vitro cold storage.

Science.gov (United States)

Faisal, Mohammad; Alatar, Abdularhaman A; Hegazy, Ahmad K

2013-01-01

Synseed technology is one of the most important applications of plant biotechnology for in vitro conservation and regeneration of medicinal and aromatic plants. In the present investigation, synseeds of Rauvolfia tetraphylla were produced using in vitro-proliferated shoots upon complexation of 3 % sodium alginate and 100 mM CaCl(2). The encapsulated buds were stored at 4, 8, 12, and 16 °C and high conversion was observed in synseeds stored at 4 °C for 4 weeks. The effect of different medium strength on in vitro conversion response of synseed was evaluated and the maximum conversion (80.6 %) into plantlets was recorded on half-strength woody plant medium supplemented with 7.5 μM 6-benzyladenine and 2.5 μM α-naphthalene acetic acid after 8 weeks of culture. Plantlets with well-developed shoot and roots were hardened and successfully transplanted in field condition. After 4 weeks of transfer to ex vitro conditions, the performance of synseed-derived plantlets was evaluated on the basis of some physiological and biochemical parameters and compared with the in vivo-grown plants. Short-term storage of synthetic seeds at low temperature had no negative impact on physiological and biochemical profile of the plants that survived the storage process. Furthermore, clonal fidelity of synseed-derived plantlets was also assessed and compared with mother plant using rapid amplified polymorphic DNA and inter-simple sequence repeats analysis. No changes in molecular profiles were found among the regenerated plantlets and comparable to mother plant, which confirm the genetic stability among clones. This synseed protocol could be useful for in vitro clonal multiplication, conservation, and short-term storage and exchange of germplasm of this antihypertensive drug-producing plant.

Effect of different intervals of x-ray split doses on shoot production of in vitro derived explants of Gerbera jamesonii Bolus

Energy Technology Data Exchange (ETDEWEB)

Walther, F; Sauer, A [Federal Research Centre for Horticultural Plant Breeding, Ahrensburg (Germany)

1990-01-01

Full text: Linearity between rising x-ray doses and mutation rate is limited by the simultaneously increasing radiation damage; induced chromosome aberrations eliminate valuable factor mutations. The application of fractionated doses provides the opportunity for repair of a distinct portion of damage. The dose of 30 Gy was fractionated into two identical parts. The periods for repair were 0.5 to 48 hs. The absolute and cumulative number of post-irradiation regenerated axillary shoots on 4 subsequent dates of cutoff were used as parameters to estimate radiosensitivity. From an economical point of view the interval of 4 hs between two dose fractions may be recommended for practice. (author)

Encapsulation of nodal segments of lobelia chinensis

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Weng Hing Thong

2015-04-01

Full Text Available Lobelia chinensis served as an important herb in traditional chinese medicine. It is rare in the field and infected by some pathogens. Therefore, encapsulation of axillary buds has been developed for in vitro propagation of L. chinensis. Nodal explants of L. chinensis were used as inclusion materials for encapsulation. Various combinations of calcium chloride and sodium alginate were tested. Encapsulation beads produced by mixing 50 mM calcium chloride and 3.5% sodium alginate supported the optimal in vitro conversion potential. The number of multiple shoots formed by encapsulated nodal segments was not significantly different from the average of shoots produced by non-encapsulated nodal segments. The encapsulated nodal segments regenerated in vitro on different medium. The optimal germination and regeneration medium was Murashige-Skoog medium. Plantlets regenerated from the encapsulated nodal segments were hardened, acclimatized and established well in the field, showing similar morphology with parent plants. This encapsulation technology would serve as an alternative in vitro regeneration system for L. chinensis.

Establishment of an efficient plant regeneration culture protocol and achievement of successful genetic transformation in Jatropha curcas L.

Science.gov (United States)

Liu, Ying; Liu, Guoxuan; Yang, Yali; Niu, Sufang; Yang, Fuguang; Yang, Shaoxia; Tang, Jianian; Chen, Jianping

2017-12-01

An efficient and reproducible protocol is described for shoot-bud regeneration and Agrobacterium tumefaciens-mediated genetic transformation of J. curcas. Treating the explants with high concentrations (5-120 mg/L) of TDZ for short durations (5-80 min) before inoculation culture increased significantly the regeneration frequency and improved the quality of the regenerated buds. The highest shoot-buds induction rate (87.35%) was achieved when petiole explants were treated with 20 mg/L TDZ solution for 20 min and inoculated on hormone-free MS medium for 30 days. Regenerated shoots of 0.5 cm or a little longer were isolated and grafted to seedling stocks of the same species, and then the grafted plantlets were planted on half-strength MS medium containing 0.1 mg/L IBA and 2 mg/L sodium nitroprusside (SNP). This grafting strategy was found to be very effective, to obtain that healthy grafted plantlets ready for acclimatization within 20 days. By the above mentioned protocol and with general Agrobacterium - mediated genetic transformation methods only 65 days were needed to obtain intact transgenic plants.

Multiplicação in vitro do porta-enxerto de Prunus spp. 'Carelli' In vitro multiplication of Prunus spp. rootstocks 'Carelli'

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Paulo de Tarso Teixeira

2004-08-01

test tubes containing 20 ml of Lepoivre culture medium supplemented with 0.0; 0.5; 1.0; 2.0 and 4.0 mg.L-1 BA. This nodal segments were originated from in vitro pre-established plants after two subcultures in Lepoivre culture medium supplemented with 0.5 mg.L-1 BA. Evaluations done after 21 days in culture showed that the different levels of BA resulted in values statistically similar for the multiplication rate and length of regenerated shoots. The values for multiplication rate ranged from 3.3 to 3.4 shoots/explant. The treatment control did not result in multiple shoots but showed shoots with the highest lengh (16.2 mm. BA significativelly affected the length of shoots and the increase in its levels reduced the shoot lengh. BA levels higher than 1.0 mg.L-1 reduced the shoot length and promoted vitrification. BA in the level of 0.5 mg.L-1 resulted in the induction of 3.3 shoots/explant with means values of 11.0 mm in length in adequate conditions for rooting.

In vitro evaluation of electrospun chitosan mats crosslinked with genipin as guided tissue regeneration barrier membranes

Science.gov (United States)

Norowski, Peter Andrew, Jr.

Guided tissue regeneration (GTR) is a surgical technique commonly used to exclude bacteria and soft tissues from bone graft sites in oral/maxillofacial bone graft sites by using a barrier membrane to maintain the graft contour and space. Current clinical barrier membrane materials based on expanded polytetrafluoroethylene (ePTFE) and bovine type 1 collagen are non-ideal and experience a number of disadvantages including membrane exposure, bacterial colonization/biofilm formation and premature degradation, all of which result in increased surgical intervention and poor bone regeneration. These materials do not actively participate in tissue regeneration, however bioactive materials, such as chitosan, may provide advantages such as the ability to stimulate wound healing and de novo bone formation. Our hypothesis is that electrospun chitosan GTR membranes will support cell attachment and growth but prevent cell infiltration/penetration of membrane, demonstrate in vitro degradation predictive of 4--6 month in vivo functionality, and will deliver antibiotics locally to prevent/inhibit periopathogenic complications. To test this hypothesis a series of chitosan membranes were electrospun, in the presence or absence of genipin, a natural crosslinking agent, at concentrations of 5 and 10 mM. These membranes were characterized by scanning electron microscopy, tensile testing, suture pullout testing, Fourier transform infrared spectroscopy, X-ray diffraction, and gel permeation chromatography, and in vitro biodegradation for diameter/morphology of fibers, membrane strengths, degree of crosslinking, crystallinity, molecular weight, and degradation kinetics, respectively. Cytocompability of membranes was evaluated in osteoblastic, fibroblastic and monocyte cultures. The activity of minocycline loaded and released from the membranes was determined in zone of inhibition tests using P. gingivalis microbe. The results demonstrated that genipin crosslinking extended the in vitro

In vitro study of stem cell communication via gap junctions for fibrocartilage regeneration at entheses.

Science.gov (United States)

Nayak, Bibhukalyan Prasad; Goh, James Cho Hong; Toh, Siew Lok; Satpathy, Gyan Ranjan

2010-03-01

Entheses are fibrocartilaginous organs that bridge ligament with bone at their interface and add significant insertional strength. To replace a severely damaged ligament, a tissue-engineered graft preinstalled with interfacial fibrocartilage, which is being regenerated from stem cells, appears to be more promising than ligament-alone graft. Such a concept can be realized by a biomimetic approach of establishing a dynamic communication of stem cells with bone cells and/or ligament fibroblasts in vitro. The current study has two objectives. The first objective is to demonstrate functional coculture of bone marrow-derived stem cells (BMSCs) with mature bone cells/ligament fibroblasts as evidenced by gap-junctional communication in vitro. The second objective is to investigate the role of BMSCs in the regeneration of fibrocartilage within the coculture. Rabbit bone/ligament fibroblasts were dual-stained with DiI-Red and calcein (gap-junction permeable dye), and cocultured with unlabeled BMSCs at fixed ratio (1:10). The functional gap junction was demonstrated by the transfer of calcein from donor to recipient cells that was confirmed and quantified by flow cytometry. Type 2 collagen (cartilage extracellular matrix-specific protein) expressed by the mixed cell lines in the cocultures were estimated by real-time reverse transcription PCR and compared with that of the ligament-bone coculture (control). Significant transfer of calcein into BMSCs was observed and flow cytometry analyses showed a gradual increase in the percentage of BMSCs acquiring calcein with time. Cocultures that included BMSCs expressed significantly more type 2 collagen compared with the control. The current study, for the first time, reported the expression of gap-junctional communication of BMSCs with two adherent cell lines of musculoskeletal system in vitro and also confirmed that incorporation of stem cells augments fibrocartilage regeneration. The results open up a path to envisage a composite

De novo regeneration of Scrophularia yoshimurae Yamazaki (Scrophulariaceae) and quantitative analysis of harpagoside, an iridoid glucoside, formed in aerial and underground parts of in vitro propagated and wild plants by HPLC.

Science.gov (United States)

Sagare, A P; Kuo, C L; Chueh, F S; Tsay, H S

2001-11-01

A protocol for de novo regeneration and rapid micropropagation of Scrophularia yoshimurae (Scrophulariaceae) has been developed. Multiple shoot development was achieved by culturing the shoot-tip, leaf-base, stem-node and stem-internode explants on Murashige and Skoog (MS) medium supplemented with 4.44 microM N6-benzyladenine (BA) and 1.07 microM alpha-naphthaleneacetic acid (NAA). Stem-node and shoot-tip explants showed the highest response (100%) followed by stem-internode (74.4%) and leaf-base (7.7%) explants. The shoots were multiplied by subculturing on the same medium used for shoot induction. Shoots were rooted on growth regulator-free MS basal medium and the plantlets were transplanted to soil and acclimatized in the growth chamber. The content of harpagoside, a quantitatively predominant iridoid glycoside, in different plant material was determined by high performance liquid chromatography (HPLC). The analysis revealed that the content of harpagoside in the aerial and underground parts of S. yoshimurae was significantly higher than the marketed crude drug (underground parts of Scrophularia ningpoensis).

Micropropagation of Plantago asiatica L. through culture of shoot-tips

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Joanna Makowczyńska

2011-01-01

Full Text Available Shoot-tip multiplication of the medicinal species - Plantago asiatica was carried on MS medium with IAA and BAP or kinetin. Best results in micropropagation were achieved by adding 0.1 mg/dm3 IAA and 1 mg/dm3 BAP. After 6 weeks shoots were transferred to MS medium for rooting. The resulting plantlets were transferred after 8 weeks into pots and after a period of adaptation into the ground (field culture. The species Plantago asiatica was propagated in vitro by shoot-tip multiplication for the first time.

Wide variety of flower-color and -shape mutants regenerated from leaf cultures irradiated with ion beams

International Nuclear Information System (INIS)

Okamura, M.; Yasuno, N.; Ohtsuka, M.; Tanaka, A.; Shikazono, N.; Hase, Y.

2003-01-01

The efficiency of ion-beam irradiation combined with tissue culture in obtaining floral mutants was investigated and compared with those of gamma rays and X-rays in carnation. Leaf segments of carnation plants in vitro were irradiated with the 220 MeV carbon ions, and cultured till the shoot regenerated. The carbon ion had the highest effect in reducing the regeneration frequency, and the RBE value with respect to gamma-rays was four. The higher mutation frequency and the wider mutation spectrum were obtained in plants irradiated with the carbon ions than low LET radiations. Three new carnation varieties developed by ion-beam irradiation were applied for the registration of the Japanese Ministry of Agriculture, Forestry and Fisheries. The results indicate that ion beam irradiation could induce wide variety of flower-color and -shape mutants, and that the combined method of ion-beam irradiation with tissue culture is useful to obtain the commercial varieties in a short time

In vitro plant regeneration of two cucumber (Cucumis sativum L. genotypes: Effects of explant types and culture medium

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Grozeva Stanislava

2014-01-01

Full Text Available The effect of different phytohormone concentrations on callusogenesis and organogenesis in two cucumber genotypes were studied. It was established that the rate of plant regeneration depends on genotype, explant type and culture medium. Hypocotyls were found to be more responsive than cotyledons in morphogenesis. In vitro planlet-regenerants have been obtained in hypocotyls explants on culture medium with 1.0 and 2.0 mgL-1 BA for cultivar Gergana and in 1.0 and 3.0 mgL-1K-line 15B. Induction of regeneration in cotyledons were established only in cultivar Gergana on culture medium supplemented with 3.0 mgL-1 BA and in combination of 0.5 mgL-1IAA.

Effect of benzylaminopurine (BAP) pulsing on in vitro shoot ...

African Journals Online (AJOL)

user

2011-03-28

Mar 28, 2011 ... Research Cluster, Faculty of Science, University Malaya, 50603 Kuala Lumpur, Malaysia. Accepted 12 ... number of bud formation in shoot cultures of Musa acuminata cv. ... In tissue culture, plant growth regulators (PGR) are.

Shoot multiplication of Paphiopedilum orchid through in vitro cutting ...

African Journals Online (AJOL)

waraporn

2012-09-20

Sep 20, 2012 ... regulators could remain higher shoot multiplication than in other media. The micropropagation ... stalk nodes, buds, root tips and rhizome segments. For mass ... plants, the future mass-market orchids will most likely be.

In vitro propagation of ginger (Zingiber officinale Rosc.) through direct organogenesis: a review.

Science.gov (United States)

Seran, Thayamini H

2013-12-15

Ginger (Zingiber officinale Rosc.) is a perennial herb. It belongs to the family Zingiberaceae and commercially cultivated in most tropical regions of the world. The underground rhizomes are the planting materials in a conventional propagation of ginger however it has a low multiplication rate. It is known that there are possible methods are available for rapid vegetative propagation of ginger through direct organogenesis or somatic embryogenesis under in vitro conditions but it is necessary to find the best protocol for in vitro multiplication of ginger. Limited studies on the tissue culture technology of ginger are available in Sri Lanka. However, significant efforts have been made in the procedure for in vitro micropropagation in the other ginger growing countries. The available literature with respect to in vitro plant regeneration has been perused and this review mainly focused on the in vitro propagation via direct organogenesis from rhizome buds or shoot tips of ginger often used as explants. This review article may be an appropriate and effective guidance for establishing in vitro cultures and subsequent production of in vitro plantlets in clonal propagation of ginger.

Anatomical, germination and in vitro studies on shea tree (Vitellaria paradoxa.) seed

International Nuclear Information System (INIS)

Iddrisu, A.

2013-07-01

In vitro and in vivo germination and regeneration studies were conducted on the development of vitellaria paradoxa seedlings as an initial effort towards its domestication. However, to achieve this objective, the morphology and anatomy of the seeds were first studied because they influence germination. Although a smooth, brown coast enclosed a V. paradoxa seed, it did not impose dormancy on the embroyo. Transverse and longitudinal sections through the seed showed that the embrayo is surrounded by latex and fat-containing tissues which made its identification difficult. Thus, the embrayo was identified by immersing transversely cut seeds in 1.0 % tetrazolium chloride (TTC) solution for 24 hours which stained it red. When V. paradoxa seeds of similar size were sown on nursery beds, the resulting seedlings developed through seven stages namely sprouting, pseudoradicle elongation, bulging, appearance of the shoot on the pseudoradicle, shoot elongation, emergence and seedling establishment. The pseudoradicle is the fused petioles of the two cotyledons and a transverse section through it revealed an outer sheath and lactiferous vessels surrounding a central hollow tube. Longitudinal section also show the lactiferous vessels surrounding the central hallow tube in which the plumule moves through until it reaches the bulge of the pseudoradicle where it develops into a rudimentary shoot. The rudimentary shoot then pseudoradicle and grows upwards. Classifying seeds into three groups based on size significantly affected days to germination and the morphology of the resulting seedlings. Large seeds germinated within one week after sowing with vigorous growth compared to small and medium seeds. Although the seedcoat of V. paradoxa never imposed dormancy, deshelling (removal of the seedcoat) significantly led to early germination and synchronous seedling emergence compared to those for intact seeds (control). In vitro culture of intact and deshelled seeds on Muarashige and Skoog

Embryogenesis induction, callogenesis, and plant regeneration by in vitro culture of tomato isolated microspores and whole anthers.

Science.gov (United States)

Seguí-Simarro, José M; Nuez, Fernando

2007-01-01

In this work, some of the different in vitro developmental pathways into which tomato microspores or microsporocytes can be deviated experimentally were explored. The two principal ones are direct embryogenesis from isolated microspores and callus formation from meiocyte-containing anthers. By means of light and electron microscopy, the process of early embryogenesis from isolated microspores and the disruption of normal meiotic development and change of developmental fate towards callus proliferation, morphogenesis, and plant regeneration have been shown. From microspores isolated at the vacuolate stage, embryos can be directly induced, thus avoiding non-androgenic products. In contrast, several different morphogenic events can be triggered in cultures of microsporocyte-containing anthers under adequate conditions, including indirect embryogenesis, adventitious organogenesis, and plant regeneration. Both callus and regenerated plants may be haploid, diploid, and mostly mixoploid. The results demonstrate that both gametophytic and sporophytic calli occur in cultured tomato anthers, and point to an in vitro-induced disturbance of cytokinesis and subsequent fusion of daughter nuclei as a putative cause for mixoploidy and genome doubling during both tetrad compartmentalization and callus proliferation. The potential implications of the different alternative pathways are discussed in the context of their application to the production of doubled-haploid plants in tomato, which is still very poorly developed.

Plants regeneration from African cowpea variety (Vigna unguiculata ...

African Journals Online (AJOL)

STORAGESEVER

2008-08-18

Aug 18, 2008 ... Vigna unguiculata (L.) Walp. plant was efficiently regenerated from cotyledonary node explants. The shoots multiplication rate was ... Africa, insect pests are often responsible for 100% losses of cowpea yields (Singh and .... 26 days after acclimatization first flower buds were observed; thus plants were ...

Effect of Plant Growth Regulators on a Shoot and Root Formation from the Leaf and Flower Culture of a Standard-type Chrysanthemum 'Jinba'

International Nuclear Information System (INIS)

Lee, J.S.; Lee, G.J.; Chung, S.J.; Kim, J.B.; Kim, D.S.; Kang, S.Y.

2008-01-01

In this study we investigated the conditions of a higher frequency for regenerated plants from different explants of a standard-type chrysanthemum 'Jinba'. In vitro culture was initiated on an MS medium containing 3% sucrose, 0.8% agar, and 5 μM benzyl adenine (BA) with naphthalene acetic acid (NAA) by using surface-sterilized leaf and flower tissues from greenhouse-grown plants. Direct shoot regeneration from the leaf and flower explants was obtained 21 to 28 days after the initial culture. Among the seven combinations of the growth regulators used for the culture, the most efficient condition for the shoot and root formation from the leaf tissue was obtained when the MS basic medium was supplemented with 0.5 mg L-¹ BA and 1.0 mg L-¹ NAA, and 0.1 mg L-¹ BA and 0.5 mg L-¹ NAA, while the culture using floret tissues was most efficient on the medium supplemented with 0.5 mg L-¹ BA and 0.5 mg L-¹ NAA, and 0.1 mg L-¹ BA and 1.0 mg L-¹ NAA. These results will provide valuable information to help set up an efficient system for a tissue culture of chrysanthemum cv. Jinba to improve one or some of its negative traits in combination with a radiation mutagenesis approach

Plant regeneration of Erigeron breviscapus (vant.) Hand. Mazz. and its chromatographic fingerprint analysis for quality control.

Science.gov (United States)

Liu, Chun-Zhao; Gao, Min; Guo, Bin

2008-01-01

An efficient micropropagation system for Erigeron breviscapus (vant.) Hand. Mazz., an important medicinal plant for heart disease, has been developed. Shoot organogenesis occurred from E. breviscapus leaf explants inoculated on a medium supplemented with a combination of plant growth regulators. On average, 17 shoots per leaf explant were produced after 30 days when they were cultured on MS basal salts and vitamin medium containing 5 microM 6-benzylaminopurine (BAP) and 5 microM 1-naphthaleneacetic acid (NAA). All the regenerated shoots formed complete plantlets on a medium containing 2.5-10 microM indole-3-butyric acid (IBA) within 30 days, and 80.2% of the regenerated plantlets survived and grew vigorously in field conditions. Based on the variation in common peaks and the produced amount of the most important bioactive component, scutellarin, a high performance liquid chromatography (HPLC) fingerprinting system was developed for quality control of these micropropagated plants. Chemical constituents in E. breviscapus micropropagated plants varied during plant development from regeneration to maturation, the latter of which showed the most similar phytochemical profile in comparison with mother plants. The regeneration protocol and HPLC fingerprint analysis developed here provided a new approach to quality control of micropropagated plants producing secondary metabolites with significant implications for germplasm conservation.

Browse Title Index

African Journals Online (AJOL)

Items 201 - 250 of 11090 ... Issue, Title. Vol 12, No 49 (2013), In vitro regeneration of selected Kenyan papaya (Carica papaya L.) lines through shoot tip culture, Abstract PDF. Naomi Nzilani Mumo, Fredah Karambu Rimberia, George Edward Mamati, Agnes Wanjiru Kihurani. Vol 7, No 12 (2008), In vitro regeneration of Turkish ...

Establishment of plant regeneration system from anther culture of ...

African Journals Online (AJOL)

ajl yemi

2011-11-28

Nov 28, 2011 ... Plant regeneration of Tagetes patula was achieved from anther explants via adventitious shoot differentiation from callus. The effects of genotype, temperature pretreatment, plant growth regulators, light regimes and sucrose concentration were studied. Eight of ten genotypes tested were successfully.

Influence of Photo period and Gamma Irradiation on Shoot Development and Chemical Composition of Yucca elephantipes Regel Plant in Vitro

International Nuclear Information System (INIS)

El-Khateeb, M.A.; El-Sharnouby, M.E.; Ragab, E.A.

2008-01-01

In-Vitro propagated plant lets of Yucca elephantipes were placed under different photo periods (24/0, 16/8, and 0/24 L/D) for two months after cultured on MS medium supplemented with 40 g/l sucrose. Growing explants of Yucca elephantipes cultured on MS medium placed under photo period 24/0 L/D gave the highest shoot length and best proliferation than other treatments especially in subculture 3. Exposure to gamma irradiation at doses 0.0, 5 ,10, 20 and 40 Gy, and placed on the same conditions .Irradiated plantlets exhibited changes in shoot growth especially on photo period at 16/8 L/D than others, also with gamma irradiation at dose 20 Gy. Using gamma irradiation at level of 20 and 40 Gy stimulated both leaf shape and thickness of stem. The contents of total phenol, total saponin and some unsap contents were increased with increasing gamma irradiation

Avaliação da capacidade de regeneração in vitro em tomateiro industrial Evaluation of the in vitro regeneration capacity in Brazilian industrial tomato

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MIKLÓS FÁRI

2000-08-01

Full Text Available Este trabalho teve por objetivo avaliar a capacidade de regeneração das cultivares de tomateiro industrial (Lycopersicon esculentum Mill IPA-5 e IPA-6, utilizando quatro composições de meio de cultura descritos na literatura e cinco variações de inoculação. Foi testada uma nova variação de inoculação, denominada cotilédone fendido. A maior freqüência de formação de gemas vegetativas foi 100% no caso de IPA-5, e 65% no caso de IPA-6. Para induzir o alongamento de brotos, foram necessários três subcultivos dos explantes apresentando gemas. No caso de IPA-5, o número de brotos obtidos foi maior quando a indução de gemas foi realizada em meio contendo BAP (2,5 mg L-1 e AIA (0,2 mg L-1 seguido de três subcultivos, em meio como zeatina (0,5 mg L-1. Usando esse protocolo, a cultivar IPA-5 produziu uma média de 5,45 brotos alongados a partir do cotilédone fendido. Essa capacidade excedeu significativamente o cotilédone aparado, que produziu 4,4 brotos alongados por explante. No caso de IPA-6, a melhor combinação de meios e método de inoculação produziu 0,87 broto alongado por explante. Os brotos alongados foram enraizados e transferidos para casa de vegetação.The objective of this work was to evaluate the regeneration capacity of the IPA-5 and IPA-6 Brazilian industrial tomato (Lycopersicon esculentum Mill cultivars using four compositions of culture media described in the literature and five inoculation methods. A new variation of inoculation, the split cotyledon method, was also tested. The largest frequency of shoot bud formation was 100% in the case of IPA-5 and 65% in the case of IPA-6. To induce shoot elongation, it was necessary to accomplish three subcultures of the explants presenting shoot buds. In the case of IPA-5, the number of the obtained shoots was higher when the induction of shoot buds was accomplished in culture medium containing BAP (2.5 mg L-1 and IAA (0.2 mg L-1 followed by three subcultures on

In vitro rescue of interspecific embryos from Elaeis guineensis x E. oleifera (Arecaceae

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Paula Cristina da Silva Angelo

2011-09-01

Full Text Available The African oil palm (Elaeis guineensis is the most effective oil producer in tons per hectare. Nevertheless, its increasing cultivation in Latin America is harmed by the “lethal yellowing”. Genetic resistance to this anomaly can be found in the germplasm of American oil palm or caiaué (E. oleifera, a native species from the Amazon rainforest. However, the procedures adopted to induce seeds of E. guineensis to germination frequently result mild for interespecific hybrids. Embryo in vitro cultivation can be a viable option. This work was aimed initially to test liquid MS medium supplemented with different glucose or sucrose concentrations for the in vitro cultivation of zygotic embryos from E. guineensis x E. oleifera controlled pollinations. Additionally we investigated different compost mixtures to acclimatize the regenerated hybrid plantlets. Concentrations of 10, 20 and 30g/L of both sugars were tested on flasks containing five mature zygotic embryos, with 15 repetitions per treatment in a total of 450 explants. The number of embryos displaying shoots and radicles at least 2mm in length per experimental unit was evaluated during phase one of in vitro cultivation. Plantlets displaying shoots and radicles were transferred to phase two of in vitro cultivation and subsequently to acclimatization, under 70% shading with manual water supply. The experiments of acclimatization were conducted with 130 plantlets randomly distributed in pure horticultural compost, 3:1 or 1:1 compost:sand mixtures and each plantlet was defined as an experimental unit. Data were submitted to ANOVA, t test and analyzes of correlation (p≤0.05. Highest emergence rates were 97% for shoots and 73% for radicles, observed in MS medium supplemented with 20g/L (110mM of glucose. This sugar in concentrations of 20 or 30g/L provided balanced shoot/root development, and this was considered one of the reasons for the higher frequency of plantlet establishment. The survival

Micropropagation of Codiaeum variegatum (L.) Blume and regeneration induction via adventitious buds and somatic embryogenesis.

Science.gov (United States)

Radice, Silvia

2010-01-01

Codiaeum variegatum (L) Blume cv. "Corazon de oro" and cv. "Norma" are successfully micropropagated when culture are initiated with explants taken from newly sprouted shoots. The establishment and multiplication steps are possible when 1 mg/L BA or 1 mg/L IAA and 3 mg/L 2iP are added to MS medium, according to the cultivar respectively selected.Adventive organogenesis and somatic embryogenesis are induced from leaf explants taken from in vitro buds of croton. On leaf-sectioned of "Corazon de oro" cultured in vitro, 1 mg/L BA stimulates continuous somatic embryos development and induces some shoots too. Replacing BA with 1 mg/L TDZ induces up to 100% bud regeneration in the same explants. On the other hand, leaf-sectioned of C. variegatum cv. Norma does not start somatic embryo differentiation if 1 mg/L TDZ is not added to the MS basal medium. Incipient callus is observed after 30 days of culture, and then, subculture to MS with 1 mg/L BA allows the same process to show on the "Corazon de oro" cultivar. Somatic embryos show growth arrest that is partially overcome by transfer to hormone-free basal medium with activated charcoal. Root induction is possible on basal medium plus 1 mg/L IBA. Plantlets in the greenhouse have variegated leaves true-to-type.

In vitro regeneration of kidney from pluripotent stem cells

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Osafune, Kenji, E-mail: osafu@cira.kyoto-u.ac.jp [Center for iPS Cell Research and Application (CiRA), Kyoto University, 53 Kawahara-cho, Shogoin, Sakyo-ku, Kyoto 606-8507 (Japan); PRESTO, Japan Science and Technology Agency (JST), 4-1-8 Honcho, Kawaguchi, Saitama 332-0012 (Japan); JST Yamanaka iPS Cell Special Project, Japan Science and Technology Agency (JST), 4-1-8 Honcho, Kawaguchi, Saitama 332-0012 (Japan)

2010-10-01

Although renal transplantation has proved a successful treatment for the patients with end-stage renal failure, the therapy is hampered by the problem of serious shortage of donor organs. Regenerative medicine using stem cells, including cell transplantation therapy, needs to be developed to solve the problem. We previously identified the multipotent progenitor cells in the embryonic mouse kidney that can give rise to several kinds of epithelial cells found in adult kidney, such as glomerular podocytes and renal tubular epithelia. Establishing the method to generate the progenitors from human pluripotent stem cells that have the capacity to indefinitely proliferate in vitro is required for the development of kidney regeneration strategy. We review the current status of the research on the differentiation of pluripotent stem cells into renal lineages and describe cues to promote this research field.

In vitro regeneration of kidney from pluripotent stem cells

International Nuclear Information System (INIS)

Osafune, Kenji

2010-01-01

Although renal transplantation has proved a successful treatment for the patients with end-stage renal failure, the therapy is hampered by the problem of serious shortage of donor organs. Regenerative medicine using stem cells, including cell transplantation therapy, needs to be developed to solve the problem. We previously identified the multipotent progenitor cells in the embryonic mouse kidney that can give rise to several kinds of epithelial cells found in adult kidney, such as glomerular podocytes and renal tubular epithelia. Establishing the method to generate the progenitors from human pluripotent stem cells that have the capacity to indefinitely proliferate in vitro is required for the development of kidney regeneration strategy. We review the current status of the research on the differentiation of pluripotent stem cells into renal lineages and describe cues to promote this research field.

Evaluation of Sugarcane (Saccharum officinarum L. Somaclonals Tolerance to Salinity Via In Vitro and In Vivo

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HAMID RAJABI MEMARI

2011-06-01

Full Text Available Tissue culture technique was used to obtain salt tolerant variants from embryogenic calluses of sugarcane (Saccharum sp. var. CP48-103 that cultured on a selective medium containing different levels of NaCl (0, 0.2, 0.4, 0.6, and 0.8% NaCl. A total of four plants regenerated from the tolerant calluses were selected but the best of them in vigor grown in in vitro and hydroponic systems under salinity stress to comparison with source variety. With increasing supply of NaCl in both systems, root growth was more adversely affected than was shoot growth. Chlorophyll contents showed a decreasing trend and dry matter yield of plants reduced but in a slow rate in tolerant somaclonal than source variety. The biochemical analysis showed that at high salt concentration, Cl- and Na+ content in shoot and root increased. With rising salt concentration from 0 to 0.8%, content of Cl- in shoot and root of tolerant variant changed lower than parent showed that this variant had genetic lowest ratio of shoot/root chloride and had minimum transport of Cl- to shoots. Also this variant had high content of Ca2+ in shoot and high K+/Na+ ratio at all salinity levels. Thus it probably has genetic potential to avoid harmful ions uptake.

The antioxidant effect of Asparagus cochinchinensis (Lour.) Merr. shoot in D-galactose induced mice aging model and in vitro.

Science.gov (United States)

Lei, Linghua; Ou, Lijun; Yu, Xiaoying

2016-04-01

An increasing number of plant components and their extracts have been shown to have beneficial health effects in humans. We aimed to explore the antioxidant effects of the aqueous extract of Asparagus cochinchinensis (Lour.) Merr. shoot in vivo and in vitro. A total of 80 Kun Ming mice were randomly divided into four groups (20/group). The mice in the control group received a daily subcutaneous injection of saline. A daily injection of D-galactose was administered to the aging model group, the vitamin C (Vc) group (positive control group), and the extract treatment group. Regular measurement of blood cells, nitric oxide synthase (NOS), catalase (CAT) activities, superoxide dismutase (SOD) activities, nitric oxide (NO), and malondialdehyde (MDA) concentration, and the expressions of NOS, SOD, and glutathione peroxidase (GPX) in serum levels were obtained. Furthermore, the microstructure of mice viscera was observed using hematoxylin and eosin staining. The aqueous extract of A. cochinchinensis (Lour.) Merr. had similar 1,1-diphenyl-2-picrylhydrazyl radical 2,2-diphenyl-1-(2,4,6-trinitrophenyl) hydrazyl (DPPH·) [or 2,2'-azino-bis (3-ethylbenzothiazoline-6-sulfonic acid) (ABTS+)] and higher hydroxyl radicals (or superoxide anion; p < 0.05) radical scavenging capabilities to Vc. Moreover, compared with the aging model group, the aqueous extract of A. cochinchinensis (Lour.) Merr. shoot could obviously increase NOS, CAT, and SOD activities and the NO content, and reduce the MDA content (p < 0.05). Additionally, the microstructure of mice viscera was obviously improved and the expressions of NOS, SOD and GPX were also manifestly increased in the treatment group (p < 0.05). The aqueous extract of A. cochinchinensis (Lour.) Merr. shoot had a strong radical scavenging capability in vivo and in vitro, and might be used to diminish radicals in the body and consequently prevent aging. Copyright © 2016. Published by Elsevier Taiwan LLC.

Factors influencing in vitro shoot regeneration of Macadamia ...

African Journals Online (AJOL)

STORAGESEVER

2008-11-19

Nov 19, 2008 ... industry that targets niche markets in Europe and the. *Corresponding author ... Guatemala, Brazil, Costa Rica and Fiji (Kiuru et al.,. 2004). Macadamia is ... months, skilled manpower, material inputs and space. (Nyakundi and ...

In vitro adventitious shoot regeneration and acclimatisation of ...

African Journals Online (AJOL)

Different potting media were assessed during plantlet acclimatization. The highest percentage of plant survival (83.33%) was on the medium that contained sand and soil (1:1), while maximum root length (4.37 cm) and plant height (7.87 cm) were attained in potting medium that consisted peat moss, perlite and vermiculite ...

Cryopreservation of in vitro shoot apices of Oxalis tuberosa Mol.

Science.gov (United States)

Gonzalez-Benito, M E; Mendoza-Condori, V H; Molina-Garcia, A D

2007-01-01

Oca (Oxalis tuberosa Mol.) is an under-utilized tuber crop from the Andean region. Cryopreservation would allow the safe and long-term preservation of the genetic resources of this crop. A protocol for the cryopreservation of in vitro grown shoots has been developed using the vitrification solution PVS2. Two genotypes were studied (G1 and G27). Nodal segments were cultured on MS medium and incubated at 10 degree C with 16 h photoperiod and 10 mol per square meter per second irradiance, for two weeks. Apices were then excised and cultured on MS+0.15 M sucrose for 3 days at 5 degree C in darkness. Subsequently, apices were immersed in a loading solution (liquid MS medium+2 M glycerol+0.4 M sucrose), and then treated with the vitrification solution PVS2 for 0 to 40 minutes. Cryovials were then immersed in liquid nitrogen. Four weeks after rewarming and culture on recovery medium, genotype G1 showed approximately 60 percent recovery (normal growth) with 20 min PVS2 treatment. Genotype G27 showed lower recovery (30 percent). Differential scanning calorimetry yielded a Tg midpoint for PSV2 solution of ca. -120 degree C. Calorimetric studies on apices at different stages of the cryopreservation protocol showed a change in calorimetric parameters consistent with a decrease in the amount of frozen water as the protocol advanced.

Induction of multiple shoots from leaf segments, in vitro-flowering and fruiting of a dwarf tomato (Lycopersicon esculentum).

Science.gov (United States)

Rao, Kokkirala Venugopal; Kiranmayee, Kasula; Pavan, Umate; Sree, Telakalapalli Jaya; Rao, Alleni V; Sadanandam, Abbagani

2005-08-01

Multiple shoots were induced from leaf explants of Lycopersicon esculentum cultivar MicroTom, within 20-25d, on MS medium supplemented with 8.9 microM benzylaminopurine (BAP)+1.14 microM indole-3-acetic acid (IAA). For rooting, elongated microshoots were excised and transferred onto MS medium supplemented with 4.9 microM indole-3-butyric acid (IBA). Well-developed roots and flower raceme were obtained on d 7 and 13, respectively, upon transfer of the microshoots onto rooting medium. The flowers self-fertilized in vitro and produced mature fruits in additional 15-17d of culture.

Regeneration of some monocotyledonous plants from subterranean organs in vitro

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Krystyna D. Kromer

2013-12-01

Full Text Available The aim in view was investigation ofthe regenerative potential of rhizomes, bulbs and corms of ten monocotyledonous plant species from four families: Amaryllidaceae (Haemanthus katharinae, Crinum abyssinicum, Leucojum vernum, Araceae (Spathiphyllum wallisii, Iridaceae (Crocus vernus, Iris germanica, Liliaceae (Hosta lancifolia, Muscari racemosum, Scilla laxiflora, Veltheimia viridifolia under conditions of in vitro culture. All the investigated species were capable of buld or, alternatively, bud and root regeneration. Different morphogenetic potential was noted between the particular families. A high regenerative potential under the conditions of culture applied was characteristic for plants of the Liliaceae and Araceae families, it was lower in plants belonging to Amaryllidaceae and lowest in those of the Iridaceae family. Plants from the Liliaceae family exhibited also the highest ability of callus formation, whereas Amaryllidaceae and Iridaceae plants possessed this ability in only a low degree. The influence of growth regulators of the auxin group (NAA, IAA, 2,4-D and of cytokinins (K on the initiation and course of organogenesis was tested. The results of the experiments indicate that auxins in interaction with kinetin gave the highest percentage of regenerating explants and also a large number of buds on the latter. Stimulation of callus tissue was highest under the influence of 2,4-D, and weaker when NAA IAA were used.

Robust regeneration protocol for the Agrobacterium tumefaciens mediated transformation of Solanum tuberosum

International Nuclear Information System (INIS)

Abbasi, A.; Bilal, M.; Hussain, J.; Shah, M. M.; Hassan, A.

2016-01-01

Plant genetic transformation requires robust regeneration system. Plant growth regulators (PGRs) such as cytokinins (CKs) play a pivotal role in organogenesis; however, CKs are the most expensive PGRs. In the current study, an efficient yet economical protocol for regeneration of potato plant was developed. Stem inter-nodal and leaf explants were cultured on different regeneration media supplemented with varying concentration of different CKs such as kinetin and zeatin. Murashige- Skoog media added with zeatin (1, 1.5 mg/L) was designated as RZ1, RZ1.5, respectively or kinetin (1.5, 2 mg/L) was designated as RK1.5 and RK2, respectively, however, concentrations of other hormones such as NAA (1-Naphthaleneacetic acid) and GA3 (Gibberellic acid A3) were kept same. RZ1 and RZ1.5 gave significantly better Results as compared to RK-type media in all aspects studied such as callus initiation, days to first shoot emergence, number of shoots per explants. RZ1 medium was then selected as regeneration media for Agrobacterium-mediated transformation of potato plants with cyanobacterial phosphoenol pyruvate carboxylase gene, which provided multiple putative transformants on selection media. The transformants were further confirmed through PCR. The current protocol is found to be cost effective and efficient for the regeneration of Solanum tuberosum and can be successfully implied for the Agrobacterium-mediated transformation. (author)

In-vitro culture performance of rice cultivars

International Nuclear Information System (INIS)

Oard, J.H.; Rutger, J.N.

1989-01-01

Full text: Four California rice varieties (L-202, S-201, M-202 and Calmochi 101) and one Texas rice (Lemont) were tested for embryogenic calli produced from mature seeds. Of these varieties S-201, M-202 and Calmochi 101 derived from mutant crosses as follows (mutant parent underlined). S-201 (Calrose 76 x Cs-M3) x S6; M-202 (IR8 x Cs-M3) x (10-7 x M-101); Calmochi 101 Tatsumi mochi x (M7 x S6). S-201 was the only cultivar that produced many shoots from virtually all portions of the embryogenic callus. M-202 produced the highest no. of shoots at the upper level of hormone (BAP). Regeneration of Calmochi 101 was rather poor. The frequency of albino shoots was also different for the varieties tested: 0.027 for L-202; 0.00 for Calmochi 101; 0.016 for S-201; 0.045 for Lemont; 0.037 for M-202. Albino shoots were found only with 0.1 mg 1 -1 BAP in the regeneration medium, except for Lemont which produced albinos without cytokinin. (author)

THE EFFICIENCY OF SHOOT AND PLANTLET FORMATION OF Cephaelis ipecacuanha AFTER THREE SUBCULTURES IN VITRO

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Osmar Alves Lameira

1994-12-01

Full Text Available Multiple adventitious shoot formed from internodal segments of Cephaelis ipecacuanha cultured 25 days on Gamborg basal medium (GAMBORG et al., 1968 supplemented with 6.66mM 6-benzylaminopurine there was a maximum of nine shoots per segment and an average of five shoots per segment formed. The presence of gibberellic acid in the subculture media promoted shoot elongation in all treatments. The shoots attained 3cm in height and rooting of 100% after 35 days of culturing upon Murashige and Skoog's basal medium (MS, added with 4.92mM indole-3-butyric acid, 0.87m gibberellic acid and 0.1% activated charcoal. Further growth was accelerated after the transfer to 1/2 MS without growth regulators. Rooted plantlets transferred to potting soil could be successfully established.

MICROPROPAGATION OF ADULT TREE OF PTEROCARPUS MARSUPIUM ROXB. USING NODAL EXPLANTS

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Shipra JAISWAL

2015-12-01

Full Text Available Attempts were made for in vitro propagation of Pterocarpus marsupium Roxb., belonging to family Fabaceae, an economically important multipurpose tree. The tree is scared with noval antidiabetic properties. The tree shows poor seed germination capacity (30% due to hard seed coat and conventional vegetative regeneration methods are a complete failure. Therefore, the propagation of this tree by tissue culture techniques is an urgent need and well justified. Nodal segments containing axillary bud from 10 years old tree of P. marsupium were evaluated for axillary shoot proliferation on Murashige and Skoog’s (MS basal medium fortified with BAP (6–benzylaminopurine and kinetin (Kn singly or in combinations with auxins at different concentrations. The best shoot proliferation was obtained with 13.95 µM Kn + additives (568 µM Ascorbic acid, 260 µM Citric acid, 605 µM Ammonium sulphate and 217 µM Adenine sulphate in MS medium where 64.44% of the axillary buds responded with development of (2.51±0.10 shoots. Multiplication of in vitro shoots were achieved on MS Medium supplemented with Kn (9.30 µM + NAA (0.54 µM and additives. Half strength MS medium supplemented with 4.92 µM IBA induced in vitro rooting of in vitro shoots. In vitro regenerated plantlets with well developed roots were successfully hardened in a greenhouse.

Micropropagation of Hedychium coronarium J. Koenig through rhizome bud.

Science.gov (United States)

Mohanty, Pritam; Behera, Shashikanta; Swain, Swasti S; Barik, Durga P; Naik, Soumendra K

2013-10-01

An optimized protocol was developed for in vitro plant regeneration of a medicinally important herb Hedychium coronarium J. Koenig using sprouted buds of rhizomes. The rhizomes with sprouted bud were inoculated on Murashige and Skoog (Physiol Plant 15:473-497, 1962) medium (MS) supplemented with either N(6)-benzyladenine (BA) alone (1.0-4.0 mg L(-1)) or in combination with 0.5 mg L(-1) naphthalene acetic acid (NAA). Of these combinations, MS supplemented with a combination of 2.0 mg L(-1) BA and 0.5 mg L(-1) NAA was most effective. In this medium, best shoots (3.6) and roots (4.0) regeneration was observed simultaneously with an average shoot and root length of 4.7 cm and 4.2 cm respectively. Regeneration of shoots and roots in the same medium at the same time (One step shoot and root regeneration) reduced the time for production of in vitro plantlets and eliminates the media cost of rooting. Cent-percent (100 %) success in plant establishment was observed in both gradual acclimatization process as well as when plants were directly transferred to outdoor in clay pots containing a mixture of garden soil and sand (2:1) without any sequential acclimatization stage.

Micropropagation of Ajuga bracteosa, a medicinal herb.

Science.gov (United States)

Kaul, Shivanee; Das, Sandip; Srivastava, P S

2013-04-01

For conservation and genetic transformation, a successful in vitro micropropagation protocol for Ajuga bracteosa, a medicinal herb has been established for the first time. MS medium supplemented with IAA (2 mg/L) and BA (5 mg/L) induced 100 % shoot regeneration with an average of 41.4 shoots of 8.4 cm per culture. Excised in vitro shoots when transferred to MS + IBA (0.5 mg/L) produced 20 roots/shoot of 20.2 cm average length in 100 % cultures. Of the three explants, leaf, petiole and root, leaf displayed quickest response followed by petiole while root was the slowest. Hardening of plantlets was achieved with 82 % survival. The hardened plants were maintained in pots with garden soil under controlled (Temp. 25 ± 2 °C) conditions. RAPD exhibited genetic fidelity with 100 % monomorphism in regenerants.

Regeneration and molecular analysis of date palm ( Phoenix ...

African Journals Online (AJOL)

Clonal plants of date palm (Poenix dactylifera L.) were regenerated from juvenile leaves on regimes consisting of the use of 2,4-D. Success depended on the concentrations of 2,4-D tested. The cultures produced adventitious shoot buds directly at the basal part of leaves as well as excessive calli. Somaclonal variation in ...

Morfogênese in vitro de brotos de macieira (Malus domestica Borkh. a partir de fragmentos delgados de folhas In vitro morphogenesis of shoots of apple tree (Malus domestica Borkh. Starting from thin fragments of leaves

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Alan Cristiano Erig

2005-06-01

plants genetic transformation, and also for the fast multiplication of the modified genotype through the micropropagation, it is a efficient regeneration protocol. The objective of this work was to study the expression of the morphogenetic potential of thin fragments of apple tree leaves, and to optimize a regeneration protocol seeking futures works of genetic transformation. The completely randomized experimental design was used, in factorial outline 3 x 2 x 6, with three cultivate of apple tree (Galaxy, Maxigala and Mastergala, two explants types (thin fragments of leaves cut in the transversal and longitudinal direction, and six concentrations of thidiazuron (TDZ in the culture medium (0; 4.54; 9.08; 13.62; 18.16 and 22.7 µM, totaling 36 treatments. The MS medium salts and vitamins were added of myo-inositol (100 mg.L-1, sucrose (30 g.L-1, agar (6 g.L-1 and 1.6 µM naphthaleneacetic acid (NAA. Flasks with capacity for 150 mL with 6 mL of culture medium were used. The explants were obtained of plants in vitro cultivated, in multiplication phase, 45 days after the inoculation, and they were constituted of fine fragments of the medium part of leaves, cut in the tranversal direction (length from 8 to 10 mm or longitudinal (length from 10 to 12 mm, both with approximate width of 1 mm. The regeneration percentage, the intensity of callus formation and the number of shoots formed by explant were evaluated by the 45 days of cultivation. Starting from the obtained results it was ended that the expression of the morphogenetic potential of the thin fragments of leaf cut in the transversal direction is higher than those cut in the longitudinal direction, and its cultivation in culture medium with 4.54 µM of TDZ, propitiates, simultaneously, high regeneration percentage, great number of shoots formation and smaller callus intensity.

Role of HTRA1 in bone formation and regeneration: In vitro and in vivo evaluation.

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Gladys Filliat

Full Text Available The role of mammalian high temperature requirement protease A1 (HTRA1 in somatic stem cell differentiation and mineralized matrix formation remains controversial, having been demonstrated to impart either anti- or pro-osteogenic effects, depending on the in vitro cell model used. The aim of this study was therefore to further evaluate the role of HTRA1 in regulating the differentiation potential and lineage commitment of murine mesenchymal stem cells in vitro, and to assess its influence on bone structure and regeneration in vivo. Our results demonstrated that short hairpin RNA-mediated ablation of Htra1 in the murine mesenchymal cell line C3H10T1/2 increased the expression of several osteogenic gene markers, and significantly enhanced matrix mineralization in response to BMP-2 stimulation. These effects were concomitant with decreases in the expression of chondrogenic gene markers, and increases in adipogenic gene expression and lipid accrual. Despite the profound effects of loss-of-function of HTRA1 on this in vitro osteochondral model, these were not reproduced in vivo, where bone microarchitecture and regeneration in 16-week-old Htra1-knockout mice remained unaltered as compared to wild-type controls. By comparison, analysis of femurs from 52-week-old mice revealed that bone structure was better preserved in Htra1-knockout mice than age-matched wild-type controls. These findings therefore provide additional insights into the role played by HTRA1 in regulating mesenchymal stem cell differentiation, and offer opportunities for improving our understanding of how this multifunctional protease may act to influence bone quality.

A filter paper-based liquid culture system for citrus shoot organogenesis - a mixture-amount experiment

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The objective of this study was to determine the effects of a static liquid culture system on shoot regeneration from citrus epicotyl explants. Two citrus types were used, Carrizo citrange and Ridge Pineapple sweet orange. A liquid culture system comprised of a Petri dish, cellulose filter paper dis...

Rapid in vitro propagation, conservation and analysis of genetic stability of Viola pilosa.

Science.gov (United States)

Soni, Madhvi; Kaur, Rajinder

2014-01-01

A protocol for in vitro propagation was developed for Viola pilosa, a plant of immense medicinal value. To start with in vitro propagation, the sterilized explants (buds) were cultured on MS basal medium supplemented with various concentrations of growth regulators. One of the medium compositions MS basal + 0.5 mg/l BA + 0.5 mg/l TDZ + 0.5 mg/l GA3 gave best results for in vitro shoot bud establishment. Although the problem of shoot vitrification occurred on this medium but this was overcome by transferring the vitrified shoots on MS medium supplemented with 1 mg/l BA and 0.25 mg/l Kn. The same medium was found to be the best medium for further in vitro shoot multiplication. 100 % root induction from in vitro grown shoots was obtained on half strength MS medium supplemented with 1 mg/l IBA. In vitro formed plantlets were hardened and transferred to soil with 83 % survival. Additionally, conservation of in vitro multiplying shoots was also attempted using two different approaches namely slowing down the growth at low temperature and cryopreservation following vitrification. At low temperature retrieval rate was better at 10 °C than at 4 °C after conservation of in vitro multiplying shoots. In cryopreservation-vitrification studies, the vitrified shoot buds gave maximum retrieval of 41.66 % when they were precooled at 4 °C, while only 16.66 % vitrified shoots were retrieved from those precooled at 10 °C. Genetic stability of the in vitro grown plants was analysed by RAPD and ISSR markers which indicated no somaclonal variation among in vitro grown plants demonstrating the feasibility of using the protocol without any adverse genetical effects.

Somatic embryogenesis and plant regeneration from leaf explants of ...

African Journals Online (AJOL)

An attempt was made to study the somatic embryogenesis and plant regeneration from the in vitro leaf explants of Rumex vesicarius L. a renowned medicinal plant, which belongs to polygonaceae family. Effective in vitro regeneration of R. vesicarius was achieved via young leaf derived somatic embryo cultures.

In vitro plant propagation and crop improvement in Lisianthus (Lisianthus Russelianus Hook.

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Rodica Pop

2016-11-01

Full Text Available Romania assists at the present time to an increase of production crops for ornamental plants and as a consequence an increased demand of planting material. Thus, improvements of the current multiplication methods are sought after. Lisianthus russelianus Hook. (Eustoma grandiflorum Grise. is a relatively new floral crop to the international market, known for beautiful flowers of various colors and for having a long vase life. This study focused on the development of an efficient protocol for rapid regeneration of this species following known basic and applied aspects of lisianthus biotechnology but exploring new potentials. In the course of experiments conducted, for in vitro multiplication there were used nodal segments (1.5 cm with axillary buds from three F1 hybrids ‘Echo Lavender’, ‘Flamenco White’, ‘Mirage Pastel Pink’ that were inoculated on MS basal medium supplemented with 0.50 mg 1-1 TDZ, 1.0 mg 1-1 BAP and 0.50 mg 1-1 AIA. The results show that the medium with BAP was most effective for obtaining the highest shoots number compared to medium containing TDZ. For rooting induction, two different concentrations of auxin IBA 0.5 mg 1-1 and 1.5 mg 1-1 were used simultaneously on MS basal medium. The highest roots number occurred when using 1.5 mg 1-1 IBA. Both the number of shoots and rooting regeneration were dependent on the cultivar. The highest shoots number was achieved for ’Mirage Pastel Pink’ hybrid (6.91 on the medium containing 1.0 mg 1-1 BAP and 0.50 mg 1-1 IAA.

In vitro regeneration of selected commercial Tanzanian open ...

African Journals Online (AJOL)

Using this regeneration system Situka M-1, Staha and TMV-1 can now be improved against various production constraints through genetic engineering. Key words: 2, 4-dichlorophexyacetic acid, embryogenic callus, immature zygotic embryos, regeneration frequency, somatic embryos, Tanzanian open pollinated maize.

Biolistic transformation of Carrizo citrange (Citrus sinensis Osb. × Poncirus trifoliata L. Raf.).

Science.gov (United States)

Wu, Hao; Acanda, Yosvanis; Jia, Hongge; Wang, Nian; Zale, Janice

2016-09-01

The development of transgenic citrus plants by the biolistic method. A protocol for the biolistic transformation of epicotyl explants and transgenic shoot regeneration of immature citrange rootstock, cv. Carrizo (Citrus sinensis Osb. × Poncirus trifoliata L. Raf.) and plant regeneration is described. Immature epicotyl explants were bombarded with a vector containing the nptII selectable marker and the gfp reporter. The number of independent, stably transformed tissues/total number of explants, recorded by monitoring GFP fluorescence 4 weeks after bombardment was substantial at 18.4 %, and some fluorescing tissues regenerated into shoots. Fluorescing GFP, putative transgenic shoots were micro-grafted onto immature Carrizo rootstocks in vitro, confirmed by PCR amplification of nptII and gfp coding regions, followed by secondary grafting onto older rootstocks grown in soil. Southern blot analysis indicated that all the fluorescing shoots were transgenic. Multiple and single copies of nptII integrations were confirmed in five regenerated transgenic lines. There is potential to develop a higher throughput biolistics transformation system by optimizing the tissue culture medium to improve shoot regeneration and narrowing the window for plant sampling. This system will be appropriate for transformation with minimal cassettes.

Development of bunchy top virus resistant banana cv lakatan in vitro culture and radiation technology

International Nuclear Information System (INIS)

Estrella, J.D.; Caymo, L.S.; Dizon, T.O.; Dela Cruz, F. Jr; Damasco, O.P.

2002-01-01

Bunchy to virus (BTV) is the most destructive virus disease of banana in the Philippines. Incorporation of resistance to this virus disease by conventional hybridization is not possible due to male and female sterility of most commercial banana cultivars. In vitro culture coupled with radiation technology can be used to develop BTV resistance in banana cv. Lakatan. The sensitivity of banana shot tip explants to gamma irradiation was determined by subjecting the shoot tips to varying doses (5, 10, 20, 25, 30, 40, 60, 80 and 100 Gy) of irradiation. The LD sub 50 for banana shoot tips determined by 50% reduction in growth and shoot proliferation, was observed to around 20-25 Gy. Bulk irradiation of shoot tip explants was conducted using 20-25 Gy. Irradiated cultures were multiplied for 3-5 cycles and plants regenerated were potted out and screened for BTV resistance. A total of 3,447 irradiated plants regenerated from the radiosensitivity experiment (1,847 plants) and bulk irradiation of 20/25 Gy (1,600 plants) were screened for BTV resistance in the greenhouse using artificial BTV inoculation using the aphid vector Pentalonia nigronervosa. One hundred eighteen plants or 3.4% (118/3,447) of the artificially irradiated plants showed seedling resistance after 4-7 months of evaluation. These plants were planted in the field and were subjected to natural BTV infection. To date, 85 (out of the 118) putative seedling resistant plants continuously expressed BTV resistance in the field after more than 10 months of evaluation. The absence of BTV infection in 39 putative resistant plants was confirmed by ELISA test. Suckers from selected putative resistance plants will be collected, propagated and evaluated for the second cycle stability of BTV resistance and detailed characterization of important horticultural traits

Bone regeneration based on nano-hydroxyapatite and hydroxyapatite/chitosan nanocomposites: an in vitro and in vivo comparative study

International Nuclear Information System (INIS)

Tavakol, S.; Nikpour, M. R.; Amani, A.; Soltani, M.; Rabiee, S. M.; Rezayat, S. M.; Chen, P.; Jahanshahi, M.

2013-01-01

Surface morphology, surface wettability, and size distribution of biomaterials affect their in vitro and in vivo bone regeneration potential. Since nano-hydroxyapatite has a great chemical and structural similarity to natural bone and dental tissues, incorporated biomaterial of such products could improve bioactivity and bone bonding ability. In this research, nano-hydroxyapatite (23 ± 0.09 nm) and its composites with variety of chitosan content [2, 4, and 6 g (45 ± 0.19, 32 ± 0.12, and 28 ± 0.12 nm, respectively)] were prepared via an in situ hybridization route. Size distribution of the particles, protein adsorption, and calcium deposition of powders by the osteoblast cells, gene expression and percentage of new bone formation area were investigated. The highest degree of bone regeneration potential was observed in nano-hydroxyapatite powder, while the bone regeneration was lowest in nano-hydroxyapatite with 6 g of chitosan. Regarding these data, suitable size distribution next to size distribution of hydroxyapatite in bone, smaller size, higher wettability, lower surface roughness of the nano-hydroxyapatite particles and homogeneity in surface resulted in higher protein adsorption, cell differentiation and percentage of bone formation area. Results obtained from in vivo and in vitro tests confirmed the role of surface morphology, surface wettability, mean size and size distribution of biomaterial besides surface chemistry as a temporary bone substitute.

In Vitro and In Vivo Study of a Novel Porcine Collagen Membrane for Guided Bone Regeneration

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Eisner Salamanca

2016-11-01

Full Text Available For years, in order to improve bone regeneration and prevent the need of a second stage surgery to remove non-resorbable membranes, biological absorbable membranes have gradually been developed and applied in guided tissue regeneration (GTR. The present study’s main objective was to achieve space maintenance and bone regeneration using a new freeze-dried developed porcine collagen membrane, and compare it with an already commercial collagen membrane, when both were used with a bovine xenograft in prepared alveolar ridge bone defects. Prior to surgery, the membrane’s vitality analysis showed statistically significant higher cell proliferation in the test membrane over the commercial one. In six beagle dogs, commercial bone xenograft was packed in lateral ridge bone defects prepared in the left and right side and then covered with test porcine collagen membrane or commercial collagen membrane. Alveolar height changes were measured. Histomorphometric results, in vitro and in vivo properties indicated that the new porcine collagen membrane is biocompatible, enhances bone xenograft osteoconduction, and reduces the alveolar ridge height reabsorption rate.

Direct organogenesis of seaside heliotrope (Heliotropium crassavicum) using stem explants.

Science.gov (United States)

Satyavani, K; Dheepak, V; Gurudeeban, S; Ramanathan, T

2013-10-15

Heliotropium crassavicum L. is a sand binder salt marsh herb with enormous traditional value and widely found in South Asia America and Europe. In the direct method of regeneration from stem explants, we observed the maximum number of shoot regeneration after four weeks culture of MS elongation medium with 2.0 mg L(-1) of 2, 4-D (17.27 +/- 0.51). It was clear that MS medium with 2.0 mg mL(-1) 2, 4-D alone suitable for shoot multiplication as well as shoot elongation then compared to other combination of auxin and cytokinin. In vitro shoots were excised from shoot clumps and transferred to rooting medium containing 2, 4-dichlorophenoxy acetic acid (0.5-3.0 mg L(-1)). The maximum number of root regeneration (6.4 +/- 0.416) and root length (6.08 +/- 0.07) were observed in MS rooting medium fortified with 2.5 mg L(-1) of 2, 4-D after 2 weeks of culture. 85% of in vitro raised plantlets with well-developed shoots and roots were transferred to ex vivo conditions into polythene bag containing sterile compost with ratio (v/v/v) of organic fertilizer: sand: peat (1:2:2; 3:1:0 or 2:2:1). Sixty five percent of acclimated plants were transferred to the pots under full sun where they grew well without any detectable phenotypic variations.

Micropropagation of Ajuga bracteosa, a medicinal herb

OpenAIRE

Kaul, Shivanee; Das, Sandip; Srivastava, P. S.

2013-01-01

For conservation and genetic transformation, a successful in vitro micropropagation protocol for Ajuga bracteosa, a medicinal herb has been established for the first time. MS medium supplemented with IAA (2 mg/L) and BA (5 mg/L) induced 100 % shoot regeneration with an average of 41.4 shoots of 8.4 cm per culture. Excised in vitro shoots when transferred to MS + IBA (0.5 mg/L) produced 20 roots/shoot of 20.2 cm average length in 100 % cultures. Of the three explants, leaf, petiole and root, l...

IN VITRO PROPAGATION OF DENDROBIUM AND PHALAENOPSIS THROUGH TISSUE CULTURE FOR CONSERVATION

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Lita Soetopo

2012-06-01

Full Text Available The studies were focused on developing an efficient and effective propagation protocol for orchid species from genera Dendrobioum and Phalaenopsis through tissue culture. The Materials used were explants from adventive shoot tip, floral stalk buds and PLBs derived from seeds. The results indicated growth and development of adventive shoot tip explants of Dendrobium: a high survival percentage for explant with green color was shown by D. racianum, followed by D. laxiflorum, D. pseudo-conantum, D. strebloceras, D. lineale, and D. veratrifolium. However, plantlets regeneration occurred only on D. pseudoconantum, and D. strebloceras. Explant regeneration from seed derived protocorm-like bodies on D. spectabile occurred 40 days after inoculation transfer and subculture. High survival percentage of explant from floral stalk shoot was shown by P. amabilis. There were several plantlets surviving in acclimatisation. Explant regeneration from seed derived from protocorm-like bodies on P. hieroglypha occurred 40 days after inoculation and subculture. It was suggested that for ex situ conservation on certain species of Dendrobium and Phalaenopsis in the category of rare germplasms, tissue culture could be applied effectively and efficiently by using explant from adventive shoot tip, floral stalk buds and seed derived protocorm-like body explant for vegetative seed multiplication.

Short communication. N-(2-chloro-4-pyridyl)-N-phenylurea (4-CPPU) enhances in vitro direct shoot organogenesis of Citrus aurantium L. epicotyl segments compared to other commonly used cytokinins

Energy Technology Data Exchange (ETDEWEB)

Roussos, P. A.; Dimitriou, G.; Voloudakis, A. E.

2011-07-01

The effect of three concentrations of five different cytokinins, i.e. 6-benzylamino purine, 2-isopentyl adenine, kinetin (Kin), thidiazuron and N-(2-chloro-4-pyridyl)-N-phenylurea (4-CPPU), was evaluated on the in vitro direct shoot organogenesis of epicotyl explants of sour orange (Citrus aurantium L.). The basal medium used was that of Murashige and Tucker and epicotyl explants were incubated in medium supplemented with the prementioned cytokinins for 45 days. The addition of Kin and 4-CPPU in the medium enhanced the direct shoot organogenesis of sour orange epicotyl segments. The concentration of each of these two cytokinins which gave the best results, was combined with indole-3-acetic acid (IAA) or {alpha}-naphthalene acetic acid ({alpha}-NAA) at concentrations ranging from 0.01 mg L{sup -}1 to 0.2 mg L{sup -}1. The inclusion of IAA at 0.2 mg L{sup -}1 in the medium with 4-CPPU at 0.05 mg L{sup -}1 resulted in 100% successful direct shoot organogenesis, while the combination of Kin at 0.25 mg L{sup -}1 with IAA or {alpha}-NAA each at 0.01 mg L{sup -}1 presented equally high organogenesis percentages (91.7%). The incubation of the produced shoots, in medium supplemented with either indole-3-butyric acid or {alpha}-NAA resulted in high rooting percentages (up to 90%) and the rooted explants were successfully acclimatized under mist (85%). Although 4-CPPU has been used in in vitro culture of various species, this is the first report on its use in the direct shoot organogenesis of citrus species and could be of great value in citrus genetic transformation protocols using epicotyl segments, since this cytokinin resulted in the absolute organogenesis percentage. (Author) 20 refs.

Embriogênese somática e regeneração de plantas a partir de embrião maduro de aveia Somatic embryogenesis and plant regeneration derived from mature embryos of oat

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Caren Regina Cavichioli Lamb

2002-02-01

Full Text Available Calo embriogênico tem sido o tecido-alvo mais utilizado para transformação genética de cereais. O objetivo deste trabalho foi investigar o estabelecimento de calos embriogênicos e a regeneração de plantas in vitro a partir de embriões maduros de genótipos de aveia (Avena sativa L.. Embriões maduros foram retirados das sementes e colocados em meio MS (Murashige & Skoog, contendo 30,0 g L-1 de sacarose e 2,0 mg L-1 de ácido 2,4-diclorofenoxiacético (2,4-D. Após o período de indução de calos, agregados embriogênicos foram isolados e subcultivados a cada 21 dias para meio fresco. Os calos embriogênicos foram então transferidos para meio de indução de parte aérea, e, na seqüência, as partes aéreas foram transferidas para meio de indução de raízes. Houve diferenças entre genótipos quanto à capacidade de embriogênese somática e regeneração de plantas in vitro a partir de embrião maduro. Este explante permitiu a indução de calos embriogênicos, que se multiplicaram, e que regeneraram in vitro um grande número de plantas de genótipos como UFRGS 7 e UFRGS 19, o que o faz passível de ser utilizado na transformação genética da aveia.Embryogenic callus has been the most used target tissue for cereal genetic transformation. Therefore, the objective of this study was to investigate the establishment of embryogenic calli and the in vitro plant regeneration from mature embryos of oat genotypes (Avena sativa L.. Mature embryos were taken out of the seeds and placed on a culture medium MS (Murashige & Skoog, containing 30,0 mg L-1 of sucrose and 2,0 mg L-1 of 2,4-dichlorophenoxyacetic acid (2,4-D. From the induction period, embryogenic aggregates were isolated and subcultivated each 21 days into a fresh medium. After this period, embryogenic calli were transferred to a medium for shoot regeneration. Subsequently, the shoot was transferred to a medium for root induction. There was variability among genotypes for somatic

Arabidopsis brassinosteroid biosynthetic mutant dwarf7-1 exhibits slower rates of cell division and shoot induction

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Schulz Burkhard

2010-12-01

Full Text Available Abstract Background Plant growth depends on both cell division and cell expansion. Plant hormones, including brassinosteroids (BRs, are central to the control of these two cellular processes. Despite clear evidence that BRs regulate cell elongation, their roles in cell division have remained elusive. Results Here, we report results emphasizing the importance of BRs in cell division. An Arabidopsis BR biosynthetic mutant, dwarf7-1, displayed various characteristics attributable to slower cell division rates. We found that the DWARF4 gene which encodes for an enzyme catalyzing a rate-determining step in the BR biosynthetic pathways, is highly expressed in the actively dividing callus, suggesting that BR biosynthesis is necessary for dividing cells. Furthermore, dwf7-1 showed noticeably slower rates of callus growth and shoot induction relative to wild-type control. Flow cytometric analyses of the nuclei derived from either calli or intact roots revealed that the cell division index, which was represented as the ratio of cells at the G2/M vs. G1 phases, was smaller in dwf7-1 plants. Finally, we found that the expression levels of the genes involved in cell division and shoot induction, such as PROLIFERATING CELL NUCLEAR ANTIGEN2 (PCNA2 and ENHANCER OF SHOOT REGENERATION2 (ESR2, were also lower in dwf7-1 as compared with wild type. Conclusions Taken together, results of callus induction, shoot regeneration, flow cytometry, and semi-quantitative RT-PCR analysis suggest that BRs play important roles in both cell division and cell differentiation in Arabidopsis.

Plant growth regulator interactions in physiological processes for controlling plant regeneration and in vitro development of Tulbaghia simmleri

Czech Academy of Sciences Publication Activity Database

Kumari, A.; Baskaran, P.; Plačková, Lenka; Omámiková, Hana; Nisler, Jaroslav; Doležal, Karel; Van Staden, J.

2018-01-01

Roč. 223, APR (2018), s. 65-71 ISSN 0176-1617 R&D Projects: GA MŠk(CZ) LO1204 Institutional support: RVO:61389030 Keywords : Endogenous hormone * Exogenous hormone application * In vitro regeneration * Ornamental and medicinal plant * Physiological process * Tulbaghia simmleri Subject RIV: EB - Genetics ; Molecular Biology OBOR OECD: Plant sciences, botany Impact factor: 3.121, year: 2016

In vitro flowering and fruiting in culture of dendrobium officinate kimura et migo (orchidaceae)

International Nuclear Information System (INIS)

Qian, X.; Wang, C.; Ouyang, T.; Tian, M.

2014-01-01

Dendrobium officinate Kimura et Migo. is a rare and endangered perennial orchid, which is valued for its attractive flowers and medicinal uses. Its three-to-five-year breeding cycle makes propagation difficult, so we investigated In vitro production of flowers and seeds for this species. Calluses were induced from shoot-tip explants of D. officinate on Murashige and Skoog (MS) medium with 0.2 mg L-1 benzyladenine (BA) and 0.05 mg L-1 naphthalene acetic acid (NAA). Multiple shoots were regenerated after protocorm-like bodies (PLBs) were transferred onto the same basal medium with 1.0 mg L-1 NAA. The plantlets, 2-4 cm in height, maintained In vitro were induced to flower, and the highest rates of inflorescence (83.2%) and normal flowers (73.6%) were produced on the MS medium with 15% (v/v) coconut water and 0.1 mg L-1 thidiazuron (TDZ) within 9 weeks. Histological analysis showed the origin of the floral primordial and normal morphologies of the pollen and female organs. The normal flowers were artificially pollinated and capsules developed. Viable seeds were produced and most of them germinated on the MS medium. This reproduction system could be used to study the molecular basis of flowering or to improve plant breeding programs. (author)

Bioactive Sr(II/Chitosan/Poly(ε-caprolactone Scaffolds for Craniofacial Tissue Regeneration. In Vitro and In Vivo Behavior

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Itzia Rodríguez-Méndez

2018-03-01

Full Text Available In craniofacial tissue regeneration, the current gold standard treatment is autologous bone grafting, however, it presents some disadvantages. Although new alternatives have emerged there is still an urgent demand of biodegradable scaffolds to act as extracellular matrix in the regeneration process. A potentially useful element in bone regeneration is strontium. It is known to promote stimulation of osteoblasts while inhibiting osteoclasts resorption, leading to neoformed bone. The present paper reports the preparation and characterization of strontium (Sr containing hybrid scaffolds formed by a matrix of ionically cross-linked chitosan and microparticles of poly(ε-caprolactone (PCL. These scaffolds of relatively facile fabrication were seeded with osteoblast-like cells (MG-63 and human bone marrow mesenchymal stem cells (hBMSCs for application in craniofacial tissue regeneration. Membrane scaffolds were prepared using chitosan:PCL ratios of 1:2 and 1:1 and 5 wt % Sr salts. Characterization was performed addressing physico-chemical properties, swelling behavior, in vitro biological performance and in vivo biocompatibility. Overall, the composition, microstructure and swelling degree (≈245% of scaffolds combine with the adequate dimensional stability, lack of toxicity, osteogenic activity in MG-63 cells and hBMSCs, along with the in vivo biocompatibility in rats allow considering this system as a promising biomaterial for the treatment of craniofacial tissue regeneration.

Ectopic expression of class 1 KNOX genes induce and adventitious shoot regeneration and alter growth and development of tobacco (Nicotiana tabacum L) and European plum (Prunus domestica L)

Science.gov (United States)

Transgenic plants of tobacco (Nicotiana tabacum L) and plum (Prunus domestica L) were produced by transforming with apple class 1 KNOX genes (MdKN1 and MdKN2) or corn KN1 gene. Transgenic tobacco plants were regenerated in vitro from transformed leaf discs cultured in a tissue medium lacking cytoki...

Insulin and IGF-II, but not IGF-I, stimulate the in vitro regeneration of adult frog sciatic sensory axons

DEFF Research Database (Denmark)

Edbladh, M; Svenningsen, Åsa Fex; Ekström, P A

1994-01-01

We used the in vitro regenerating frog sciatic nerve to look for effects of insulin and insulin-like growth factors I and II (IGF-I, IGF-II) on regeneration of sensory axons and on injury induced support cell proliferation in the outgrowth region. In nerves cultured for 11 days, a physiological...... dose (10 ng/ml, approximately 2 nM) of insulin or IGF-II increased ganglionic protein synthesis (by 20% and 50%, respectively) as well as the level of newly formed, radiolabelled axonal material distal to a crush injury (both by 80%), compared to untreated, paired controls. In addition, insulin...... increased the outgrowth distance of the furthest regenerating sensory axons by 10%. The preparation was particularly sensitive to insulin during the first 5 days of culturing. Furthermore, both insulin and IGF-II were found to inhibit proliferation of support cells in the outgrowth region in a manner...

C4 photosynthesis in Euphorbia degeneri and E. remyi: a comparison of photosynthetic carbon metabolism in leaves, callus cultures and regenerated plants

International Nuclear Information System (INIS)

Ruzin, S.E.

1984-04-01

Based on analysis of 14 CO 2 fixation kinetics and assays of enzymes related to C 4 metabolism (NAD-ME, NADP-ME, NAD-MDH, NADP-MDH, AST, ALT), leaves and regenerated plants of Euphorbia degeneri exhibit a modified NADP-ME-type photosynthesis. Apparently, both aspartate and malate are used for transport of CO 2 to bundle sheath cells. Callus grown on either non-shoot-forming or shoot-forming media fixes CO 2 into RPP-cycle intermediates and sucrose, as well as malate and aspartate. 14 CO 2 pulse/chase kinetics show no significant loss of label from C 4 acids throughout a one minute chase. Analysis of PEPCase revealed the presence of 2 isoenzymes in both leaf and regenerated plant tissues (K/sub m/ [PEP] = 0.080 and 0.550) but only one isoenzyme in callus (K/sub m/ = 0.100). It appears that C 4 photosynthesis does not occur in callus derived from this C 4 dicot but is regenerated concomitant with shoot regeneration, and β-carboxylation of PEP in callus, mediated by the low K/sub m/ isoenzyme of PEPCase, produces C 4 acids that are not involved in the CO 2 shuttle mechanism characteristic of C 4 photosynthesis. 161 references, 19 figures, 12 tables

Effect of γ-ray irradiation on in vitro culture and plant regeneration of alfalfa

International Nuclear Information System (INIS)

Zhang Xiaodong; Lin Tingan

1992-01-01

60 Co γ-ray irradiation ranged 0-16 kR was used to treat the cotyledons and hypocotyls of 5 cultivars of a alfalfa (Medicago sativa L.). The effects of irradiation on the frequency of callus, fresh weight of callus, the frequency of somatic embryo induction and plantlet regeneration were studied. The results showed as follows: the radiosensitivities of cotyledon was significantly higher than that of hypocotyl. Exposure of 2 kR could improve the growth of callus, embryogenesis and plantlet regeneration at various levels. Exposure of 12 kR completely inhibited the callus formation from explants of cotyledon. Callus from two cultivars, England 648 and Jining alfalfa, were irradiated with 0-8 kR. The results showed that exposure below 2 kR could promote the growth of callus and the formation of somatic embryo at different levels. The exposure of 8 kR had the effect of lethal. The optimum exposure for genetic improvement of alfalfa in vitro by inducing mutation was considered to be 4-6 kR