Stress and defense responses in plant secondary metabolites production.

In the growth condition(s) of plants, numerous secondary metabolites (SMs) are produced by them to serve variety of cellular functions essential for physiological processes, and recent increasing evidences have implicated stress and defense response signaling in their production. The type and concentration(s) of secondary molecule(s) produced by a plant are determined by the species, genotype, physiology, developmental stage and environmental factors during growth. This suggests the physiological adaptive responses employed by various plant taxonomic groups in coping with the stress and defensive stimuli. The past recent decades had witnessed renewed interest to study abiotic factors that influence secondary metabolism during in vitro and in vivo growth of plants. Application of molecular biology tools and techniques are facilitating understanding the signaling processes and pathways involved in the SMs production at subcellular, cellular, organ and whole plant systems during in vivo and in vitro growth, with application in metabolic engineering of biosynthetic pathways intermediates.

Proteome study of somatic embryogenesis in Nothapodytes nimmoniana (J. Graham) Mabberly.

Somatic embryogenesis (SE) is the most suitable biotechnological tool for the rapid clonal propagation of endangered woody plants, but many bottlenecks limit understanding its molecular and physiological processes in Nothapodytes nimmoniana. Combinations of two-dimensional electrophoresis (2-DE) and mass spectrometry (MaSp) were used to study proteomic expression changes during SE of the forest tree. Callus was induced from mature seed embryos, and embryogenic callus (EC) obtained at very low frequency after about 6 month culture. Globular embryos were induced from the seed embryo-derived EC and the subsequent stages of the SE. Analysis of the extracted proteins from globular, heart/torpedo-shaped, and maturing embryo stages resolved in the 2-DE gels showed increased protein expression across developmental stages of the somatic embryos. The mass spectrometric analysis with database search aided identification of 55 out of 100 and 54 selected protein spots. Identified proteins classified by the cellular role which they perform are involved in aspects of stress responses, energy metabolism, carbon fixation, secondary metabolism, and other metabolic functions, while three proteins are of unknown cellular role. The putative role of the expressed proteins during SE provided insight into the physiology of somatic embryo development in N. nimmoniana.

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

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

Changes in the biochemical parameters of albino, hyperhydric and normal green leaves of Caladium bicolor cv. "Bleeding hearts" in vitro long-term cultures.

Caladium bicolor is an excellent landscape and pot plant grown for its ornamental value due to the varied shapes and multi-colored foliage. However, the ornamental value of in vitro clonally propagated plants is affected by the occurrence of morpho-physiological anomalies in the long-term cultures. During the long-term culture of rhizome-derived shoot cultures of C. bicolor cv. "Bleeding hearts", some incidences of hyperhydricity and albinism were observed. Hyperhydricity occurred at high frequency in the shoot cultures and was more prevalent in the liquid over solid medium cultivated plantlets. The addition of silver nitrate at 7.5 µM reduced prevalence of the condition over other treatment concentrations but, changes in the gelling agent concentration and photoperiodic incubation were ineffective. Albinism also occurred at a higher frequency but, was more prevalent in the cultures incubated below or above 12 h light/dark photoperiods. Evaluation of the physiological, biochemical parameters showed differences between the leaves of albino, hyperhydric and normal green C. bicolor micro shoots. The experimental results obtained suggested an effective role of pigment molecules, cellular osmolytes, and enzymatic and non-enzymatic antioxidant systems in the development of albino and hyperhydric leaves in C. bicolor micro shoot cultures. Photoperiodic incubation and culture conditions showed some effect on the incidences of hyperhydricity and albinism in the shoot cultures.

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

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

Stress and defense responses in plant secondary metabolites production

In the growth condition(s) of plants, numerous secondary metabolites (SMs) are produced by them to serve variety of cellular functions essential for physiological processes, and recent increasing evidences have implicated stress and defense response signaling in their production. The type and concentration(s) of secondary molecule(s) produced by a plant are determined by the species, genotype, physiology, developmental stage and environmental factors during growth. This suggests the physiological adaptive responses employed by various plant taxonomic groups in coping with the stress and defensive stimuli. The past recent decades had witnessed renewed interest to study abiotic factors that influence secondary metabolism during in vitro and in vivo growth of plants. Application of molecular biology tools and techniques are facilitating understanding the signaling processes and pathways involved in the SMs production at subcellular, cellular, organ and whole plant systems during in vivo and in vitro growth, with application in metabolic engineering of biosynthetic pathways intermediates.

Anticancer Alkaloids from Trees: Development into Drugs.

Trees have made an enormous phytochemical contribution in anticancer drugs' development more than any other life form. The contributions include alkaloids that are biosynthesized in various ways and yield. Lead alkaloids isolated from the trees are taxol and camptothecins that currently have annual sales in billion dollars. Other important alkaloids isolated from these life forms include rohitukine, harringtonine, acronycine, thalicarpine, usambarensine, ellipticine, and matrines. Studies on their mechanism of action and target on the DNA and protein of cancerous cells aided the development of potent hemisynthesized congeners. The molecules and their congeners passed/are passing a long period of historical development before approved as antineoplastic drugs for cancer chemotherapy. Some of them did not find the application as anticancer drugs due to ineffectiveness in clinical trials; others are generating research interest in the antineoplastic activity at the present and have reached clinical trial stages. Potentials in antineoplastic molecules from trees are high and are hoped to be commensurate with cancer types afflicting human society in the future.

Rethinking Ginkgo biloba L.: Medicinal uses and conservation.

Ginkgo biloba (G. biloba) is an ancient medicinal tree species that has been in existence for millennia without undergoing modifications due to its resistance to environmental stresses. Palaeobotanical history showed a wide distribution of the species across the globe but declined over geological time, becoming restricted to narrow geographical range with few surviving individuals in the modern day. The tree is slow growing, adapted to many ecological conditions and shows numerous adaptation in developmental patterns. Medicinal use of the species is attracting research interest, especially the various parts of the tree that are used in orthodox or traditional medicine to treat diseases due to the many bioactive compounds. The primary compounds receiving increasing research interest are the triterpene lactones and flavonoids; these are the target of biotechnological strategies being employed to enhance production. Many genetic and environmental factors have contributed to the endangered status of the species; conservation measures are required to protect it from extinction. In many countries, the cultivation of plantations for the supply of ginkgo leaf-based pharmaceutical formulations is in progress, and efforts to standardize ginkgo leaf extract as herbal medication for human use are being made. Microcuttings and cuttings, cryopreservation, and plant tissue culture have all aided to conserve G. biloba.

Somatic embryo mediated mass production of Catharanthus roseus in culture vessel (bioreactor) - A comparative study.

The purpose of this study was to evaluate and compare the use of liquid and solid Murashige and Skoog (MS) medium in different culture vessels for mass production of Catharanthus roseus, an important source of anticancerous compounds, vincristine and vinblastine. Three media conditions i.e. agar-solidified medium (S), liquid medium in agitated conical flask (L) and growtek bioreactor (B) were used. Rapid propagation was achieved through in vitro somatic embryogenesis pathway. The process of embryogenesis has been categorized into induction, proliferation, maturation and germination stages. All in vitro embryogenesis stages were conducted by withdrawing spent liquid medium and by adding fresh MS medium. In optimized 4.52 µM 2,4-D added MS, the callus biomass growth was low in solid (1.65 g) compared to liquid medium in agitated conical flask (1.95 g) and in bioreactor (2.11 g). The number of normal somatic embryos was more in solid medium (99.75/50 mg of callus mass) compared to liquid medium used in conical flask (83.25/callus mass) and growtek bioreactor (84.88/callus mass). The in vitro raised embryos maturated in GA3 (2.60 µM) added medium; and in bioreactor the embryo growth was high, a maximum length of 9.82 mm was observed at the end of four weeks. These embryos germinated into seedlings in BAP (2.22 µM) added medium and the embryo germination ability was more (59.41%) in bioreactor compared to liquid medium in conical flask (55.5%). Shoot length (11.25 mm) was also high in bioreactor compared to agitated conical flask. The liquid medium used in agitated conical flask and bioreactor increased seedling production efficiency, at the same time it also reduced plant recovery time. The embryo generated plants grew normally in outdoor conditions. The exploitation of medium to large culture vessel or bioreactor may make the process more efficient in getting large number of Catharanthus plant as it is the only source of anti-cancerous alkaloids, vincristine and vinblastine.