A Method for Demonstrating Prophenoloxidase after Electrophoresis

The demonstration of prophenoloxidase after electrophoresis is based on its activation by sodium dodecyl sulfate (SDS) or sodium oleate and staining the activated phenoloxidase with dopamine and 3-methyl-2-benzothiazolinone hydrazone hydrochloride (MBTH). A rapid method is presented for demonstrating the presence of activated phenoloxidase using polyacrylamide gel electrophoresis followed by staining in the presence of SDS or sodium oleate.     

Isolation of cold stress-responsive genes from Lepidium latifolium by suppressive subtraction hybridization

Suppression subtraction hybridization (SSH) libraries were constructed from RNA isolated from leaves of control and cold stress-induced Lepidium latifolium, a cold-tolerant plant species from high altitudes for isolation of cold-responsive genes. A total of 500 clones were obtained from the cold stress library. Dot blot expression analysis identified 157 clones that were upregulated and 75 that were downregulated during cold stress. These clones selected on the basis of their expression patterns on dot blot were sequenced. As much as 27 and 17 genes were identified from the forward and reverse libraries, respectively. The genes identified revealed homology with genes involved in diverse processes such as gene regulation/signaling, photosynthesis, DNA damage repair protein, pathogenesis-related protein, senescence-associated proteins and proteins with unknown functions.     

The cowpea trypsin inhibitor promoter drives expression in response to cellular maturation in Arabidopsis thaliana

The bowman-birk type trypsin inhibitors accumulate in high concentration in legume and cereal seeds, especially during seed maturation and are considered to be involved in insect tolerance. The 5′ flanking sequences of the trypsin inhibitor was isolated from cowpea genomic DNA using anchor PCR. Analysis of sequences showed presence of seed specific RY elements and also other elements associated with seed development such as abscisic acid responsive elements (ABA responsive elements; ABRE) and dehydration responsive elements (DRE). Spatial and temporal control of the promoter driven expression pattern was analyzed using gus as reporter. Expression was found to occur both in embryo and endosperm; starting from torpedo stage of embryogenesis and continuing till the stage of final maturation i.e. bent cotyledon stage. Additional expression analyses showed that the promoter actually drives expression in tissues like leaves, roots, stipules, etc., but followed a specific pattern. Comparative analysis of expression in seeds and other organs indicated that the promoter driven expression is in response to cellular maturation.     

First insights into the biochemical and molecular response to cold stress in <Emphasis Type="Italic">Cicer microphyllum</Emphasis>, a crop wild relative of chickpea (<Emphasis Type="Italic">Cicer arietinum</Emphasis>)

Identifying a potential crop wild relative (CWR) of legumes, especially one with high abiotic stress tolerance, has been a priority of plant breeders for many decades. Traditionally CWRs have been selected based on biometrical traits observed in the field, however this methodology is insufficient for research into nonmorphological traits such as stress tolerance. Biochemical and molecular analysis of potential CWRs allows for more informed selection. Specifically, we focus on Cicer microphyllum Benth, a CWR of cultivated chickpea Cicer arietinum L., which is distributed in Trans Himalayan ranges adjacent to glaciers of India and Pakistan at the alpine altitude gradient between 2700 to 6000 m. The objective of this study is to begin characterization of the biochemical and molecular bases of adaptation of C. microphyllum to cold stress and compare it to its cultivated relative (Cold susceptible genotype ILC533). Significant differences were recorded in terms of malondialdehyde (MDA) concentration, electrolyte leakage and proline accumulation in C. microphyllum, as compared to C. arietinum, upon cold exposure (4°C/24h). C. microphyllum exhibits more membrane stability under cold stress. Furthermore, proline overaccumulation and an increase in the enzymatic activities of antioxidants including superoxide dismutase, catalase, and ascorbate peroxidase were also observed in C. microphyllum under cold stress treatment. Expression of pyrroline-5-carboxylate synthetase, chalcone reductase, flavonoid 3',5'-hydroxylase and flavonoid 3'-monooxygenase are all upregulated under cold treatment in C. microphyllum. The characteristics recommend C. microphyllum both as a model for plant response to cold stress and as a potential source for abiotic stress resistant germplasm for chickpea breeding programs.     

Pure red upconverted and near-infrared luminescence properties of Er3+-doped SnO2 nanocrystals for lighting applications

Tin oxide (SnO₂) nanocrystalline powders doped with erbium ion (Er³⁺) in different molar ratios (0, 3, 5, and 7 mol%) were prepared using a solid-state reaction technique. These samples were characterized by X-ray diffraction (XRD), scanning electron microscopy (SEM), ultraviolet-visible absorption, visible upconversion, and near-infrared luminescence techniques. XRD analysis revealed the tetragonal rutile structure of SnO₂ and the average crystallite size was about 32 nm. From Tauc's plots, it was confirmed that the substitution of Er³⁺ ions into the SnO₂ host lattice resulted in the narrowing its band gap. Optical absorption bands at 520 and 654 nm correspond to the 4f electron transitions of Er³⁺ further confirming visible light absorption. Infrared luminescence spectra showed a broad band centred at 1536 nm which is assigned to the ⁴I_13/2 → ⁴I_15/2 transition of Er³⁺. Visible upconverted emission spectra under 980 nm excitation exhibit a strong red luminescence with a main peak at 672 nm which is attributed to the ⁴F_9/2 → ⁴I_15/2 transition of Er³⁺. Power-dependent upconversion spectra confirmed that two photons participated in the upconversion mechanism. Enhancement in the intensities of both visible and infrared luminescence was observed when raising the concentration. The results pave the way for the potential applications of these nanocrystalline powders in energy harvesting applications such as infrared light upconverting layer in solar cells, light emitting diodes, infrared broadband sources and amplifiers, and biological labelling.     

Beneficial effects of alpha linolenic acid rich flaxseed oil on growth performance and hepatic cholesterol metabolism in high fat diet fed rats

The purpose of the study was to investigate the effect of flaxseed oil (FO), rich in alpha-linolenic acid (ALA) (18:3 n-3) on growth parameters and lipid metabolism of rats fed with high fat diet. High fat diet (HFD) resulted in significant alterations in hepatic lipids, increase in body weight gain and negative effect on lipoprotein metabolism. FO supplementation significantly lowered the increase in body weight gain, liver weight, plasma cholesterol, triglycerides, phospholipids, free fatty acids, high-density lipoprotein (HDL), low-density lipoprotein-cholesterol (LDL-C), very low-density lipoprotein (VLDL), LDL/HDL and TC/HDL ratio in HFD fed rats. FO significantly reduced the hepatic and plasma lipid levels indicating its hypolipidemic activity. On the other hand, oral administration of FO exhibited lower plasma lipoprotein profile as compared to HFD rats. Hepatic protection by FO is further substantiated by the normal liver histological findings in HFD fed rats. These data suggest that FO participate in the normal regulation of plasma lipid concentration and cholesterol metabolism in liver. No adverse effect of FO on growth parameters and plasma lipids in rats fed with fat-free diet. The results of the present study demonstrate that FO may be developed as a useful therapy for hyperlipidemia through reducing hepatic lipids, thereby proving its hypolipidemic activity.     

A simple flow cytometry-based assay to study global methylation levels in onion,a non-model species

DNA methylation is an important epigenetic mark and global methylation dynamics regulate plant developmental processes. Even though genome sequencing technologies have made DNA methylation studies easier, it is difficult in non-model species where genome information is not available. Therefore in this study, we developed a simple assay for analysing global methylation levels in plants by washless immunolabelling of unfixed nuclei using flow cytometry. Onion leaf tissue was used as a model system, and mean fluorescence intensity due to anti-5- methyl cytosine (5-mC) antibodies were used as a measure of global methylation levels. Among three nuclear isolation buffers evaluated, the highest nuclear yield with the low background was obtained with LB01. To maintain a balance between high DNA fluorescence value and low coefficient of variation of DNA peaks, 45 min of hydrolysis with 0.2 N hydrochloric acid was used for chromatin denaturation resulting in six-fold increase in 5-mC fluorescence compared to control. This method was used successfully to detect 5-Azacytidine induced DNA hypomethylation in onion leaf tissues.Supplementary InformationThe online version contains supplementary material available at 10.1007/s12298-021-01047-6.