Sankaran KrishnaRaj

Tryptophan is a precursor for melatonin and serotonin biosynthesis in in vitro regenerated St. John's wort (Hypericum perforatum L. cv. Anthos) plants

Hypericum perforatum cv. Anthos) is presented. Isotope tracer experiments were performed on plantlets regenerated from thidiazuron-induced stem explants and grown on MS basal medium for 2 months. Radiolabel from 14C-tryptophan was recovered as 14C-indoleacetic acid, 14C-tryptamine, 14C-5-hydroxytryptophan, 14C-serotonin and 14C-melatonin in the treated St. Johns wort plantlets. Chromatographic peak identity was confirmed by high performance liquid chromatography-mass spectrometry-mass spectrometry and quantification of melatonin by radioimmunoassay. Significantly more radiolabel was recovered in serotonin relative to melatonin under low light conditions with this ratio being reversed under increased lighting, indicating that the rate of flow through this biosynthetic pathway is regulated, at least in part, by light.

Phytoremediation of Heavy Metal Contaminated and Polluted Soils

Most plants and animals depend on soil, as a growth substrate, for their sustained growth and development. In many instances the sustenance of life in the soil matrix is adversely affected by the presence of deleterious substances or contaminants. These pollutants can be broadly classified into two groups: (1) organic, which contain carbon, and (2) inorganic, devoid of carbon (Webber and Singh 1995). The focus of this chapter is to provide an overview of the plant-based remediation strategies for inorganic pollutants, while the use of such strategies for organic pollutants is also briefly discussed (for an indepth review see Cunningham et al. 1995).

Somatic embryogenesis and organogenesis in peanut: The role of thidiazuron and N6-benzylaminopurine in the induction of plant morphogenesis

Intact peanut (Arachis hypogaea L.) seeds, incubated on media containing N6-benzylaminopurine (BAP) or thidiazuron (TDZ) exhibited de novo regeneration at the hypocotyledonary notch region. Regeneration was observed when seeds were cultured on either TDZ or BAP but the optimal level of media supplementation was 10 μmol·L−1 for TDZ and 50 μmol−L−1 for BAP. Light microscopic observations revealed that the regenerants induced by TDZ were somatic embryos while those induced by BAP were shoots. An alternative approach of exposing the seeds to TDZ was through vacuum infiltration followed by culture on basal media but BAP did not induce regeneration by this method. Although TDZ has often been classified as a synthetic cytokinin, our results clearly demonstrate that seedlings treated with TDZ undergo a different morphological route of development than that induced by purine cytokinins.

Role of endogenous purine metabolism in thidiazuron-induced somatic embryogenesis of peanut (Arachis hypogaea L.)

Somatic embryogenesis was induced at the hypocotyledonary notch region of intact peanut (Arachis hypogaea L.) seedings cultured on a medium containing 10 μmol·L−1 thidiazuron (TDZ). Inclusion of the purine analogs 2,6-diaminopurine (DAP), azaadenine or azaguanine to the thidiazuron amended medium inhibited the embryogenic response of the seedlings. DAP-mediated inhibition was not overcome by the addition of adenine sulphate. Inhibition of the embryogenic response by DAP provides evidence that the TDZ-induced accumulation of purine cytokinins is an essential component of TDZ-induced somatic embryogenesis process. Analyses of the endogenous level of purine metabolites indicated that supplementation of the media with TDZ resulted in an overall increase in the endogenous cytokinins while DAP inhibited the purine recycling resulting in decreased levels of endogenous adenine and zeatin.

Inhibitory effect of GA3 on the development of thidiazuron-induced somatic embryogenesis in geranium (Pelargonium xhortorum Bailey) hypocotyl cultures

Somatic embryogenesis in geranium (Pelargonium xhortorum Bailey cv ‘Scarlet Orbit Improved’) can be achieved by incubating hypocotyl explants on MS medium supplemented with thidiazuron (TDZ; 10 μM for 3 days followed by subculture on medium devoid of any plant growth regulators. The presence of gibberellins (GAs) during both the induction and expression phases of embryogenesis was significantly detrimental to somatic embryo formation on the hypocotyl explants. The addition of the GA-synthesis inhibitors paclobutrazol, uniconazole or ancymidol during the period of growth and differentiation of somatic embryos increased the number of somatic embryos formed on each explant. However, paclobutrazol added during the period of induction had no significant influence on somatic embryo formation. Results suggest that both exogenously supplied as well as endogenous GAs play a role, albeit a negative one, on somatic embryogenesis of geranium.

Morphological and Physiological Changes during Thidiazuron-Induced Somatic Embryogenesis in Geranium (Pelargonium x hortorum Bailey) Hypocotyl Cultures

Somatic embryogenesis in geranium (Pelargonium x hortorum Bailey cv. Scarlet Orbit Improved) was achieved by culturing hypocotyl explants on media supplemented with either thidiazuron (TDZ) or indol-3-yl-acetic acid (IAA) and 6-benzylaminopurine (BAP). The physiological relationships between the morphological changes that lead to complete development of somatic embryos and the endogenous levels of plant growth regulators were studied. TDZ induced similar but more pronounced and earlier morphological changes in cultured tissues than did IAA and BAP, but after 21 d in culture, both treatments promoted the formation of somatic embryos of different developmental stages that were similar to zygotic embryos. TDZ induced a higher number of somatic embryos, with similar but greater increases in the endogenous levels of auxins and cytokinins, than did IAA and BAP. This may indicate that TDZ may be a more potent plant growth regulator in inducing physiological and morphological changes than combined auxin and cytokinin, during the process of somatic embryogenesis in geranium.

Recent advances in Pelargonium in vitro regeneration systems

Recent advances in the development of protocols for in vitro culture and genetic manipulation have provided new avenues for the development of novel varieties of Pelargonium and for use as model systems for investigating the factors controlling plant morphogenesis. Optimized techniques of meristem culture have supplemented the culture indexing methods in commercial greenhouse production resulting in availability of large-scale pathogen indexed planting material. Currently, technologies are available for the mass in vitro propagation of F1 hybrid Pelargonium through both organogenesis and somatic embryogenesis. The somatic embryogenesis model system has allowed researchers to identify critical factors controlling plant morphogenesis in vitro such as regulation of regeneration by growth regulators, choice of explant and characterization of induction and expression phases of morphogenesis in Pelargonium. Also, optimization of technologies for genetic transformation of Pelargonium opened up the possibilities for developing genotypes with novel characters, including resistance to some of the major diseases. Finally, the development of regeneration systems for Pelargonium spp. has facilitated conventional crop improvement programs, thereby providing a valuable resource to the horticultural industry.

Metal Tolerance of Scented Geranium (Pelargonium sp. ‘Frensham’): Effects of Cadmium and Nickel on Chlorophyll Fluorescence Kinetics

ABSTRACT The ability of scented geraniums (Pelargonium sp. Frensham) to tolerate metal stress was assessed using chlorophyll a fluorescence kinetics. The effects of various concentrations of cadmium and nickel in the culture solution on photosynthetic efficiency in scented geranium was evaluated in comparison to two well-established metal accumulators, the Indian mustard (Brassica juncea), and the sunflower (Helianthus annuus), under greenhouse conditions. The efficiency of the photosynthetic apparatus was affected to varying degrees at all metal concentrations for the plants tested. High concentrations of cadmium (1000 mg L-1) did not significantly affect the efficiency of photosystem II activity, expressed as the ratio of variable fluorescence to maximal fluorescence (Fv/Fm), which remained high (0.738) in scented geraniums, but decreased significantly (P < 0.05) in Indian mustard (0.089) and sunflower (0.026) plants following 4 days of metal exposure. Similar trends were observed for nickel treatments. Also, the number and size of active photosynthetic reaction centers, as measured by the Fv/Fo ratio, was not significantly affected by metal exposure in scented geranium plants, while the ratio significantly decreased in Indian mustard and sunflower seedlings. The results suggest that scented geranium plants were able to overcome metal stress through (1) maintaining an efficient photosystem II activity, which is required for plant metabolism and physiological functions, as well as to overcome metal ion mediated stress, and (2) restricting damage to the photosynthetic apparatus (reaction centers) by metal ions.

The role of proline in thidiazuron-induced somatic embryogenesis of peanut

SummaryPeanut seeds germinated on media supplemented with thidiazuron [TDZ: N-phenyl-N′-(1,2,3-thiadiazol-yl)urea], formed somatic embryos at the hypocotyledonary notch region by Day 35 of the culture period. Supplementation of the culture media with proline, thioproline, or glutamine reduced the total number of embryos formed, but the resulting embryos were larger, greener and had a more synchronous development than the regenerants formed on media containing TDZ alone. Analysis of the endogenous amino acid content of the germinating seeds during the induction phase of somatic embryogenesis revealed accumulation of proline to 6% of the dry seed weight. Concurrent with the emergence of the radicle, the proline concentration remained significantly elevated throughout the expression phase of embryogenesis. Several other amino acids including alanine, aspartate, asparagine, glutamate, glutamine, γ-aminobutyrate (GABA), hydroxyproline, isoleucine, threonine and valine accumulated to peak values approximately 10-fold higher than those of the controls. These results indicate that proline plays a key role in directing the route of TDZ-induced somatic embryogenesis and that TDZ effectively stimulates a cascade of metabolic events resulting in the production of specific metabolites, including amino acids, required for the regenerative process.

A Fragrant Solution to Soil Remediation

ABSTRACT The ability of scented geraniums (Pelargonium sp. ‘Frensham’) plants to tolerate, uptake, and accumulate lead was assessed compared with two well-established metal accumulators, Indian mustard (Brassica juncea) and sunflower (Helianthus annuus), under greenhouse conditions. The efficiency of the photosynthetic apparatus and the number and size of active photosynthetic reaction centers (expressed as chlorophyll a fluorescence ratios of variable fluorescence to maximal fluorescence [Fv/Fm] and variable fluorescence to unquenchable portion of fluorescence [Fv/Fo], respectively) were affected to varying degrees at all metal concentrations in all the plants tested. Lead exposure did not significantly affect the efficiency of photosystem II activity or the number and size of the photosynthetic reaction centers in scented geraniums, but the ratios decreased significantly in Indian mustard and sunflower plants following lead exposure. In addition to tolerating high levels of lead, the scented geraniums accumulated in excess of 3000 mg of lead per kg DW of shoot and above 60,000 mg of lead per kg DW of root tissue. Additionally, scented geraniums exposed to a mixture of metals (lead + cadmium + nickel) had the ability to uptake in excess of 4.72% Pb + 0.44% Cd + 0.52% Ni per kg of root DW, as well as 0.17% Pb + 0.07% Cd + 0.14% Ni per kg of shoot DW within 14 d, indicated the potential for existence of more than one functional tolerance and metal uptake mechanism(s). The cellular localization of lead was assessed using transmission electron microscopy coupled with an X-ray microanalyzer. Lead accumulation was observed in the apoplasm and in the cytoplasm, vacuoles, and as distinct globules (potentially as lead-lignin or lead-phosphate complexes) on the cell membrane and cell wall. We conclude that the ability of scented geraniums to tolerate high lead accumulation in its biomass is due, in part, to limiting damage to photosynthetic apparatus and metal detoxification by formation of metal complexes.