Optimisation of growth medium for the production of cyclodextrin glucanotransferase from Bacillus stearothermophilus HR1 using response surface methodology

Cyclodextrin glucanotransferase (CGTase) activity was observed when the bacterium was grown in the medium at various initial pH values, containing carbon, nitrogen, phosphorus and mineral salt sources at 50 °C for 24 h in the shake flasks. The optimisation of this growth medium was carried out using response surface methodology. The design contains a total of 32 experimental trials involving 10 star points and 6 replicates at the centre points. The design was employed by selecting sago starch, peptone from casein, K₂HPO₄, CaCl₂ and initial pH as five independent variables in this study. The optimal calculated values of tested variables for maximal production of CGTase were found to be comprised of: sago starch, 16.02 g/l; peptone from casein, 20 g/l; K₂HPO₄, 1.4 g/l; CaCl₂, 0.2 g/l and initial pH, 7.54 with a predicted CGTase activity of 14.20 U/ml. These predicted optimal parameters were tested in the laboratory and the final CGTase activity obtained was very close to the predicted value at 14.80 U/ml.     

Application of loop-mediated isothermal amplification (LAMP)-based technology for authentication of <Emphasis Type="Italic">Catharanthus roseus</Emphasis> (L.) G. Don

In this study, loop-mediated isothermal amplification (LAMP)-based molecular marker was developed for authentication of Catharanthus roseus, a medicinal plant. Samples of this plant were collected from different geographical locations in India. Random amplified polymorphic deoxyribonucleic acid (DNA) analysis of collected samples was carried out with 25 random primers. A 610-bp DNA fragment, common to all accessions, was eluted, cloned, and sequenced. Four LAMP primers were designed on the basis of sequence of 610 bp DNA fragment. LAMP reaction, containing 10× Bst DNA polymerase reaction buffer, Bst DNA polymerase, four in-house designed primers, dNTPs, MgSO₄, and betaine, was incubated at 65°C for 1 h. The resulting amplicon was visualized by adding SYBR Green I to the reaction tube. The data showed confirmatory results. Since the assay method is simple, sensitive, and cost-effective, it is a feasible method for identifying and authentication of C. roseus.     

Genetic diversity analysis of Zingiber Officinale Roscoe by RAPD collected from subcontinent of India

The present investigation was undertaken for the assessment of 12 accessions of Zingiber officinale Rosc. collected from subcontinent of India by RAPD markers. DNA was isolated using CTAB method. Thirteen out of twenty primers screened were informative and produced 275 amplification products, among which 261 products (94.90%) were found to be polymorphic. The percentage polymorphism of all 12 accessions ranged from 88.23% to 100%. Most of the RAPD markers studied showed different levels of genetic polymorphism. The data of 275 RAPD bands were used to generate Jaccard’s similarity coefficients and to construct a dendrogram by means of UPGMA. Results showed that ginger undergoes genetic variation due to a wide range of ecological conditions. This investigation was an understanding of genetic variation within the accessions. It will also provide an important input into determining resourceful management strategies and help to breeders for ginger improvement program.     

Optimization of methods for peripheral blood mononuclear cells isolation and expansion of human gamma delta T cells

Human Vg9/Vδ2 T cells (γδ T cells) are immune surveillance cells both in innate and adaptive immunity and are a possible target for anticancer therapies, which can induce immune responses in a variety of cancers. Small non-peptide antigens such as zoledronate can do activation and expansion of T cells in vitro. It is evident that for adoptive cancer therapies, large numbers of functional cells are needed into cancer patients. Hence, optimization of methods needs to be carried out for the efficient expansion of these T cells. Standardization of peripheral blood mononuclear cells (PBMCs) isolation was devised. Cytokines (interleukin 2 (IL-2) and interleukin 15 (IL-15)) and zoledronate were also standardized for different concentrations. It was found that an increased number of PBMCs were recovered when washing was done at 1100 revolution per minute (rpm). Significantly high expansion fold was (2524 ± 787 expansion fold) achieved when stimulation of PBMCs was done with 1 µM of zoledronate and both cytokines IL-2 and IL-15 supported the expansion and survival of cells at the concentrations of 100 IU/ml and 10 ng/ml respectively. 14-day cultures showed highly pure (91.6 ± 5.1%) and live (96.5 ± 2.5%) expanded γδ T cells. This study aimed to standardize an easy to manipulate technique for the expansion of γδ T cells, giving a higher yield.     

Imaging techniques for the detection of stored product pests

Stored grains are subject to deterioration and losses through various factors, but mainly insects and fungi. Various techniques are employed to detect stored product pests; however, there is an urgent need for an industrial-scale on-line detection technique. Near-infrared hyperspectroscopic imaging and soft X-rays have shown the potential for real-time application. These techniques are particularly effective for detecting internal infestations of stored grains. The digital images of the scanned objects are analyzed for various spectral and image features using statistical techniques such as complex multivariate tools. Classification accuracies as high as 80–100 % have been achieved for various pest and grain combinations. Dual-energy X-rays have been shown to detect the concealed eggs of stored product insect pests. The main threats to stored cereals come from Aspergillus spp., Penicillium spp., and Fusarium spp., which may produce mycotoxins. These imaging techniques have shown good results in the detection of fungal infections of stored grain.     

Oil palm (Elaeis guineensis) protoplasts: isolation, culture and microcallus formation

Procedures are deseribed for the efficient isolation of protoplasts from a variety of oil palm (Elaeis guineensis Jacq.) tissues. Various factors including donor source, composition of enzyme mixture and culture medium affected the yield and viability of the protoplasts Polyembryogenic cultures of oil palm were the most suitable starting material in terms of yield, viability and metabolic competence. Pectolyase Y-23 in association with cellulase and hemicellulase was required for the efficient release of protoplasts from the oil palm tissues. Limited cell division to form microcallus was observed at very low frequency (<0.01%) when glutathione and catalase were incorporated in the culture medium.Abbreviations 2,4-d dichlorophenoxyacetic acid - DTT dithiothreitol - MES 2[N-morpholino] ethanesulphonic acid - NAA 1-naphthalene acetic acid - PVP polyvinylpyrrolidone     

Fatigue effect on cross-talk in mechanomyography signals of extensor and flexor forearm muscles during maximal voluntary isometric contractions

Objective:This paper presents the analyses of the fatigue effect on the cross-talk in mechanomyography (MMG) signals of extensor and flexor forearm muscles during pre- and post-fatigue maximum voluntary isometric contraction (MVIC).Methods:Twenty male participants performed repetitive submaximal (60% MVIC) grip muscle contractions to induce muscle fatigue and the results were analyzed during the pre- and post-fatigue MVIC. MMG signals were recorded on the extensor digitorum (ED), extensor carpi radialis longus (ECRL), flexor digitorum superficialis (FDS) and flexor carpi radialis (FCR) muscles. The cross-correlation coefficient was used to quantify the cross-talk values in forearm muscle pairs (MP1, MP2, MP3, MP4, MP5 and MP6). In addition, the MMG RMS and MMG MPF were calculated to determine force production and muscle fatigue level, respectively.Results:The fatigue effect significantly increased the cross-talk values in forearm muscle pairs except for MP2 and MP6. While the MMG RMS and MMG MPF significantly decreased (p<0.05) based on the examination of the mean differences from pre- and post-fatigue MVIC.Conclusion:The presented results can be used as a reference for further investigation of cross-talk on the fatigue assessment of extensor and flexor muscles’ mechanic.     

Unsteady Boundary Layer Flow and Heat Transfer of a Casson Fluid past an Oscillating Vertical Plate with Newtonian Heating

In this paper, the heat transfer effect on the unsteady boundary layer flow of a Casson fluid past an infinite oscillating vertical plate with Newtonian heating is investigated. The governing equations are transformed to a systems of linear partial differential equations using appropriate non-dimensional variables. The resulting equations are solved analytically by using the Laplace transform method and the expressions for velocity and temperature are obtained. They satisfy all imposed initial and boundary conditions and reduce to some well-known solutions for Newtonian fluids. Numerical results for velocity, temperature, skin friction and Nusselt number are shown in various graphs and discussed for embedded flow parameters. It is found that velocity decreases as Casson parameters increases and thermal boundary layer thickness increases with increasing Newtonian heating parameter.     

Antioxidative responses to mercury in the cell cultures of Sesbania drummondii.

The effect of mercury (Hg) on the growth and the response of antioxidative systems have been investigated in Sesbania cell cultures to determine the tolerance limits and the mechanisms of metal (Hg) tolerance in plant cells. Cell cultures of Sesbania were developed in different concentrations (0-50 microM) of mercury. Cultures tolerated Hg up to a concentration of 40 microM and showed an increase in the fresh weight growth by 620% in 3 weeks. The levels of antioxidants: glutathione (GSH) and non-protein thiols (NPSH) and the activities of antioxidative enzymes: superoxide dismutase (SOD, EC 1.15.1.1), ascorbate peroxidase (APX, EC 1.11.1.11) and glutathione reductase (GR, EC 1.6.4.2) were influenced by Hg treatments. The contents of GSH, NPSH and GSH/GSSG ratio increased up to a concentration of 40 muM Hg and then severely declined at 50 microM Hg. The activities of antioxidative enzymes, SOD, APX and GR followed the same trends as antioxidants, first increased up to a concentration of 40 muM Hg and then declined in the presence of 50 microM Hg.