Preincubation of cut tobacco leaf explants promotes Agrobacterium-mediated transformation by increasing vir gene induction

The effects of preincubation of cut tobacco leaf explants on Agrobacterium transformation efficiency and induction of Agrobacterium virE-lacZ fusion were evaluated. Transformation efficiency was evaluated by histochemical and fluorometric analysis of beta-glucuronidase in leaf rings transformed with Agrobacterium tumefaciens strain LBA4404(pKIWI105). The transformation efficiency increased by 2-fold, 5-fold, and 4.3-fold upon preincubation for 24, 48, and 72 h, respectively. Preincubation for 24, 48, and 72 h increased the ability of tobacco leaf segments to induce Agrobacterium virE by 2.3-fold, 3.5-fold and 4.5-fold, respectively. The requirement of preincubation for increased transformation efficiency was obviated by the addition of 100 mu M acetosyringone to the freshly cut leaf rings cocultivated with Agrobacterium. The production of vii gene inducers by the leaf rings during the preincubation period is an important factor that contributes to increased transformation efficiency of Agrobacterium upon preincubation. (C) 1999 Elsevier Science Ireland Ltd. All rights reserved.

Expression pattern of Cubitus interruptus from the mulberry silkworm Bombyx mori in late developmental stages

A partial genomic clone of Bombyx mori homologue of the segment polarity gene Cubitus interruptus (BmCi), encoding the conserved zinc finger domain and harbouring two introns, has been characterized. BmCi was expressed in the silkglands of B. mori from embryonic to the late larval stages(3rd, 4th and 5th intermoults). The expression was confined to the anterior region of the middle silkglands, overlapping with the domain of sericin-2 expression and excluding the domains of Bm invected expression, namely the middle and posterior regions of the middle silkglands. In the wing discs, the expression was restricted to the anterior compartment, which increased from 4th to 5th larval intermoults and declined later in the pupal wing buds. In gonadal tissues (both ovaries and testes) BmCi was expressed from the larval to pupal stages. The transcripts were localized to the sperm tubes containing spermatogonia in the testis of Bombyx larvae. BmCi expression, however, was not detected in any of these tissues during the moulting stages. Expression of Ci in the wing discs and gonads is evolutionarily conserved, while the silkgland represents a novel domain. Our results imply that BmCi is involved in the specification and maintenance of micro-compartment identity within the middle silkglands.

Spatial filter based 3D resolution improvement and polarization properties of multiphoton multiple-excitation-spot-optical microscopy

Three-dimensional (3D) resolution improvement in multi-photon multiple-excitation-spot-optical microscopy is proposed. Specially designed spatial filter is employed for improving the overall 3D resolution of the imaging system. An improvement up to a factor of 14.5 and sub-femto liter volume excitation is achieved. The system shows substantial sidelobe reduction (<4%) due to the non-linear intensity dependence of multiphoton process. Polarization effect on x-oriented and freely rotating dipoles shows dramatic change in the field distribution at the focal-plane. The resulting point-spread function has the ability to produce several strongly localized polarization dependent field patterns which may find applications in optical engineering and bioimaging.

High resolution multiple excitation spot optical microscopy

We propose fundamental improvements in three-dimensional (3D) resolution of multiple excitation spot optical microscopy. The excitation point spread function (PSF) is generated by two interfering counter-propagating depth-of-focus beams along the optical axis. Detection PSF is obtained by coherently interfering the emitted fluorescent light (collected by both the objectives) at the detector. System PSF shows upto 14-fold reduction in focal volume as compared to confocal, and almost 2-fold improvement in lateral resolution. Proposed PSF has the ability to simultaneously excite multiple 3D-spots of sub-femtoliter volume. Potential applications are in fluorescence microscopy and nanobioimaging. Copyright 2011 Author(s). This article is distributed under a Creative Commons Attribution 3.0 Unported License. [doi:10.1063/1.3598413]

High resolution facies record on late Holocene flood plain sediments from lower reaches of Narmada Valley, Western India

A high resolution quantitative granulometric record for site Uchediya 21A degrees 43'2.22aEuro(3) N, 73A degrees 6'26.22aEuro(3) E; 10 m a. s. l.] gives understanding towards accretion history of the late Holocene flood plain in the lower reaches of Narmada River. Two sediment facies (sandy and muddy) and seven subfacies (sandy subfacies: St(MS+FS+CS), SmFS+MS, Sl(FS+VFS), and St(MS + CS); muddy subfacies: FmSILT+VFS+FS, FmSILT+VFS (O) and FmSILT+VFS (T)) are identified based on cluster analysis supplemented with sedimentary structures observed in field and other laboratory data. Changes in hydrodynamics are further deduced based on various sedimentological parameters and their ratios leading to arrive at a depositional model.

Anticancer property of Bryophyllum pinnata (Lam.) Oken. leaf on human cervical cancer cells

Background: Bryophyllum pinnata (B. pinnata) is a common medicinal plant used in traditional medicine of India and of other countries for curing various infections, bowel diseases, healing wounds and other ailments. However, its anticancer properties are poorly defined. In view of broad spectrum therapeutic potential of B. pinnata we designed a study to examine anti-cancer and anti-Human Papillomavirus (HPV) activities in its leaf extracts and tried to isolate its active principle. Methods: A chloroform extract derived from a bulk of botanically well-characterized pulverized B. pinnata leaves was separated using column chromatography with step-gradient of petroleum ether and ethyl acetate. Fractions were characterized for phyto-chemical compounds by TLC, HPTLC and NMR and Biological activity of the fractions were examined by MTT-based cell viability assay, Electrophoretic Mobility Shift Assay, Northern blotting and assay of apoptosis related proteins by immunoblotting in human cervical cancer cells. Results: Results showed presence of growth inhibitory activity in the crude leaf extracts with IC50 at 552 mu g/ml which resolved to fraction F4 (Petroleum Ether: Ethyl Acetate:: 50: 50) and showed IC50 at 91 mu g/ml. Investigations of anti-viral activity of the extract and its fraction revealed a specific anti-HPV activity on cervical cancer cells as evidenced by downregulation of constitutively active AP1 specific DNA binding activity and suppression of oncogenic c-Fos and c-Jun expression which was accompanied by inhibition of HPV18 transcription. In addition to inhibiting growth, fraction F4 strongly induced apoptosis as evidenced by an increased expression of the pro-apoptotic protein Bax, suppression of the anti-apoptotic molecules Bcl-2, and activation of caspase-3 and cleavage of PARP-1. Phytochemical analysis of fraction F4 by HPTLC and NMR indicated presence of activity that resembled Bryophyllin A. Conclusions: Our study therefore demonstrates presence of anticancer and anti-HPV an activity in B. pinnata leaves that can be further exploited as a potential anticancer, anti-HPV therapeutic for treatment of HPV infection and cervical cancer.

Biochemical characterization of C4 protein of cotton leaf curl Kokhran virus-Dabawali

Background: Cotton leaf curl Kokhran Virus-Dabawali (CLCuKV-Dab) is a monopartite begomovirus encoding two proteins V1 and V2 in the virion sense and four proteins Cl, C2, C3 and C4 in the complementary sense. The C4 protein of monopartite begomoviruses has been implicated to play a role in symptom determination and virus movement. The present work aims at the biochemical characterization of this protein. Methods: The C4 protein of CLCuKV-Dab was purified in fusion with GST and tested for the ability to hydrolyze ATP and other phosphate containing compounds. ATPase activity was assayed by using radiolabeled gamma-32P]-ATP and separating the product of reaction by thin layer chromatography. The hydrolysis of other compounds was monitored by the formation of a blue colored phosphomolybdate complex which was estimated by measuring the absorbance at 655 nm. Results: The purified GST-C4 protein exhibited metal ion dependent ATPase and inorganic pyrophosphatase activities. Deletion of a sequence resembling the catalytic motif present in phosphotyrosine phosphatases resulted in 70% reduction in both the activities. Mutational analysis suggested arginine 13 to be catalytically important for the ATPase and cysteine 8 for the pyrophosphatase activity of GST-C4. Interaction of V2 with GST-C4 resulted in an increase in both the enzymatic activities of GST-C4. Conclusions: The residues important for the enzymatic activities of GST-C4 are present in a motif different from the classical Walker motifs and the non-classical ATP binding motifs reported so far. General significance: The C4 protein of CLCuKV-Dab, a putative natively unfolded protein, exhibits enzymatic activities.

Phylogenetic analysis and molecular dating suggest that hemidactylus anamallensis is not a member of the hemidactylus radiation and has an ancient late cretaceous origin

Background of the Work: The phylogenetic position and evolution of Hemidactylus anamallensis (family Gekkonidae) has been much debated in recent times. In the past it has been variously assigned to genus Hoplodactylus (Diplodactylidae) as well as a monotypic genus `Dravidogecko' (Gekkonidae). Since 1995, this species has been assigned to Hemidactylus, but there is much disagreement between authors regarding its phylogenetic position within this genus. In a recent molecular study H. anamallensis was sister to Hemidactylus but appeared distinct from it in both mitochondrial and nuclear markers. However, this study did not include genera closely allied to Hemidactylus, thus a robust evaluation of this hypothesis was not undertaken. Methods: The objective of this study was to investigate the phylogenetic position of H. anamallensis within the gekkonid radiation. To this end, several nuclear and mitochondrial markers were sequenced from H. anamallensis, selected members of the Hemidactylus radiation and genera closely allied to Hemidactylus. These sequences in conjunction with published sequences were subjected to multiple phylogenetic analyses. Furthermore the nuclear dataset was also subjected to molecular dating analysis to ascertain the divergence between H. anamallensis and related genera. Results and Conclusion: Results showed that H. anamallensis lineage was indeed sister to Hemidactylus group but was separated from the rest of the Hemidactylus by a long branch. The divergence estimates supported a scenario wherein H. anamallensis dispersed across a marine barrier to the drifting peninsular Indian plate in the late Cretaceous whereas Hemidactylus arrived on the peninsular India after the Indian plate collided with the Eurasian plate. Based on these molecular evidence and biogeographical scenario we suggest that the genus Dravidogecko should be resurrected.

Failure modes and fatigue behavior of conventional and refilled friction stir spot welds in AA 6061-T6 sheets

The fatigue behavior of conventional friction stir spot welding (FSSW) and friction stir spot welding refilled by the friction forming process (FSSW-FFP) in aluminum 6061-T6 lap shear specimens, are investigated based on the experimental observations. Optical micrographs of the welds after fatigue failure in both the cases are examined to study the fatigue crack propagation and failure modes. Experimental results indicate that the fatigue strength of the FSSW-FFP weld samples is higher than that of the conventional FSSW samples at all loads. Fracture surfaces are analyzed in detail using the scanning electron microscope. (C) 2013 Elsevier Ltd. All rights reserved.

Static shear strength and fatigue life of refill friction stir spot welded AA 6061-T6 sheets

This study was aimed at evaluating the static shear strength and fatigue properties of the newly developed refilled friction stir spot welded AA 6061-T6 joints. The keyhole, the process disadvantage of conventional friction stir spot welding, was refilled successfully, using an additional filler plate, with specially designed tools. Two different tool profiles, namely, convex and concave, were used for the refilling process. Sound and defect free joints were obtained by the refilling process. Joints refilled with convex tools showed better static shear strength than those with the concave ones. The variation of microhardness in different regions of the weld was analysed. Fatigue tests were conducted on the lap shear specimens at a stress ratio of R=0.1. The optical micrographs of the welds after fatigue failure in both the conventional and refilled processes were examined to study the fatigue crack propagation and failure modes.

Anti-inflammatory activity of the ethanol extract of Dictamnus dasycarpus leaf in lipopolysaccharide-activated macrophages

Background: Dictamnus dasycarpus is widely used as a traditional remedy for the treatment of eczema, rheumatism, and other inflammatory diseases in Asia. The current study investigates the molecular mechanism of anti-inflammatory action of the ethanol extract of Dictamnus dasycarpus leaf (DE) in lipopolysaccharide (LPS)-stimulated RAW 264.7 cells. Methods: Nitric oxide (NO) production was assessed by Griess reaction and the mRNA and protein expressions of pro inflammatory cytokines, transcription factor, and enzymes were determined by real-time RT-PCR and immunoblotting analysis. Results: DE (0.5 and 1 mg/mL) suppressed the NO production by 10 and 33%, respectively, compared to the untreated group in LPS-stimulated RAW 264.7 cells. DE (0.5 and 1 mg/mL) reduced the mRNA expression of key transcription factor nuclear factor-kappa B by 7 and 24%, respectively compared to the untreated group in LPS activated macrophage. The pro inflammatory cytokines such as tumor necrosis factor a and interleukin 1 beta were also decreased by DE treatment. Moreover, the protein expression of pro inflammatory enzymes, inducible nitric oxide synthase and cyclooxygenase 2 were also dramatically attenuated by DE in a dose dependent manner. Conclusions: These results suggest that Dictamnus dasycarpus leaf has a potent anti-inflammatory activity and can be used for the development of new anti-inflammatory agents.