Neera Bhalla Sarin

Ameliorative effects of proline on salt stress-induced lipid peroxidation in cell lines of groundnut ( Arachis hypogaea L.)

Abstract. Cell lines of a salt-sensitive cultivar of groundnut (Arachis hypogaea L. cv. JL24) were selected on media amended with high NaCl concentrations. Comparative analyses of the water status and ionic relations of salt-sensitive and salt-tolerant cell lines showed a decrease in the water potential (ψw) and solute potential (ψπ) with increases in salt stress. However, the maintenance of cellular turgor indicated active osmotic adjustments in response to salinity stress. In addition to the extrusion of Na+ in the NaCl-selected cell lines, a significant accumulation of proline was observed, which was probably associated with osmotic adjustments and the protection of membrane integrity. The addition of proline to the culture medium alleviated the salt stress-induced decline in fresh weight accumulation and reduced peroxidative damage to the lipid membranes, both in a concentration-dependent manner.

Differential response of arsenic stress in two varieties of Brassica juncea L.

Present study showed the toxicity caused by Arsenite (As(III)) and its detoxification responses in two varieties (Varuna and Pusa Bold) of Brassica juncea. Comparisons were made in leaves and roots of both the varieties, which showed that the accumulation pattern in both the varieties were dose and duration dependent, being more in roots for two days and in leaves for four days. Increase/decrease of antioxidant enzymes activities (SOD, CAT, GPX) showed not much changes at the given concentrations except that the enzyme activities showed significant increase at the lower concentrations. Semi quantitative RT-PCR analysis of PCS showed more expression of its transcript in P. Bold as compared to Varuna variety due to As(III) stress. The analysis of isoenzyme pattern in leaves of P. Bold showed five and two major bands of SOD and GPX, respectively. As(III) treatment leads to the activation of MAPK activity indicating role of this important cascade in transducing As(III) mediated signals. The data presented indicates the differential responses in both the varieties and also that the increased tolerance in P. Bold may be due to the defensive role of antioxidant enzymes, induction of MAPK and up regulation of PCS transcript which is responsible for the production of metal binding peptides.

The effect of TDZ on organogenesis and somatic embryogenesis in pigeonpea (Cajanus cajan L. Millsp)

Abstract The effect of TDZ was studied on seedlings of pigeonpea. Seedlings raised from decoated seeds on MS basal medium supplemented with a low concentration of TDZ (0.05–1.0 μM) induced multiple shoots, and an intermediary concentration (5.0 μM) produced clusters of leafy structures, and a higher concentration (10.0 or 20.0 μM) completely switched the regeneration pathway by inducing somatic embryos at the cotyledonary nodal region that developed into mature plants. Seedlings raised from seeds with intact seed coats, failed to produce multiple shoots or embryos under the same conditions. Multiple shoots were also induced from seeds exposed for short duration (24 h–14 days) on MS basal medium containing 10.0 μM TDZ and then transferred to MS basal medium. However, continuous exposer to TDZ is required for somatic embryogenesis. Histological examination revealed direct shoot organogenesis and somatic embryogenesis. All the shoots developed prolific roots at their basal ends on MS basal medium containing 2.5 μM IBA. The well-developed plantlets were established in pots containing soil with 95% survival rate, and all the surviving plants were morphologically normal to plants raised from seeds. The present protocol is simple, rapid (the initiation of tissue cultures to transplantation of regenerants to soil completed in 8 week) with high regeneration frequency (75%) and applicable to seven genotypes.

Antioxidant value addition in human diets: genetic transformation of Brassica juncea with γ-TMT gene for increased α -tocopherol content

Abstractα-Tocopherol, the most biologically active form of vitamin E, is implicated in decreasing the risk of several types of cancers, coronary heart disease and a number of degenerative human conditions, when taken in excess of the recommended daily allowance. Natural α-tocopherol has twice the bioavailability of the synthetic isomer. This study describes a successful attempt at fortifying human diets with natural α-tocopherol by taking recourse to genetic engineering of an important oilseed crop, Brassica juncea. γ-Tocopherol methyl transferase cDNA from Arabidopsis thaliana, coding for the enzyme catalysing the conversion of the large γ-tocopherol pool to α-tocopherol, was overexpressed in B. juncea plants. The successful integration of the transgene was confirmed by PCR and Southern blot analysis, while the enhanced transcript level was evident in the northern blot analysis. HPLC analysis of the seeds of the T1 transgenic lines showed a shift in tocopherol profile with the highest over-expressors having α-tocopherol levels as high as sixfold over the non-transgenic controls. This study discusses the production of a transgenic oilseed crop with high α-tocopherol levels, which can provide a feasible, innocuous, and inexpensive way of taking the beneficial effects of high α-tocopherol intake to the masses.

Modulation of antioxidant machinery in α-tocopherol-enriched transgenic Brassica juncea plants tolerant to abiotic stress conditions

The antioxidant machinery in plants consists of several components with unique or overlapping functions that combat the deleterious production of reactive oxygen species (ROS) induced by stress conditions. Tocopherols are a group of powerful antioxidants having additional roles in signaling and gene expression, with α-tocopherol being the most potent form. In the present study, we used wild-type (WT) and α-tocopherol-enriched transgenic (TR) Brassica juncea plants grown under salt, heavy metal, and osmotic stress to compare their relative tolerance to these stresses and to assess the effects of increased α-tocopherol content on the other antioxidative enzymes and molecules. The oxidative damage caused by induced stress was lower in TR plants compared to WT plants as assessed by their higher relative water content and lower electrolyte leakage, malondialdehyde content as well as H2O2 accumulation. Lesser superoxide and H2O2 accumulation was also observed by histochemical staining in TR seedlings exposed to stress. Though no significant differences were evident under normal growth conditions, TR plants showed higher activities and transcript levels of antioxidant enzymes superoxide dismutase, catalase, ascorbate peroxidase, and glutathione reductase than WT plants under similar stress conditions. A decrease in ascorbate and glutathione content with marginally higher reductive ratios of these compounds was also observed in TR plants under the stress conditions. Our findings implicate the role of higher α-tocopherol levels in conferring better tolerance against salt, heavy metal, and osmotic stresses and also establish the existence of interplay between this lipid-soluble antioxidant and other water-soluble components of plant antioxidant defense.

Heavy metal induced DNA changes in aquatic macrophytes: Random amplified polymorphic DNA analysis and identification of sequence characterized amplified region marker

Plants have been used as good bio-indicators and genetic toxicity of environmental pollution in recent years. In this study, aquatic plants Hydrilla verticillata and Ceratophyllum demersum treated with 10 micromol/L Cd, 5 micromol/L Hg, and 20 micromol/L Cu for 96 h, showed changes in chlorophyll, protein content, and in DNA profiles. The changes in DNA profiles included variation in band intensity, presence or absence of certain bands and even appearance of new bands. Genomic template stability test performed for the qualitative measurement of changes in randomly amplified polymorphic DNA (RAPD) profiles, showed significant effect at the given concentration of metals. Cloning and sequencing of bands suggested that these markers although may not be homologous to any known gene but its conversion as a sequence characterized amplified region (SCAR) marke is useful in detecting the effects of genotoxin agents.

Generation of protective immune response against anthrax by oral immunization with protective antigen plant-based vaccine

In concern with frequent recurrence of anthrax in endemic areas and inadvertent use of its spores as biological weapon, the development of an effective anthrax vaccine suitable for both human and veterinary needs is highly desirable. A simple oral delivery through expression in plant system could offer promising alternative to the current methods that rely on injectable vaccines extracted from bacterial sources. In the present study, we have expressed protective antigen (PA) gene in Indian mustard by Agrobacterium-mediated transformation and in tobacco by plastid transformation. Putative transgenic lines were verified for the presence of transgene and its expression by molecular analysis. PA expressed in transgenic lines was biologically active as evidenced by macrophage lysis assay. Intraperitoneal (i.p.) and oral immunization with plant PA in murine model indicated high serum PA specific IgG and IgA antibody titers. PA specific mucosal immune response was noted in orally immunized groups. Further, antibodies indicated lethal toxin neutralizing potential in-vitro and conferred protection against in-vivo toxin challenge. Oral immunization experiments demonstrated generation of immunoprotective response in mice. Thus, our study examines the feasibility of oral PA vaccine expressed in an edible plant system against anthrax.

The Xerophyta viscosa Aldose Reductase (ALDRXV4) Confers Enhanced Drought and Salinity Tolerance to Transgenic Tobacco Plants by Scavenging Methylglyoxal and Reducing the Membrane Damage

We report the efficacy of an aldose reductase (ALDRXV4) enzyme from Xerophyta viscosa Baker in enhancing the prospects of plant’s survival under abiotic stress. Transgenic tobacco plants overexpressing ALDRXV4 cDNA showed alleviation of NaCl and mannitol-induced abiotic stress. The transgenic plants survived longer periods of water deficiency and salinity stress and exhibited improved recovery after rehydration as compared to the wild type plants. The increased synthesis of aldose reductase in transgenic plants correlated with reduced methylglyoxal and malondialdehyde accumulation and an elevated level of sorbitol under stress conditions. In addition, the transgenic lines showed better photosynthetic efficiency, less electrolyte damage, greater water retention, higher proline accumulation, and favorable ionic balance under stress conditions. Together, these findings suggest the potential of engineering aldose reductase levels for better performance of crop plants growing under drought and salt stress conditions.

Litchi Breeding for Genetic Improvement

Litchi chinensis Sonn. ranks (high) among the most important horticultural crops, belongs to the family Sapindaceae and widely grown in tropical and subtropical regions (Menzel 1985). The tree produces delicious top quality fruits that are in great demand for their wholesome taste, sweet aroma and attractive colour. They are mainly consumed as a fresh table fruit worldwide but in China, dried litchis, called litchi nuts with the taste of the raisin are quite popular. They are also preserved and canned in syrup or used as squash. In Florida, frozen fruits are consumed on a limited scale. China, India and Taiwan are the major producers of litchi whereas in the last 40 years substantial increase in production in South Africa, Australia, Thailand, Vietnam, USA and Israel has led litchis to become a significant commodity in the international trade, (Underhill et al. 1997). Major thrust of litchi research has been on prevention of physiological browning and retention of bright red colouration of litchi fruits in several countries. Exotic litchi fruits have received worldwide attention. Increase in popularity has necessitated litchi cultivation in a wide range of environmental conditions. In fact litchi is generally adapted to various soil types via alluvial sands, loams, heavy clay, organic soil and calcareous soil with 30% lime and rock files (Chapman 1984a). In China, the best litchi trees are prevalent in Gwanagdong province close to the rivers on alluvial sands with good drainage and access to the water table (Chapman 1984b). They are also grown in gravelly sandy loam to loam soils as well as in swampy areas. However, soil in Fijian province is very high in clay, poorly drained and acidic in reaction (Winks et al. 1983). In South Africa, trees are more vigorous in growth on acid soils rather than on neutral or alkaline soils (Marloth 1947). Under Indian conditions, litchi cultivation in Bihar state is common on calcareous soils containing more than 40% free calcium carbonate and trees flourish well in a moist subtropical climate and in deep loamy soil with high moisture content. However,

Stress-inducible overexpression of glyoxalase I is preferable to its constitutive overexpression for abiotic stress tolerance in transgenic Brassica juncea

The glyoxalase system catalyzes the conversion of cytotoxic methylglyoxal to d-lactate via the intermediate S-d-lactoylglutathione. It comprises two enzymes, Glyoxalase I (Gly I) and Glyoxalase II (Gly II), and reduced glutathione which acts as a cofactor by anchoring the substrates in the active sites of the two enzymes. The overexpression of both Gly I and Gly II, either alone or in combination, has earlier been reported to confer tolerance to multiple abiotic stresses. In the present study, we sought to evaluate the consequences of constitutive and stress-induced overexpression of Gly I on the performance and productivity of plants. Towards this end, several Gly I transgenic Brassica juncea lines (designated as R and S lines) were generated in which the glyoxalase I (gly I) gene was expressed under the control of either a stress-inducible rd29A promoter or a constitutive CaMV 35S promoter. Both the R and S lines showed enhanced tolerance to salinity, heavy metal, and drought stress when compared to untransformed control plants. However, the S lines showed yield penalty under non-stress conditions while no such negative effect was observed in the R lines. Our results indicate that the overexpression of the gly I gene under the control of stress-inducible rd29A promoter is a better option for improving salt, drought and heavy metal stress tolerance in transgenic plants.