High efficiency in planta transformation of Indian mustard (Brassica juncea) based on spraying of floral buds

Plant Cell, Tissue and Organ Culture (PCTOC) - Brassica juncea (Indian mustard) and its sub-varieties include whole range of oilseed and vegetable mustard in China, Canada, Australia, Europe and...     

Fabrication of iron oxide nanocolloids using metallosurfactant-based microemulsions: antioxidant activity,cellular, and genotoxicity toward Vitis vinifera

The present work aims at the fabrication of iron oxide nanocolloids using biocompatible microemulsion and their cytotoxic, genotoxic effect on Vitis vinifera plant has been evaluated. The three iron-based metallosurfactant complexes were synthesized. Nanosuspensions (Ns) were prepared using microemulsion technique and for the purpose, the microemulsion was prepared using oleic acid, butanol, tween 80 and as synthesized iron metallosurfactant. In this technique, no additional capping agent and/or reducing agent was added. Tween 80 which is a biocompatible surfactant acted as a reducing agent as well as stabilizing for the iron oxide Ns. Characterization of Ns’s was done using TEM, FESEM, EDX, XRD, AFM, and zeta potential. Mixed type of iron oxide nanoparticles i.e. magnetite (Fe₃O₄), and maghemite (Fe₂O₃) with a size range of 1–16 nm was found to be present in the nanosuspensions prepared from all the three precursors. The antioxidant activity of the Fe Ns was also confirmed using DPPH assay, with order of activity FeDDA > FeCTAC > FeHEXA. The cellular toxicity of Ns was evaluated by observing the morphological changes on V. vinifera plant (petiole) using a light microscope. Further, the interactions of iron oxide Ns with V. vinifera’s DNA (plant-DNA) was assessed using circular dichroism (CD) and gel electrophoresis. For the case of FeCTAC Ns, a decrease in the intensity of bands was observed indicating fragmentation or adduct formation resulting in DNA damage. In the case of FeDDA, a modest decrease in the intensity of bands was observed. However, for FeHEXA Ns, complete neutralization of bands was confirmed implying maximum damage to the plant DNA. CD, gel electrophoresis and antioxidant activity confirmed that FeHEXA Ns were most toxic and FeDDA Ns were safest among the three as-fabricated nanosuspensions.     

Salt stress enhanced antioxidant response in callus of three halophytes (Salsola baryosma, Trianthema triquetra, Zygophyllum simplex) of Thar Desert

Effects of salinity on growth, protein content, proline, catalase and antioxidant enzyme activity in callus of three halophytes of the Thar Desert; Salsola baryosma, Trianthema triquetra and Zygophyllum simplex were evaluated. Callus tissues were cultured on Murashige and Skoog’s medium containing different concentrations of NaCl (50, 100 and 200 mM). Increase in dry weight and soluble proteins were observed in the callus exposed to lower salinity (50 and 100 mM NaCl) in all the three species, whereas on the medium containing 200 mM NaCl, significant decrease in these two growth parameters was recorded. Under the salinity stress maximum proline accumulation was found in S. baryosma with parallel increase in soluble sugars. Among the three species, T. triquetra callus showed maximum CAT activity with 50 and 100 mM NaCl treatment, whereas the enzyme activity decreased at 200 mM NaCl treatment in all three species. The antioxidant potential steadily elevated under salt treatment in all the above three species using 1, 1-diphenyl-2-picrylhydrazyl (DPPH) and ferric reducing antioxidant potential (FRAP) assay. Whereas, superoxide dismutase (SOD) quenching were recorded maximum at low (50 and 100 mM) concentrations in all the three species. However, T. triquetra callus showed maximum total phenolic content (TPC) 15 mg GAE g^?1 with the elevated concentration of NaCl up to 200 mM, and S. baryosma callus showed lower TPC as compared to both species. A significant correlation between antioxidant capacity and TPC was observed indicating that phenolic compounds are the major contributors to the antioxidant potential in these halophyte species. FRAP and DPPH activity of Z. simplex showed maximum correlation (R = 0.992), as compared to other two species. We can conclude that all the three species exhibit a protection mechanism by sustaining growth parameters and antioxidant capacity. Due to high antioxidant property of all these species, the plant extracts may be included in nutraceutical formulations.     

Micropropagtion of Terminalia bellerica from nodal explants of mature tree and assessment of genetic fidelity using ISSR and RAPD markers

The present study reports an efficient in vitro micropropagation protocol for a medicinally important tree, Terminalia bellerica Roxb. from nodal segments of a 30 years old tree. Nodal segments taken from the mature tree in March-April and cultured on half strength MS medium gave the best shoot bud proliferation response. Combinations of serial transfer technique (ST) and incorporation of antioxidants (AO) [polyvinylpyrrolidone, PVP (50 mg l⁻¹) + ascorbic acid (100 mg l⁻¹) + citric acid (10 mg l⁻¹)] in the culture medium aided to minimize browning and improve explant survival during shoot bud induction. Highest multiplication of shoots was achieved on medium supplemented with 6-benzyladenine (BA, 8.8 μM) and α-naphthalene acetic acid (NAA, 2.6 μM) in addition to antioxidants. Shoot elongation was obtained on MS medium containing BA (4.4 μM) + phloroglucinol (PG, 3.9 μM). Elongated shoots were transferred to half strength MS medium containing indole-3-butyric acid (IBA, 2.5 μM) for root development. The acclimatization of plantlets was carried out under greenhouse conditions. The genetic fidelity of the regenerated plants was checked using inter simple sequence repeats (ISSR) and randomly amplified polymorphic DNA (RAPD) analysis. Comparison of the bands among the regenerants and mother plant confirmed true-to-type clonal plants.     

Recombinant Modified Vaccinia Virus Ankara Expressing Glycoprotein E2 of Chikungunya Virus Protects AG129 Mice against Lethal Challenge

Chikungunya virus (CHIKV) infection is characterized by rash, acute high fever, chills, headache, nausea, photophobia, vomiting, and severe polyarthralgia. There is evidence that arthralgia can persist for years and result in long-term discomfort. Neurologic disease with fatal outcome has been documented, although at low incidences. The CHIKV RNA genome encodes five structural proteins (C, E1, E2, E3 and 6K). The E1 spike protein drives the fusion process within the cytoplasm, while the E2 protein is believed to interact with cellular receptors and therefore most probably constitutes the target of neutralizing antibodies. We have constructed recombinant Modified Vaccinia Ankara (MVA) expressing E3E2, 6KE1, or the entire CHIKV envelope polyprotein cassette E3E26KE1. MVA is an appropriate platform because of its demonstrated clinical safety and its suitability for expression of various heterologous proteins. After completing the immunization scheme, animals were challenged with CHIV-S27. Immunization of AG129 mice with MVAs expressing E2 or E3E26KE1 elicited neutralizing antibodies in all animals and provided 100% protection against lethal disease. In contrast, 75% of the animals immunized with 6KE1 were protected against lethal infection. In conclusion, MVA expressing the glycoprotein E2 of CHIKV represents as an immunogenic and effective candidate vaccine against CHIKV infections.     

Terminal end bud maintenance in mammary gland is dependent upon FGFR2b signaling

We previously demonstrated that Fibroblast Growth Factor 10 (FGF10) and its receptor FGFR2b play a key role in controlling the very early stages of mammary gland development during embryogenesis [Mailleux, A.A., Spencer-Dene, B., Dillon, C., Ndiaye, D., Savona-Baron, C., Itoh, N., Kato, S., Dickson, C., Thiery, J.P., and Bellusci, S. (2002). Role of FGF10/FGFR2b signaling during mammary gland development in the mouse embryo. Development 129, 53-60. Veltmaat, J. M., Relaix, F., Le, L.T., Kratochwil, K., Sala, F.G., van Veelen, W., Rice, R., Spencer-Dene, B., Mailleux, A.A., Rice, D.P., Thiery, J.P., and Bellusci, S. (2006). Gli3-mediated somitic Fgf10 expression gradients are required for the induction and patterning of mammary epithelium along the embryonic axes. Development 133, 2325-35.]. However, the role of FGFR2b signaling in postnatal mammary gland development is still elusive. We show that FGF10 is expressed at high level throughout the adipose tissue in the mammary gland of young virgin female mice whereas its main receptor FGFR2 is found mostly in the epithelium. Using a rtTA transactivator/tetracycline promoter approach allowing inducible and reversible attenuation of the FGFR2b signaling throughout the adult mouse, we are now reporting that FGFR2b signaling is also critical during postnatal mammary gland development. Ubiquitous attenuation of FGFR2b signaling in the postnatal mouse for 6 weeks starting immediately after birth is not lethal and leads to minor defects in the animal. Upon dissection of the mammary glands, a 40% reduction in size compared to the WT control is observed. Further examination shows a rudimentary mammary epithelial tree with completely absent terminal end buds (TEBs), compared to a well-branched structure observed in wild type. Transplantation of mammary gland explants into cleared fat pad of wild type mouse recipients indicates that the observed abnormal branching results from defective FGFR2b signaling in the epithelium. We also demonstrate that this rudimentary tree reforms TEBs and resumes branching upon removal of doxycycline suggesting that the regenerative capacities of the mammary epithelial progenitor cells were still functional despite long-term inactivation of the FGFR2b pathway. At the cellular level, upon FGFR2b attenuation, we show an increase in apoptosis associated with a decrease in the proliferation of the mammary luminal epithelium. We conclude that during puberty, there is a differential requirement for FGFR2b signaling in ductal vs. TEBs epithelium. FGFR2b signaling is crucial for the survival and proliferation of the mammary luminal epithelial cells, but does not affect the regenerative potential of the mammary epithelial progenitor cells.     

Differential protection against MPTP or methamphetamine toxicity in dopamine neurons by deletion of ppN/OFQ expression

Nociceptin (N/OFQ) is an endogenous neuropeptide that plays a role in the behavioral deficits associated with Parkinson's disease (PD). The purpose of the present study was to characterize the protective effects of prepro (pp)N/OFQ gene deletion against two dopamine toxins, MPTP and methamphetamine (METH). Results demonstrate that ppN/OFQ gene deletion attenuates the loss of both the number of tyrosine hydroxylase (TH)-positive neurons in the substantia nigra pars compacta (SNpc) and loss of TH and vesicular monoamine transporter-2 (VMAT) immunoreactivity in the caudate putamen (CPu) of MPTP-treated mice. This protection was unaffected by age or gender, although, when loss of TH exceeded 90% in 5-6 month-old mice, the protective effect was greatly diminished. In contrast, METH administration preferentially damaged dopaminergic terminals in the CPu with little effect on dopamine neurons in the SNpc, an effect not reversed by ppN/OFQ gene deletion. To determine if N/OFQ and MPP+ act directly and synergistically on dopamine neurons, differentiated SH-SY5Y cells were incubated with N/OFQ and/or MPP+. N/OFQ did not increase MPP+-mediated cell loss, suggesting an indirect action of N/OFQ. These studies demonstrate that inhibition of the endogenous N/OFQ system may represent a new therapeutic target for prevention of neuronal loss associated with PD.