Showing papers on "Murashige and Skoog medium published in 1999"

Plant regeneration from seedling explants of Eucalyptus grandis × E. urophylla

Abstract: Hypocotyls, cotyledons, cotyledonary nodes and primary leaves were used as explants to establish a regeneration protocol for Eucalyptus grandis × E. urophylla. These seedling-derived explants were incubated on a modified MS medium (SP medium), supplemented with 2.0 μM TDZ. After 1 month, the calluses obtained were transferred to SP medium containing different concentrations of BA and NAA or zeatin and NAA. Shoots were induced from these calluses at a high frequency. Shoot elongation was then stimulated on SP medium supplemented with BA, NAA and GA3 for 20 to 30 days. For rooting, 50 mm long shoots were cultivated on root induction medium containing IBA (2.5 μM) for different periods and then transferred to the same medium but without auxin, for 30 days. Plantlets were then successfully transplanted to greenhouse conditions.

Multiple shoot regeneration and tissue culture studies on Bacopa monnieri (L.) Pennell

Abstract: Adventitious shoot buds were induced from leaf and stem explants of Bacopa monnieri on Murashige and Skoog medium supplemented with benzyladenine or kinetin. The source of the explants as well as different gelling agents in the medium were found to influence shoot induction and eventual shoot growth. The best response was obtained in leaf explants taken from shoot cultures grown in medium supplemented with 2 μM benzyladenine and gelled with 0.2% gelrite. A transverse section of the leaf explant incubated in this medium showed several shoot primordia emerging from the leaf surface. This system exhibited a potential for repeated harvesting of the shoots from the original leaf explant as the latter continued to expand and regenerate new shoots, upon repeated periodical subculturing onto fresh medium. However, the callusing response of the plant was very low. Qualitative TLC studies of the regenerated shoots revealed a phytochemical profile similar to that of the field grown-plants.

Regeneration of plants from seed-derived callus of Hybanthus enneaspermus L. Muell., a rare ethnobotanical herb

Abstract: A procedure was developed for plant regeneration of Hybanthus enneaspermus, a rare ethnobotanical herb from the Deccan peninsula in India, through seed-derived callus Seeds demonstrated a high induction frequency (694±28%) and a high yield (3644±25 mg) of light-yellow friable callus on Murashige and Skoog's (MS) medium containing 26 μm NAA and 22 μm BA within 4 weeks of incubation After 1 year of subculture, yellow friable and light-green compact calli types were established from initial light-yellow friable callus Shoot differentiation was achieved from light-green compact callus, but not from yellow friable callus Shoot differentiation resulted when light-green compact callus was transferred to MS medium supplemented with 88 μm BA and 26 μm NAA; the highest percentage of calli forming shoots (666±48%) and the highest number of shoots (89±03) were achieved in this medium Differentiated shoot buds elongated to 4–5 cm within 4 weeks The addition of casein hydrolysate (500 mg/l) and more potassium phosphate (186 mm) to the culture medium enhanced shoot differentiation Rooting was achieved on the shoots using half-strength MS medium containing 48 μm IBA About 70% of the plants were established in pots containing pure garden soil after 2 weeks of hardening The regenerated plants were morphologically uniform and exhibited normal seed set

Proline Accumulation, Protein Pattern and Photosynthesis in Bacopa Monniera Regenerants Grown under NaCl Stress

Abstract: Shoots of Bacopa monniera exhibited 100 % regeneration on Murashige and Skoog medium with 2 % sucrose, 0.2 mg dm−3 1-naphthaleneacetic acid, 0.5 mg dm−3 6-benzylaminopurine and 50 mg dm−3 glutamine. When the medium was supplied with various concentrations (5 - 15 g dm−3) of sodium chloride, proline content in regenerants was six times higher than in the control. With increasing NaCl concentration photosynthetic rate decreased and fresh mass and root length of regenerants declined. NaCl also induced formation of new proteins.

In vitro callus induction and regeneration studies in Withania somnifera

Abstract: Callus cultures were initiated from axillary leaves, axillary shoots, hypocotyls, and root segments on Murashige and Skoog (MS) (1962) medium supplemented with 2,4-D (2 mg l−1) and KN (0.2 mg l−1). Shoots differentiated best from axillary shoot base callus on MS medium containing BA (2 mg l−1). Regenerated shoots rooted best on MS medium containing IBA (2 mg l−1) alone, and IBA (2 mg l−1) with IAA (2 mg l−1). Plantlets were transferred to pots containing sand and soil mixture, acclimatized in a culture room and afterwards transferred to the glasshouse.

A combination of thidiazuron and benzyladenine promotes multiple shoot production from cotyledonary node explants of faba bean (Vicia faba L.)

Abstract: A procedure for multiple shoot formation from cotyledonary node explants of faba bean (Vicia faba LcvSGhdar) cultured on MS medium containing benzyladenine (BA) and thidiazuron (TDZ) was developed Explants on medium with TDZ in combination with BA produced a higher number of shoots than with either cytokinin alone The highest number of shoots was obtained when explants from 7-day-old seedlings were cultured on MS medium supplemented with TDZ and BA (2 mgl−1 each) for 31 days before transfer to hormone-free MS medium for elongation Shoots produced in vitro were rooted on half-strength agar-solidified MS basal medium or with 025 or 05 mgl−1 naphthalenacetic acid (NAA) prior to transfer to green house conditions This procedure was found to be applicable to seven other cultivars of faba bean from widely diverse provenances Thus, it can be advantageously applied to the production of transgenic faba bean plants

Phenylacetic acid improves bud elongation and in vitro plant regeneration efficiency in Capsicum annuum L.

Abstract: We have developed a highly efficient three-stage protocol for plant regeneration in sunflower (Helianthus annuus L.) from embryonal cotyledons. This protocol uses phenylacetic acid (PAA) for both shoot-bud induction and the elongation of smaller buds. The medium used for inducing bud formation from the cotyledons was modified MS medium supplemented with 3 mg/l 6-benzylaminopurine (BAP) and 0.5 mg/l PAA. Buds were elongated on MS medium supplemented either with only 0.2 mg/l gibberellic acid (GA3) or with 0.2 mg/l GA3 + 0.1 mg/l PAA + 0.3 mg/l BAP. The elongated shoots were then transferred onto rooting medium containing 1 mg/l PAA. The complete plantlets with well-developed roots were transferred to field conditions where they survived and set normal seeds. The induction of shoot buds from embryonal cotyledons was also observed on modified MS medium supplemented with 0.5–5 mg/l BAP in combination with 0.5–5 mg/l α-naphthaleneacetic acid (NAA). In this case, the formation of callus took place along with shoot-bud formation, which hindered further development of the latter. The presence of PAA with BAP in the primary bud induction medium promoted normal development and elongation of shoot buds.

Somatic embryogenesis from mature leaves of rose (Rosa sp.)

Abstract: Several plant growth regulators (0.3–53.3 μm 6-benzyladenine, 2,4-dichlorophenoxyacetic acid, gibberellic acid, 3-indoleacetic acid, p-chlorophenoxyacetic acid, kinetin and α-naphthylacetic acid), alone or in combination, and culture conditions were tested for their capacity to induce somatic embryogenesis from mature leaf and stem explants of rose (Rosa sp.) of four commercial rose cultivars (Baccara, Mercedes, Ronto and Soraya). Somatic embryos were only induced from mature leaf explants derived from Soraya on Murashige and Skoog (MS) medium supplemented with 53.5 μm p-chlorophenoxyacetic acid and 4.6 μm kinetin, although satisfactory callus induction rates were obtained from all cultivars. After subculturing on the same medium, embryos at various developmental stages (globular, heart and torpedo shaped) were transferred for maturation onto a MS medium supplemented with 5.2 μm 6-benzyladenine and 5.7 μm 3-indoleacetic acid. Germination of mature embryos took place after subculturing them onto medium of the same composition. Plantlets regenerated from embryos and bearing three to four leaves were transferred to a greenhouse.

Regeneration and large-scale propagation of bamboo (Dendrocalamus strictus Nees) through somatic embryogenesis

Abstract: A complete protocol for large-scale propagation of Dendrocalamus strictus Nees by somatic embryogenesis has been developed. Seeds cultured on agar-solidified Murashige and Skoog (MS) medium supplemented with 2,4-dichlorophenoxyacetic acid (2,4-D; 3×10–5 m) produced embryogenic callus from proliferation of the embryo. Somatic embryos formed in vitro multiplied rapidly (two- to five fold every 5 weeks) on semi-solid MS medium containing 2,4-D (1×10–5 m), kinetin (Kn) (5×10–6 m), 1-indolebutyric acid (IBA) (2×10–6 m) and soluble polyvinylpyrrolidone (PVP) (250 mg l–1), or MS with 2,4-D (1×10–5 m), 6-benzylaminopurine (BAP) (1×10–5 m), and soluble PVP (250 mg l–1). Upon transfer to MS containing 1-naphthaleneacetic acid (NAA) (5×10–6 m), Kn (5×10–6 m) and soluble PVP (250 mg l–1), the dark-green embryos developed into healthy plantlets. Unrooted shoots, if any, obtained on the multiplication media were rooted on MS major salts reduced to half strength supplemented with NAA (3×10–6 m) and IBA (2.5×10–6 m). The rooted plants were successfully transferred to soil in polythene bags with over 80% survival. Using this methodology, more than 100,000 plants have been produced.

Micropropagation of Dendrocalamus asper by shoot proliferation using seeds

Abstract: An efficient and reproducible procedure for the large-scale propagation of Dendrocalamus asper is described. High-frequency direct shoot proliferation was induced in aseptic seed cultures of D. asper on modified Murashige and Skoog's (1962) medium supplemented with 1.0–10.0 mg/l benzyladenine (BA). Multiple shoots (1–25) were formed within 5 weeks of seed culture without root formation. The shoot-forming capacity of seeds was influenced by the BA concentration in the medium. Proliferating shoot cultures were established by repeatedly subculturing shoots in propagules of 3 shoots each. A multiplication rate of 15–16 fold was achieved on MS medium +3.0 mg/l BA. Roots were formed on excised propagules of 3–5 shoots when transferred to MS medium containing 10.0 mg/l indole-3-butyric acid (IBA) or 3.0 mg/l 1-naphthaleneacetic acid (NAA). Plantlets were hardened, acclimatized and established in soil, where they exhibited normal growth.

Somatic embryogenesis and in vitro rosmarinic acid accumulation in Salvia officinalis and S. fruticosa leaf callus cultures

Abstract: The effect of explant age, plant growth regulators and culture conditions on somatic embryogenesis and rosmarinic acid production from leaf explants of Salvia officinalis and S. fruticosa plants collected in Greece was investigated. Embryogenic callus with numerous spherical somatic embryos could be induced on explants derived from both species and cultured for 3 weeks on a Murashige and Skoog (MS) medium supplemented with 1.8–18 μm 2,4-dichlorophenoxyacetic acid (2,4-D) and kinetin (Kin) or 10.5–21 μm 1-naphthalenacetic acid and 6-benzyladenine. Only explants from young plants (with six to eight leaves) responded to the culture treatments and, in general, low light intensities (50 μmol m–2 s–1) favoured callus formation and induction of somatic embryos. Somatic embryos were further developed on the same medium. Heart- and torpedo-shaped embryos (1–2 mm long) were subcultured on a growth-regulator-free MS medium for maturation. Maximum rosmarinic acid accumulation in S. officinalis and S. fruticosa callus cultured on 4.5 μm 2,4-D and 4.5 μm Kin was 25.9 and 29.0 g/l, respectively.

In vitro regeneration of a mature leguminous liana (Bauhinia vahlii Wight & Arnott)

Abstract: An in vitro propagation protocol has been developed from mature lianas of Bauhinia valii. Browning was the major obstacle in the establishment of cultures. Explants collected during the growing season (April–June) showed maximum browning; however, browning was minimal during the dormant phase. This problem was circumvented by soaking the sterilized explants in a solution of antioxidant (50 mg l–1 ascorbic acid+75 mg l–1 citric acid). The explants were thereafter transferred to culture room conditions after an initial incubation in the dark at 4 °C for 48 h. Shoot proliferation (58%), shoot number (4.5) and shoot length (35 mm) was best in Murashige and Skoog (MS) medium supplemented with 2.5 μM kinetin + 100 mg l–1 adenine sulfate. Seasonal fluctuations significantly affected the proliferation potential of the explants. March– April was found to be the best season for shoot initiation. Microshoots were rooted on a half-strength, growth regulator-free, agar-gelled Murashige and Skoog medium after a dip in half-strength MS liquid medium containing 1-naph-thaleneacetic acid + indole-3-butyric acid (10 μM). Rooted plantlets were potted and acclimatized under culture room conditions for 4 weeks before transfer to a polyhouse.

Micropropagation of the threatened Nepalese medicinal plant Swertia chirata Buch.-Ham. ex Wall.

Abstract: Multiplication by adventitious shoot regeneration from root explants was found to be the most suitable method for the propagation of Swertia chirata. A two-step system consisting of an initial 3-week cultivation on modified Murashige and Skoog medium supplemented with 3 μM 6-benzylaminopurine (BAP), followed by another period of 3 weeks on hormone-free medium was used. After rooting and acclimatization micropropagated plants could be successfully cultivated in Nepal.

In vitro propagation of Gladiolus hybridus Hort.: Synergistic effect of heat shock and sucrose on morphogenesis

Abstract: Three cultivars (cvs.) of Gladiolus hybridus Hort., namely ‘Her Majesty’, ‘Aldebaran’ and ‘Bright Eye’ were successfully micropropagated. The cultures were established using intact cormels or segments of cormels and inflorescence axes on Murashige and Skoog (1962; MS) medium. The response depended on media supplements; both callus formation or direct induction of shoot buds was observed. Shoot differentiation from callus could be obtained on MS medium containing 1.0 μM BA (6–benzyladenine) and 10.0 μM NAA (α-naphthalene acetic acid) in all three cultivars. The same could be achieved by giving a heat shock (HS; 50 °C, 1h) to callus cultures (in case of ‘Her Majesty’ and ‘Aldebaran’ only) maintained on the basal medium. In these two cultivars, high sucrose concentration (0.232, 0.290 or 0.348 M) also favoured growth and proliferation of shoot cultures on a plant growth regulator-free medium at 20 °C in comparison to the cultures kept at 25 °C. On the other hand, shoot cultures maintained on the basal medium at 25 °C containing normal (0.058 M, i.e., 2.0%, w/v) sucrose concentration responded similar to those maintained at 20 °C on a high sucrose medium; reduced response was observed on normal sucrose containing medium at 20 °C. Heat shock enhanced shoot proliferation in the cultures maintained on basal medium, but induced prolific rooting in shoot cultures, within 5 days of HS, on high sucrose (optimum 0.232 M) medium. While the number of roots increased at higher sucrose concentrations in the medium in case of cvs. ‘Her Majesty’ and ‘Aldebaran’, the same was found to be independent of sucrose concentration in cv. ‘Bright Eye’. Generally the rooted plants produced on high sucrose (0.232 M) medium in comparison to medium with normal sucrose concentration showed better survival (ca. 90% as against 40%) in the soil.

Micropropagation from inflorescence stems of the Spanish endemic plant Centaurea paui Loscos ex Willk. (Compositae)

Abstract: Tissue culture techniques have been established as a useful approach for ex situ conservation of rare, endemic or threatened plant species. This report describes the micropropagation of Centaurea paui Loscos ex Willk (Compositae), an extremely endangered plant species endemic to the Valencia Community (eastern Spain), as a conservation measure which does not cause damage to the wild plants used as explant source. Inflorescence nodal segments of C. paui were selected as explants for in vitro establishment. The best rate of shoot proliferation was obtained on Murashige and Skoog (MS) mineral medium supplemented with 0.5 mg/l 6-benzyladenine or with 2 mg/l kinetin. Maximum shoot elongation was achieved without growth regulators, and the addition of cytokinins significantly decreased their size. In vitro rooting of shoots was difficult after 6 weeks on rooting media. The combination of 2 mg/l indole-3-acetic acid plus 2 mg/l indole-3-butyric acid on MS medium yielded the best results. In this medium, 40% of shoots rooted before 30 days of culture. About 70% of the rooted plants were successfully transferred to pots and acclimatized to ex vitro conditions.

Effects of sucrose concentration, supporting material and number of air exchanges of the vessel on the growth of in vitro coffee plantlets

Abstract: Growth of coffee (Coffea arabusta) plantlets cultured in vitroas affected by sugar, types of supporting material and number of air exchanges of the vessel was investigated. Single node cuttings of in vitro coffee plantlets were cultured on half strength MS medium with or without 20 g l−1 sucrose. Two types of supporting material, agar and Florialite, and two levels of air exchange expressed by number of air exchanges per vessel, 0.2 and 2.3 h−1, were studied. At the end of a 40-day culture period, fresh weight, shoot length, root length and leaf area of plantlets when cultured on Florialite soaked in sugar-free medium and under the higher number of air exchanges were greater than those in sugar containing medium. Callus was observed at the shoot base of plantlets grown on agar medium containing sucrose. Photosynthetic ability of coffee plantlets in vitro was also significantly increased when grown on sugar-free medium with the high number of air exchanges and Florialite as a supporting material.

In vitro production of potato microtubers in liquid medium using temporary immersion

Abstract: CIRAD developed a new apparatus for plant tissue culture, using temporary immersion in a liquid medium. This apparatus was adapted to the microtuber production in potato. The procedure is as follows: single node cultivation on MS medium containing 30 g/l sucrose in the light for 2 weeks, induction of microtuberisation with 80 g/l sucrose over a 2 week period in the light, followed by a further 6 weeks in the dark. All experiments were performed at 20 °C. The basic vessel had a capacity of approximately 11;30 nodes were cultivated per vessel. Depending on the cultivars tested (Bintje, Ostara and Desiree) 47 to 115 microtubers were harvested per vessel. Between 30 and 60% of the microtubers weighted over 0.5 g and between 10 and 40% over 0.8 g. Sprouting is still under investigation. Preliminary results indicate that the dormancy period was relatively short and several stems were obtained per microtuber. These results seem to be better than those usually reported. Only one simple protocol has been tested and further improvements are probably easy to obtain.

Somatic embryogenesis and plant regeneration from pistil thin cell layers of Citrus

Abstract: Callus induction, somatic embryogenesis and plant regeneration were obtained in six different citrus species [Citrus deliciosa Ten. (cv 'Avana'), C.limon (L.) Burm. (cv 'Berna'), C.madurensis Lour. (cv 'CNR P9'), C.medica L. (cv 'Cedro di Trabia'), C.tardiva Hort. ex Tan. (cv 'CNR P6'), C.sinensis (L.) Osb. (cv 'Ugdulena 7')] from cultures of pistil transverse thin cell layer explants [(t)TCL]. Explants were cultured on three different media: the nutrients and vitamins of Murashige and Skoog medium alone (MS) or MS supplemented with either 500 mg l–1 malt extract (MS I) or 500 mg l–1 malt extract and 13.3 μM 6-benzylaminopurine (MS II). Sucrose (146 mM) was used as the carbon source. Somatic embryos were visible 2–5 months after culture initiation. The different genotypes showed a different embryogenic frequency from stigma, style and ovary (t)TCL explants. All of the cultivars regenerated somatic embryos. Percentages of style (t)TCL explants producing somatic embryos ranged from 0% (C.deliciosa, C.madurensis, C.sinensis and C.tardiva on the three different media) to 5.2% (C.limon on MS II). Embryo formation in stigma (t)TCL explants ranged from 0% (C.madurensis on MS and MS I, C.sinensis on MS, C.deliciosa and C.tardiva on the three different media) to 42.4% (C.limon on MS II). Embryo formation in ovary (t)TCL explants ranged from 0% (C.deliciosa on MS, C.limon, C.medica, and C.sinensis on the three different media) to 9.3% (C.tardiva on MS I). After about 12 weeks somatic embryos developed into plantlets at a high frequency.

Rapid clonal propagation of Plumbago zeylanica Linn.

Abstract: An efficient protocol was developed for in vitro clonal propagation of Plumbago zeylanica Linn. through nodal culture. Multiple shoots were induced from nodal explants of P. zeylanica on Murashige and Skoog's (1962) medium supplemented with 0.5 mg L−1 to 1.0 mg.L−1 6-benzyladenine and 3% (w/v) sucrose. Inclusion of IAA (0.01 mg L−1) in the culture medium improved the frequency of production of multiple shoots. Rooting was readily achieved upon transferring the shoots onto half-strength MS medium supplemented with 0.25 mg L−1 IBA and 2% (w/v) sucrose. Micropropagated plantlets were hardened in the greenhouse and successfully established in soil.

Agrobacterium tumefaciens-mediated transformation of greenhouse-grown Brassica rapa ssp. oleifera

Abstract: Greenhouse-grown plants of turnip rape Brassica rapa ssp. oleifera (syn. B. campestris) cv. Valtti and Sisu were transformed by Agrobacterium tumefaciens infection. Of the three A. tumefaciens strains tested (C58C1, EHA105 and LBA4404), LBA4404 gave the best results. Segments excised from one to two upper internodes of an inflorescence-carrying stem served as explants for the Agrobacterium infection. Cultivation of the explants horizontally during the first 3 days of co-cultivation with A. tumefaciens following immediate selection of transformed tissue of the stem segments placed vertically basal side down were critical. Use of silver nitrate (5–10 mg/l) in the culture medium and Micropore (3 M) paper tape for sealing plates was also beneficial. Transgenic shoots were recovered using either hygromycin or kanamycin (20–25 mg/l) selection. Hygromycin was preferable, as the proportion of `escapes' was 90% under kanamycin and 10% under hygromycin selection. Regeneration was achieved by culturing the explants for 3–6 days on 0.5 mg/l of 2,4-di-chlorophenoxyacetic acid and 1–2 weeks on 2–3 mg/l of 6-benzyl aminopurine with/without 0.05 mg/l α-naphthaleneacetic acid. Recovered shoots were then cultured on hormone-free MS medium. This culture program gave 60–80% shoot regeneration. Regenerants were tested by histological β-glucuronidase staining and Southern blotting. The recovery rate of transgenic shoots was 4–9% of the number of explants used in the experiments.

Micropropagation of Lippia junelliana (Mold.) Tronc.

Abstract: A method for the micropropagation of Lippia junelliana (Mold.) Tronc. from shoot tips or nodal segments was developed. Proliferating microshoot cultures were obtained by placing shoot tips or nodal segments on full strength Murashige and Skoog medium (MS) supplemented with 4.4 μM benzyladenine (BA) or 0.04 μM indolebutyric acid- (IBA) plus 4.4 μM BA. The rooting of shoots was better on full-strength MS medium without growth regulators. Rooted plantlets were successfully acclimatized to soil. The shoot cultures showed a lower essential oil accumulation in comparison with parent plants. Essential oil accumulation is closely related with growth and shows a negative correlation with shoot proliferation.

Induction and growth characteristics of embryogenic avocado cultures

Abstract: Embryogenic cultures were induced from immature avocado zygotic embryos representing different botanical races and complex hybrids. The optimum induction medium consisted of B5 major salts, MS minor salts, 0.4 mg l−1 thiamine HCl, 100 mg l−1 myo-inositol, 30 g l−1 sucrose, 0.41 μM picloram and 8 g l−1 TC agar. Somatic embryogenesis occurred directly from the explants on induction medium, and secondary embryos and proembryonic masses proliferated in liquid and on semisolid maintenance medium. Embryogenic culture maintainance was optimized in liquid, filter-sterilized MS medium, supplemented with 30–50 mg l−1 sucrose, 4 mg l−1 thiamine HCl and 0.41 μM picloram. Two types of embryogenic cultures were recognized: –genotypes that proliferated as proembryonic masses in the presence of auxin (PEM-type) and; –genotypes in which the heart stage and later stages of somatic embryos developed in the presence of auxin(SE-type). Embryogenic suspension cultures became increasingly disorganized over time, and this was associated with progressive loss of embryogenic potential.

Efficient in vitro propagation of Agave parrasana Berger

Abstract: An efficient method for the in vitro propagation of Agave parrasana Berger, an important ornamental plant species native to the state of Coahuila, Mexico, was developed. Proliferation of good quality shoots was achieved on agar-solidified basal MS medium supplemented with L2 vitamins and 13.3 μM benzyladenine. Rooting was successful in the basal medium with no growth regulators; however, a light intensity of 100 μmol m-2 s-1 was found to promote better rooting than 25 μmol m-2 s-1.

High-efficiency plant production via direct somatic single embryogenesis from preplasmolysed cotyledons of Panax ginseng and possible dormancy of somatic embryos

Abstract: Cotyledon explants of immature ginseng zygotic embryos cultured on Murashige and Skoog medium lacking growth regulators formed somatic embryos directly, most in a multiple state, fused together and to the parent cotyledon explants. When the cotyledon explants of ginseng were pretreated with 1.0 m sucrose for 24–72 h, all the somatic embryos developed individually from all surfaces of the cotyledons and the number of somatic embryos per explant was enhanced fourfold. Histological observation revealed that all the single somatic embryos from preplasmolysed cotyledons originated from epidermal single cells, whereas all the multiple embryos from cotyledons without pretreatment originated from epidermal and subepidermal cell masses. When the somatic embryos matured to the cotyledonary stage, further growth ceased and they remained white, probably indicating dormancy. Gibberellic acid (GA3) (over 1.0 mg/l) or chilling treatment (–2°C for over 8 weeks) were prerequisites for the germination of somatic embryos. Ultrastructural observation revealed that the cotyledon cells of somatic embryos without chilling or GA3 treatment contained numerous lipid reserves, dense cytoplasm, proplastids and non-activated mitochondria, whereas the cotyledon cells of somatic embryos after chilling or GA3 treatment were highly vacuolated and contained well-developed chloroplasts and active-state mitochondria enclosing numerous cristae, indicating that in-vitro-developed somatic embryos of P. ginseng may be dormant after maturing in a manner similar to zygotic embryos.

Rosavin as a product of glycosylation by Rhodiola rosea (roseroot) cell cultures

Abstract: The paper discusses glycosylation of trans-cinnamyl alcohol to obtain the biologically active compound rosavin and possibly other cinnamoylglycosides. Cell suspension cultures of Rhodiola rosea were established from callus of leaf origin cultured under light in a modified Murashige and Skoog medium. Under these conditions, no rosavin was formed. However, when trans-cinnamyl alcohol (2.5 mM; in MeOH) was added to the medium, after 72 h incubation cells transformed over 90% of the cinnamyl alcohol into a number of unidentified products. The structure of potential rosavin accumulated in intracellular spaces was elucidated as [3-phenyl-2-propenyl-O-(6'-O-α-L-arabinopyranosyl) -β-D-glucopyranoside] by means of chemical and spectral analysis using TLC, HPLC, UV, LSIMS and NMR methods. Rosavin yields of 0.03–1.01% dry weight were obtained. The actual amount depended on the cell strain cultured and the biotransformation period.