High Concentrations Of Kinetin Research Articles

In vitro Barnardia japonica (Thunb.) Schult. et Schult. fil. micropropagation by direct regeneration

In vitro micropropagation of rare Far Eastern species Barnardia japonica by the direct regeneration method was studied for the first time. The effect of different concentrations of 6-benzylaminopurine and kinetin in combination with auxins on the regeneration potential of young seedlings using Murashige and Skoog media was estimated. It was found that the largest number of new plants (61 new shoots on average) is formed when explants are cultivated on the medium with a high concentration of kinetin (10 mg/L) and a minimum of α-naphthylacetic acid (0.1 mg/L). The resulting plants are rooted easily on a hormone-free medium of the same mineral composition. The least effective for the mass production of plants by direct regeneration was a medium with a combination of 2 mg/L kinetin and 1 mg/L α-naphthylacetic acid. The developed approach can be applied for the effective propagation of Barnardia japonica and returning the species to its natural habitat.

Comparative Study of In Vitro Root and Shoot Proliferation from the Node Explants of Asparagus racemosus Willd

Asparagus racemosus Willd. locally known as Kurilo or Shatavari is found throughout the tropical and subtropical regions of Nepal. It is a high value herb because of its medicinal and nutritional values which is leading this species decrease in its natural habitat. Hence, to conserve this species, an in vitro tissue culture method of its multiplication has been applied as an effective method of ex situ conservation. In the experiment, although most of the treatments induced highly insignificant number of adventitious shoots, low concentration of IBA (0.1-0.5 mg/l) in combination with relatively higher concentrations of Kinetin (1.0-2.0 mg/l) in MS medium were found to be significantly inducing up to 8.33±1.308 shoots/node. Among the treatments where hormones were used singly, IAA 0.5 and Kinetin 1.0 induced 4.83±1.08 and 4.66±1.43 shots/node respectively. Hence, a protocol for the effective multiplication of this species has been developed which can be used accordingly.

Media composition affects seed dormancy, apical dominance and phenolic profile of Knautia sarajevensis (Dipsacaceae), Bosnian endemic

AbstractKnautia sarajevensisis an endemic plant of the Dinaric Alps and is mainly distributed on Bosnian Mountains. Due to the quite large flower heads and easy maintenance, this plant has a potential use as a substitute ornamental plant forK. arvensisin perennial beds. The current study evaluated the germination process in different treatments in an attempt to suppress dormancy and increase germination rate, and to develop a successful protocol for micropropagation. An over 60% germination rate was achieved through cultivation of seeds on MS basal medium with reduced mineral nutrient composition and the absence of sucrose. On the other hand, a below 10% germination rate was achieved with untreated seeds. Suppression of apical dominance was achieved through application of high concentrations of kinetin, apical shoot decapitation or cultivation of shoots in liquid media. Overall, liquid cultures were more successful as a micropropagation system for this plant. Shoots spontaneously developed roots on multiplication treatments and were successfully acclimatized. Moreover, phenolic compound profile was analysed in the light of the possible medicinal potential of this plant. Variable amounts of total phenolic compounds as well as individual phenolics were recorded, according to treatment and solidification of media. An increase in rosmarinic acid content was reported for kinetin treatments and acclimatized plants comparing to mother plants in natural habitat. The present study shows that choice of cytokinin concentration, explant type as well as culture type influences not only shoot proliferation and apical dominance suppression but alsoin vitroproduction of phenolics.

Effects of light and cytokinin on in vitro micropropagation and microrhizome production in turmeric (Curcuma longa L.)

Using clonal in vitro propagated plantlets of Curcuma longa, the effects of light and cytokinin on micropropagation and microrhizome formation were investigated. The results showed the promotive effects of low light intensity for micropropagation and of short day condition for well-developed microrhizome induction. In the tissue of well-developed microrhizomes, differentiation of yellowish-pigmented cells similar to the secretory cells of rhizome of the mother plant was observed. Staining of the cells with nile red suggested the accumulation of essential oil. Rapidly growing callus was isolated from the root tip of micropropagated plantlets which retained totipotency during subculture. High concentration of kinetin induced microrhizome formation in this callus. Yellow pigmented cells were also observed in the tissue of microrhizome, although far less frequency.

In Vitro Regeneration of Four Commercial Cotton Cultivars ( Gossypium hirsutum L.) Grown in Xinjiang, China

Genetic improvement of cotton through biotechnology has been limited by lack of an efficient regeneration system. An efficient somatic embryo production and maturation procedure was thus developed to regenerate plantlets from hypocotyls of cotton cultivars Xinluzhong 20, Xinluzao 24, Xinluzao 33, and 03298 grown in Xinjiang. Calli were effectively produced by 0.01 to 0.10 mg L −1 kinetin (KT) and 0.10 mg L −1 2,4-dichlorophenoxyacetic acid (2,4-D), with a better result by 0.02 or 0.10 mg L −1 KT and 0.10 mg L −1 2,4-D. Split hypocotyl segments and double amounts of KNO 3 during induction of calli were beneficial to the emerge of embryogenic calli. Embryogenic calli and globular-stage somatic embryos were effectively initiated with high concentration of KT and low concentration of 2,4-D in ECM media (0.05 or 0.10 mg L −1 KT and 0.01 mg L −1 2,4-D). Embryos were further developed into plantlets in MSBF medium under the conditions of dehydration and ventilation, which were achieved using filter paper on medium and cotton tampon sealing flask. Using this protocol, normal plantlets with strong roots were developed from these cotton cultivars in 6 to 8 months. The successful regeneration protocol established in this study can be used to improve cotton cultivars by genetic engineering.

Effects of tissue culture conditions during embryoid multiplication on the incidence of "mantled" flowering in clonally propagated oil palm.

New and established embryoid lines of oil palm (Elaeis guineensis Jacq.) clones, classified with regard to the expected risk of mantled flowering, were included in experiments to determine to what extent culture conditions during 48 weeks of embryoid multiplication could influence the incidence and severity of mantled flowering. Variables tested included clone risk status, duration of transfer interval, and plant growth regulator composition of the culture medium. Clones classified as high risk at the outset gave a significantly higher incidence of mantled flowering in the field than clones classified as low/medium risk, confirming that data on terminal inflorescences produced in vitro allows effective screening of material with regard to the risk of mantled flowering. The incidence of mantled flowering ranged from 0 to 100% depending on risk status of the clones, and the treatments applied. In general, a relatively long transfer interval (8 weeks) and a high auxin/cytokinin ratio resulted in the lowest incidence of mantled flowering with all types of material. Reducing the transfer interval down to 2 or 4 weeks, and/or using media with relatively high concentrations of kinetin (0.25 mg l−1), and low concentrations of naphthalene acetic acid (0 or 0.1 mg l−1), resulted in a high incidence of mantled flowering (above 30%), even with low/medium risk clones. Exclusion of plant growth regulators from the embryoid multiplication medium did not prevent some mantled flowering (12%) of new embryoid lines of low/medium risk clones following embryoid multiplication on a 4 week transfer interval, and a much higher incidence of mantling (above 90%) of lines from high risk clones.

A simple and efficient procedure to improve plant regeneration from protoplasts isolated from long-term cell-suspension cultures of indica rice

A simple and efficient method has been developed to improve plant regeneration from protoplasts isolated from >1-yr-old cell-suspension culture of Indica rice (Oryza sativa) cv. Pusa Basmati 1. A two-step regeneration procedure, involving the transfer of calluses to shoot-regeneration medium containing 1% (w/v) agarose prior to culture on a medium containing 0.4% (w/v) agarose, was found to improve plant regeneration. High concentrations of kinetin in the regeneration medium were also found to be beneficial. The two-step regeneration procedure, combined with a high concentration of kinetin (10.0 mgl−1) in the medium, significantly increased plant regeneration. By this method, even though protoplasts were isolated from over 1-yr-old cell-suspension cultures, protoplast-derived plant regeneration frequency reached 16.1% compared with <4% regeneration frequency without such treatment. Use of a similar protocol might improve plant regeneration from other plant species, especially recalcitrant species.

Production of ginseng saponin and polysaccharide by cell cultures ofPanax notoginseng andPanax ginseng

The effects of various combinations of the two kinds of phytohormones, auxin and cytokinin, on cell growth and production of ginseng saponin and polysaccharide were investigated in suspension cultures of Panax notoginseng. It was found that a high concentration of kinetin (KT) (7 mg/L) seriously inhibited cell growth, but that of benzyl adenine (BA) did not. Under 0.7 mg/L of cytokinin (i.e., KT and BA), 2,4-dichlorophenoxy acetic acid (2,4-D) at 0.2 mg/L was better for the cell cultures than that at 2 or 20 mg/L; and for both naphthalene acetic acid (NAA) and indole acetic acid (IAA), 20 mg/L was their best level for the cell cultures. The highest cell concentration of 11.9 g/L (by dry wt) was obtained with the combination of 0.2 mg/L of 2,4-D and 0.7 mg/L of BA. The highest saponin content of 13.9% was achieved under 2.0 mg/L IAA and 0.07 mg/L KT; its highest production, i.e., 1.13 g/L, was obtained at 0.2 mg/L of 2,4-D and 0.7 mg/L of KT. Under 20 mg/L NAA and 0.7 mg/L KT, the highest polysaccharide content and production were reached, i.e., 16.4% and 1.86 g/L, respectively. In this work, the effects of phytohormones on P. ginseng cell cultures were also studied. A high saponin production of 1.78 g/L was observed at 10 mg/L of indole butyric acid and 0.1 mg/L of BA, and the highest production of polysaccharide (1.95 g/L) was reached with the combination of 10 mg/L NAA and 0.1 mg/L KT.

Agrobacterium rhizogenes-mediated transformation and plant regeneration of four Gentiana species

Shoots of micropropagated Gentiana acaulis, G. cruciata, G. lutea, and G. purpurea were inoculated with suspensions of Agrobacterium rhizogenes cells, strains ATCC 15834 or A4M70GUS. Adventitious roots appeared at the sites of inoculation in all 4 species. Root tips were excised and cultured on growth regulator-free media for 2-6 years. They exhibited very high branching and plagiotropism. Spontaneous bud initiation occurred in roots of G. cruciata. Roots of G. lutea, G. acaulis and G. purpurea were cultured on media with high kinetin concentration, which induced the formation of friable callus tissues. Only in G. purpurea were these calluses organogenic. Regenerated shoots of G. cruciata and G. purpurea gave rise to plants, that displayed the typical phenotypes of A. rhizogenes-transformed plants: short internodes and rolled leaves. In the roots of G. acaulis and G. cruciata, transformed with A. rhizogenes A4M70GUS, a positive reaction with X-gluc indicated the activity of β-glucuronidase. The DNA extracted from hairy roots and from the roots of transgenic plants hybridized with the appropriate genomic probes in Southern blotting. This is taken as evidence of the stable genetic transformation in the 4 Gentiana species.

The Effects of ABA and Kinetin on the Developmental Period of Drosophila melanogaster

The effects of the plant growth regulators, ABA and kinetin, on the fecundity, number of pupae and mature young offspring female, and the viability of D. melanogaster were tested. For this reason, various concentrations of ABA and kinetin (10-3 M, 10-4 M and 10-5 M) were added into the standard food-medium of Drosophila. The imagoes female which fed on the hormoned food-medium for the first three days of their adult life were used. Experimental results show that ABA and low concentrations of kinetin (10-5 M) inhibit fecundity and the number of pupae and mature young adults; however, higher concentrations of kinetin (10-3 M) stimulates these factors.

Efficient culture method for production of plantlets from mechanically cut horseradish hairy roots

Effects of the initial density of roots fragmented by blender cutting and the addition of kinetin on plantlet formation were studied. The number of plantlets increased with an increasing initial density of root fragments (1.7 g-f.w./ l to 13.3 g-f.w./ l) at 0.1 mg/ l of kinetin, but the number of plantlets with elongated roots unavailable for use as artificial seed dramatically increased after kinetin depletion in the medium. The condition at 0.8 mg/ l of kinetin in the medium and 13.3 g-f.w./ l of initial density of root fragments increased the number of the plantlets without roots (suitable for artificial seed) compared with the condition at 0.1 mg/ l of kinetin in the medium and 13.3 g-f.w./ l of initial density of roots. To avoid the toxic effect of a high concentration of kinetin at the beginning stage of the culture, 0.1 mg/ l of kinetin was periodically supplemented in the medium 8 times (on the initial day and from 4 d to 10 d of the culture). Under this condition, the total number of plantlets increased and reached a maximum on 12 d, and the number of plantlets 2–4 mm in size without roots (6,270 l −1) was the highest of all other cultures.

Differential alcohol dehydrogenase and malate dehydrogenase isozyme expression in long-term callus tissue cultures of Cereus peruvianus (Cactaceae).

Alcohol dehydrogenase (ADH) and mitochondrial malate dehydrogenase (mMDH) isozymes were tested as markers to study the effect of a high kinetin concentration on isozyme phenotypes and on the development of Cereus peruvianus callus tissue culture. Three-year-old callus tissues were used as samples. Callus tissue samples grown on 4.0 mg/L 2,4-dichlorophenoxyacetic acid (2,4-D) and on 4.0 and 8.0 mg/L N-(2-furanylmethyl)-1H-purine-6 amine (kinetin) were cut and transferred to fresh medium containing 4.0 mg/L 2,4-D and 4.0, 8.0, 16.0, and 32 mg/L kinetin combinations. The pattern of changes observed in the ADH and mMDH isozymes as well as the growth of callus tissues was independent of the concentrations tested. The various ADH and mMDH isozymes seem to be products of differential association of subunits of the two Adh and two mMdh genes. Both genes are active throughout callus tissue development; however, gene expression changed with various callus culture condictions. This study addresses how long-term callus culture condictions affect constitutive and differential gene expression of the Adh and mMdh genes in C. peruvianus.

Lignin release and photomixotrophism in suspension cultures of Picea abies

The effect of different concentrations of sucrose (0‐4%) and of two growth regulators (0–50 μM 2,4‐D and 0–25 μM kinetin) was tested on growth and chlorophyll content of suspension cultures of Picea abies (L.) Karst. originating from chlorophyllous embryo callus in an elevated CO2 (2%) atmosphere. A continuous chlorophyllous suspension culture was achieved on a medium containing 2% sucrose and a low level of organic nitrogen (0.25 mM arginine and 0.5 mM glutamine) supplemented with 2,4‐D (0.5 μM) and kinetin (2.5 μM). The same medium with 4% sucrose gave the best growth response, but a negative correlation between chlorophyll level and growth was observed. The chlorophyllous cultures grew slowly in a medium with low (0.5%) sucrose or without any carbohydrate source, suggesting photomixotrophism. A high concentration of kinetin inhibited both growth and chlorophyll synthesis. Release of lignin into the nutrient medium was observed in several experiments, especially in slow‐growing cultures supplemented with sucrose. Only a few successive passages of suspensions that produced lignin could be cultured before cell death. The cultures releasing lignin may be unique for studies on synthesis and biodegradation of this very complex compound.

Plant regeneration from tissue cultures of Persian buttercup (Ranunculus asiaticus L.)

Different plant explants of Persian buttercup (Ranunculus asiaticus L.) were screened for callus induction and adventitious shoot regeneration on different media to establish totipotent cultures. Murashige & Skoog (MS) medium was used, supplemented with different concentrations of the following growth regulators: kinetin, benzyladenine (BA), naphthaleneacetic acid (NAA) and indoleacetic acid (IAA). Callus was induced and adventitious buds regenerated only from cotyledonary explants after 4–5 weeks. Subculture of the regenerated buds on the same basal medium in presence of gibberellic acid (GA3) and BA produced well-organized shoots. Rooting was obtained by transferring shoots to growth regulator-free MS medium. A high rate of shoot multiplication has been achieved on medium with high concentration of kinetin and long-day photoperiod. Finally the plants were successfully transferred to soil and grown in a greenhouse.

The Essential Oil of Coleonema album (Rutaceae) and of a Photomixotrophic Cell Culture Derived thereof

Abstract The volatile shoot oil of Coleonema album was found to be composed of monoterpenes, ses­quiterpenes and phenylpropanoids. Heterotrophic and photom ixotrophic callus cultures were established from green parts of the plant. A photom ixotrophic cell line contained a heteroge­neous population of plastids and synthesized an essential oil, which differed qualitatively and quantitatively from the green parts of the intact plant. Substitution of 2,4-D and pCPA by a lower amount of NAA and high kinetin concentrations favoured the formation of essential oil. The heterotrophic callus, when grown on the same nutrient medium, was devoid of any differ­ entiated plastids and of essential oil constituents. Thus, a close correlation of light-induced plastid differentiation and essential oil synthesis was concluded.

The initiation and maintenance of callus from tubers of Colocasia esculenta and Dioscorea rotundata

An attempt was made to initiate callus from tuber segments of Dioscorea rotundata (yam) and Colocasia esculenta (taro) on six different agar media. Callus production was achieved on a medium containing Linsmaier and Skoog's salt mixture fortified with various concentrations of kinetin, napthalene acetic acid, and coconut milk. The tissues were then subcultured on media to which Kaden's vitamins were added. The subcultured tissues grew after a few weeks into masses of amber, semispherical tissue thus suggesting that high concentrations of kinetin, napthalene acetic acid indole-3-acetic acid, and coconut milk were essential for in vitro callus production from yam and taro tubers.

The role of hormones in regulating the germination of polymorphic seeds and early seedling growth of Atriplex triangularis under saline conditions

Germination of the polymorphic seeds and seedling growth of Atriplex triangularis under various salinity, gibberellic acid and kinetin treatments were determined. Gibberellic acid (GA3; 2.9 mM) promoted germination and growth at high NaCl concentrations (345 mM). Kinetin (4.7 μM) stimulated germination at all salinities and seed sizes tested. GA3 and kinetin generally increased seedling growth at all concentrations of salinity studied. Higher concentrations of kinetin were found to be inhibitory.

Effect of Culture Conditions on DOPA Accumulation in a Callus Culture ofStizolobium hassjoo

Effects of passage of subculture and several culture conditions, nutritional, hormonal and light conditions on 3,4-dihydroxyphenylalanine (DOPA) content were examined in callus of STIZOLOBIUM HASSJOO P IPER and T RACY. The favourable conditions for DOPA accumulation were found to be suboptimal conditions for growth, i.e., low concentration of phosphate, low concentration of 2,4-D, high concentration of kinetin. Light promoted DOPA accumulation. The best condition for DOPA accumulation was discussed in relation to protein synthesis.

The effect of plant growth regulators on formation of crown gall tumors on potato tuber discs

The effect of several plant growth regulators on the number of tumors developing on potato tuber discs (Solatium tuberosum L. cv. Radka) inoculated withAgrobacterium tumefaciens, strain C 58 was studied. The plant growth regulators used in appropriate range of concentrations stimulated the formation of tumors byA. tumefaciens. Naphthaleneacetic acid (NAA) was most active in concentration of 10−4 mg ml−1. Kinetin gave a biphasic response with optimal promotions of tumor initiation at 10−4 − 2 × 10−3 mg ml−1. High kinetin concentration (10−1 mg ml−1) inhibited the formation of tumors completely. Indoleacetic acid (IAA) stimulated the initiation of tumors in the same range of concentrations as kinetin, except that very high concentrations did not inhibit but enhanced tumor formation. 2,4-diehlorphenoxyacetic acid (2,4-D) showed a biphasic response with maxima in 10−4 mg ml−1 and 10−1 mg ml−1. All the tumors scored for nopaline production showed nopaline synthase activity independently whether their formation was stimulated by l0−1 mg ml−1 IAA or they were initiated without any treatment by plant growth regulators.

Regulation of in Vitro Androgenesis of Tobacco: Relationship between Concentration of Iron Ions and Kinetin

Summary Studying tobacco anthers ( Nicotiana tabacum L. cv. Samsun) cultured on the media containing 0–30 μ M FeEDTA we have recorded the development of globular embryoids only, while on the media with 40–150 μ M complete plants occured. The threshold concentration of 40 μ M, inducing transformation of the globular embryoids into plants was the same both in light and dark. Darkening of the anthers was directly proportional to FeEDTA concentration of the medium. The effect of FeEDTA was in all cases superior to that of kinetin, both at the levels of anthers and isolated embryoids. Frequency of occurrence of embryogenic anthers on the media without FeEDTA and their colour were not influenced by kinetin. The high kinetin concentration (10 mg per 1 liter medium) induced a secondary multiplication of androgenic plants. The relationship between morphogenetic effects of iron and developmental stages of embryoids, activation of their photosynthetic apparatus and respiration is discussed.