NOPR Collection: Pages 557-620

A microdroplet cell culture based high frequency somatic embryogenesis system for pigeonpea, Cajanus cajan (L.) Millsp.

2016-07-20T05:26:30Z

Title: A microdroplet cell culture based high frequency somatic embryogenesis system for pigeonpea, Cajanus cajan (L.) Millsp. Authors: Kumar, Nagan Udhaya; Gnanaraj, Muniraj; Sindhujaa, Vajravel; Viji, Maluventhen; Manoharan, Kumariah Abstract: A protocol for high frequency production of somatic embryos was worked out in pigeonpea, Cajanus cajan (L.) Millsp. The protocol involved sequential employment of embryogenic callus cultures, low density cell suspension cultures and a novel microdroplet cell culture system. The microdroplet cell cultures involved culture of a single cell in 10 µl of Murashige and Skoog’s medium supplemented with phytohormones, growth factors and phospholipid precursors. By employing the microdroplet cell cultures, single cells in isolation were grown into cell clones which developed somatic embryos. Further, 2,4-dichlorophenoxyacetic acid, kinetin, polyethylene glycol, putrescine, spermine, spermidine, choline chloride, ethanolamine and LiCl were supplemented to the low density cell suspension cultures and microdroplet cell cultures to screen for their cell division and somatic embryogenesis activity. Incubation of callus or the inoculum employed for low density cell suspension cultures and microdroplet cell cultures with polyethylene glycol was found critical for induction of somatic embryogenesis. Somatic embryogenesis at a frequency of 1.19, 3.16 and 6.51 per 10⁶cells was achieved in the callus, low density cell suspension cultures and microdroplet cell cultures, respectively. Advantages of employing microdroplet cell cultures for high frequency production of somatic embryos and its application in genetic transformation protocols are discussed. Page(s): 600-610

Characterization of coal fly ash nanoparticles and their induced in vitro cellular toxicity and oxidative DNA damage in different cell lines

2016-07-20T05:16:27Z

Title: Characterization of coal fly ash nanoparticles and their induced in vitro cellular toxicity and oxidative DNA damage in different cell lines Authors: Sambandam, Bharathi; Devasena, Thiyagarajan; Islam, Villianur Ibrahim Hairul; Prakhya, Balkrishna Murthy Abstract: Coal combustion generates considerable amount of ultrafine particles and exposure to such particulate matter is a major health concern in the developing countries. In this study, we collected nano sized coal fly ash (CFA) and characterized them by scanning electron microscope-energy dispersive X-ray analysis (SEM-EDX), particle size analyzer (PSA) and transmission electron microscope (TEM), and investigated its toxicity in vitro using different cell lines. The imaging techniques showed that the coal fly ash nanoparticles (CFA-NPs) are predominately spherical shaped. The analyses have revealed that the CFA-NPs are 7-50 nm in diameter and contain several heavy metals associated with CFA particles. The studies showed significant amount of toxicity in all cell lines on treatment with CFA-NPs. The cytotoxicity and oxidative DNA damage caused by CFA-NPs were determined by inhibition of cellular metabolism (MTT), total intracellular glutathione (GSH), reactive oxygen species (ROS) and DNA fragmentation in cultured cell lines (Chang liver, HS294T and LL29). The cellular metabolism was inhibited in a dose-dependent manner in CFA-NPs treated cell lines. The CFA-NPs induced ROS and decreased the total intracellular glutathione with increased dose. Further, the CFA-NPs treated cells showed severe DNA laddering as a result of DNA fragmentation. Page(s): 585-593

Ginger extract ameliorates phosphamidon induced hepatotoxicity

2016-07-20T05:20:45Z

Title: Ginger extract ameliorates phosphamidon induced hepatotoxicity Authors: Mukherjee, Suprabhat; Mukherjee, Niladri; Saini, Prasanta; Roy, Priya; Babu, Santi P. Sinha Abstract: Organophosphorus (OP) compounds commonly used as pesticides in agriculture cause serious health problems to living beings. The present study enumerates the ameliorating effect of ginger extract (GE) against phosphamidon (PHO, an organophosphorus insecticide) induced hepatotoxicity. GE was prepared from dried ginger and characterized for compound profile and antioxidant activity. Eight groups of albino rats (n=6) were treated with 1/5^th lethal dose of PHO for 5-20 days. Out of the treated 8 groups, 4 were simultaneously fed with GE (1 mg/kg body wt.) along with PHO. Alterations in the levels of hepatocellular oxidative stress (OS) markers in the treated groups indicated an enhanced generation of reactive oxygen species (ROS) and oxidative stress (OS). Upregulation of apoptotic markers, DNA fragmentation and appearance of apoptotic nuclei suggested induction of apoptosis in the liver cell that was found to be attenuated after GE treatment. Moreover, no toxicity and mortality was observed up to 100 mg/kg dose of GE for 30 days in the rat model studied. Thus, GE can be considered as an effective, economical and safe extract to circumvent PHO-induced hepatotoxicity. Page(s): 574-584

Q2N and E64G double mutation of ubiquitin confers a stress sensitive phenotype on Saccharomyces cerevisiae

2016-07-20T05:31:29Z

Title: Q2N and E64G double mutation of ubiquitin confers a stress sensitive phenotype on Saccharomyces cerevisiae Authors: Sharma, Mrinal; Prabha, C Ratna Abstract: The eukaryotic protein, ubiquitin harbours a parallel β-bulge in its structure which is formed by residues Glu64(1), Ser65(2) and Gln2(X). Despite their low % frequency of occurrence in parallel β-bulges, the residues Gln2 and Glu64 have been totally conserved in ubiquitin. In a previous study, two single mutants UbQ2N and UbE64G were constructed by replacing the residues Gln2 and Glu64 with Asn and Gly, respectively to understand their importance. The choice of the residues for substitution was made on the basis of their high preference for existence in parallel β-bulge, so that the structure of mutants remains unaltered and any functional differences observed would highlight the importance of Gln2 and Glu64 in ubiquitin biology. The results from this study established that yeast cells expressing either UbQ2N or UbE64G, displayed functional differences with respect to survival upon exposure to cycloheximide and degradation of substrates by ubiquitin fusion degradation (UFD) pathway. It describes construction of the double mutant UbQ2N-E64G and its characterization. Our results showed expression of UbQ2N-E64G in stress hypersensitive SUB60 cells led to significant decrease in growth rate and prolonged half-life of substrates of UFD pathway, besides failure of complementation under heat and antibiotic stresses, providing the reason for conservation of Gln2 and Glu64 in ubiquitin sequence. Page(s): 617-620