Light-up fluorophore--DNA aptamer pair for label-free turn-on aptamer sensors.

We developed a light-up fluorophore-DNA aptamer pair for label-free aptamer sensors that fluoresce upon binding to the analyte. A 42mer DNA aptamer binding to the environment-sensitive fluorophore, dapoxyl, which increased the fluorescence by more than 700-fold upon binding, was successfully used to construct aptamer sensors by fusion with analyte-binding DNA aptamers.

Advancing the development of glycated protein biosensing technology: next-generation sensing molecules.

Research advances in biochemical molecules have led to the development of convenient and reproducible biosensing molecules for glycated proteins, such as those based on the enzymes fructosyl amino acid oxidase (FAOX) or fructosyl peptide oxidase (FPOX). Recently, more attractive biosensing molecules with potential applications in next-generation biosensing of glycated proteins have been aggressively reported. We review 2 such molecules, fructosamine 6-kinase (FN6K) and fructosyl amino acid-binding protein, as well as their recent applications in the development of glycated protein biosensing systems.

Low-Vacuum Deposition of Glutamic Acid and Pyroglutamic Acid: A Facile Methodology for Depositing Organic Materials beyond Amino Acids.

Thin layers of pyroglutamic acid (Pygl) have been deposited by thermal evaporation of the molten L-glutamic acid (L-Glu) through intramolecular lactamization. This deposition was carried out with the versatile handmade low-vacuum coater, which was simply composed of a soldering iron placed in a vacuum degassing resin chamber evacuated by an oil-free diaphragm pump. Molecular structural analyses have revealed that thin solid film evaporated from the molten L-Glu is mainly composed of L-Pygl due to intramolecular lactamization.

Growth and physiological adaptation of whole plants and cultured cells from a halophyte turf grass under salt stress.

Understanding the mechanisms used by halophytic members of the Poaceae to cope with salt stress will contribute to the knowledge necessary to genetically engineer salt-tolerant crops. In this study, we identified a genotype of Sporobolus virginicus, a halophytic turf grass collected in Japan, and investigated its growth rate, ion concentration and secretion, and proline concentration in comparison with the reported properties of genotypes collected from the USA, South Africa and Egypt. Surprisingly, the Japanese genotype showed a salinity tolerance up to 1.

Detection of single methylated cytosine using junction-forming DNA probes.

DNA methylation is an epigenetic mechanism for transcriptional regulation. The methylation process controls cellular differentiation and is defective in many diseases including cancer. Therefore, the development of a simple method for analysing cytosine methylation in a target gene is required.

Sequence-selective modification of DNA cytosine by using junction-forming DNA probes and its application to the detection of single cytosine methylation.

Methylated DNA plays an important role in epigenetic gene regulation, and could therefore serve as a potentially promising biomarker for the diagnosis of cancer and other diseases. Therefore, the development of a simple method for analyzing cytosine methylation in a target gene is required. Here, we report on a new method for the sequence-selective chemical modification of a single cytosine in single-stranded DNA (ssDNA).

Involvement of elevated proline accumulation in enhanced osmotic stress tolerance in Arabidopsis conferred by chimeric repressor gene silencing technology.

Arabidopsis plants transformed with a chimeric repressor for 6 transcription factors (TFs), including ADA2b, Msantd, DDF1, DREB26, AtGeBP, and ATHB23, that were converted by Chimeric REpressor gene Silencing Technology (CRES-T), show elevated salt and osmotic stress tolerance compared with wild type (WT) plants. However, the roles of TFs in salt and osmotic signaling remain largely unknown. Their hyper-osmotic stress tolerance was evaluated using 3 criteria: germination rate, root length, and rate of seedlings with visible cotyledons at the germination stage.

Long-term electrophysiological activity and pharmacological response of a human induced pluripotent stem cell-derived neuron and astrocyte co-culture.

Human induced pluripotent stem cell (hiPSC)-derived neurons may be effectively used for drug discovery and cell-based therapy. However, the immaturity of cultured human iPSC-derived neurons and the lack of established functional evaluation methods are problematic. We here used a multi-electrode array (MEA) system to investigate the effects of the co-culture of rat astrocytes with hiPSC-derived neurons on the long-term culture, spontaneous firing activity, and drug responsiveness effects.

Thymoquinone protects cultured hippocampal and human induced pluripotent stem cells-derived neurons against α-synuclein-induced synapse damage.

The seeds of Nigella sativa are used worldwide to treat various diseases and ailments. Thymoquinone (TQ) that is present in the essential oil of these seeds mediates most of the plant's diverse therapeutic effects. The present study aimed to determine whether TQ protects against α-synuclein (αSN)-induced synaptic toxicity in rat hippocampal and human induced pluripotent stem cell (hiPSC)-derived neurons.

Carbon nanotube multi-electrode array chips for noninvasive real-time measurement of dopamine, action potentials, and postsynaptic potentials.

Multi-electrode arrays (MEAs) can be used for noninvasive, real-time, and long-term recording of electrophysiological activity and changes in the extracellular chemical microenvironment. Neural network organization, neuronal excitability, synaptic and phenotypic plasticity, and drug responses may be monitored by MEAs, but it is still difficult to measure presynaptic activity, such as neurotransmitter release, from the presynaptic bouton. In this study, we describe the development of planar carbon nanotube (CNT)-MEA chips that can measure both the release of the neurotransmitter dopamine as well as electrophysiological responses such as field postsynaptic potentials (fPSPs) and action potentials (APs).

Control of neural network patterning using collagen gel photothermal etching.

Two-dimensional (2D) micropatterning techniques have been developed to guide dissociated neurons into predefined distributions on solid substrates, such as glass and plastic. Micropatterning methods using three-dimensional (3D) substrates or scaffolds that reproduce aspects of the in vivo microenvironment could facilitate the engineering of functional tissues for transplantation or more robust experimental models. We developed a 3D collagen gel photothermal etching method using an infrared laser that precisely controls the area of cell adhesion and neurite projection by etching a small targeted section of the collage gel.

Cloning and characterization of fructosamine-6-kinase from Arthrobacter aurescens.

Fructosamine-6-kinases (FN6Ks) that catalyze phosphorylation of glycated amino acids, i.e., fructosyl amino acids (FAs), have been shown as a potential recognition element for glycated protein detection.

Thymoquinone protects cultured rat primary neurons against amyloid β-induced neurotoxicity.

Thymoquinone (TQ) is the main constituent of the oil extracted from Nigella sativa seeds, which is known to be the active constituent responsible for many of the seed antioxidant and anti-inflammatory effects. The present study was designed to investigate whether TQ can protect against Alzheimer's amyloid-β peptide (Aβ) induced neurotoxicity in rat primary neurons. Cultured hippocampal and cortical neurons were treated with Aβ1-42 and TQ simultaneously for 72 h.

Fiber-Optic Fluoroimmunoassay System with a Flow-Through Cell for Rapid On-Site Determination of Escherichia coli O157:H7 by Monitoring Fluorescence Dynamics.

Dynamic fluoroimmunoassay with a flow-through system using optical fiber probes consisting of polystyrene was developed and applied to a quantitative detection of E. coli O157:H7. The system measures E.

A simplified micropatterning method for straight-line neurite extension of cultured hippocampal neurons.

We report a simplified micropatterning method for the straight-line extension of the neurites of cultured neurons. We prepared a poly-D-lysine (PDL)-patterned surface using a polydimethylsiloxane microfluidic stamp. Hippocampal neurons were cultured on the PDL-bound substrate with the stamp removed, allowing for conventional cell seeding and detailed optical observation without fluorescent label.

Identification and functional analysis of fructosyl amino acid-binding protein from Gram-positive bacterium Arthrobacter sp.

Aim: Fructosyl amino acid-binding protein (FABP) is a substrate-binding protein (SBP), which recognizes fructosyl amino acids (FAs) as its ligands. Although FABP has been shown as a molecular recognition tool of biosensing for glycated proteins, the availability of FABP is still limited and no FABP was reported from Gram-positive bacteria. In this study, a novel FABP from Gram-positive bacteria, Arthrobacter spp.

Identification of Chimeric Repressors that Confer Salt and Osmotic Stress Tolerance in Arabidopsis.

We produced transgenic Arabidopsis plants that express chimeric genes for transcription factors converted to dominant repressors, using Chimeric REpressor gene-Silencing Technology (CRES-T), and evaluated the salt tolerance of each line. The seeds of the CRES-T lines for ADA2b, Msantd, DDF1, DREB26, AtGeBP, and ATHB23 exhibited higher germination rates than Wild type (WT) and developed rosette plants under up to 200 mM NaCl or 400 mM mannitol. WT plants did not grow under these conditions.

Development of lipid A-imprinted polymer hydrogels that selectively recognize lipopolysaccharides.

To remove lipopolysaccharide (LPS) from pure water, we developed polymer hydrogels that selectively recognize LPS. A molecular imprinting technique was used to prepare the polymer hydrogels. We prepared the polymer hydrogels with LPS-binding sites by using acryloyllysine and acryloylphenylalanine as functional monomers and used lipid A as a template because it is the biologically active part of LPS and contains two phosphate groups.

Hexavalent chromium reduction by an actinobacterium Flexivirga alba ST13T in the family Dermacoccaceae.

An isolated strain Flexivirga alba ST13(T) was examined for the reducing activity of hexavalent chromium (Cr(VI)). Strain ST13(T) grew in the liquid medium containing 0.33 mM Cr(VI) under aerobic cultivation, even though growth was inhibited on agar minimal media containing 0.

Fluorescence emission and polarization analyses for evaluating binding of ruthenium metalloglycoclusters to lectins and tetanus toxin C-fragment.

We develop a fluorescent ruthenium metalloglycocluster for use as a powerful molecular probe in evaluating the binding between carbohydrates and lectins by fluorescence emission (FE) and fluorescence polarization (FP) analyses. Changes in the FE and FP of these metalloglycoclusters are measured following the addition of lectin [peanut agglutinin (PNA), Ricinus communis agglutinin 120, Concanavalin A (ConA), or wheat germ agglutinin] or tetanus toxin c-fragment (TCF). After the addition of PNA, the FE spectrum of [Ru(bpy-2Gal)(3)] shows a new emission peak and the FP value of [Ru(bpy-2Gal)(3)] increases.

A shRNA library constructed through the generation of loop-stem-loop DNA.

Background: Short hairpin RNA (shRNA) libraries are considered to comprise a powerful tool in genetic screening. Several library construction methods have been proposed; these methods require between four and nine tedious reaction steps to construct the library. In the present study, we describe a method of generating shRNA-coding DNA efficiently from randomized oligonucleotides based on four reaction steps.

Fluorescence emission and polarization for analyzing binding of ruthenium metalloglycocluster to lectin and tetanus toxin C-fragment.

We have designed and synthesized ruthenium complexes bearing clustered galactose Ru(bpy-2Gal)(3) and glucose Ru(bpy-2Glc)(3). Changes in fluorescence emission (FE) and fluorescence polarization (FP) of the metalloglycoclusters were measured by adding each lectin (peanut agglutinin (PNA), Ricinus communis agglutinin 120 (RCA), concanavalin A (ConA), or wheat germ agglutinin (WGA)) or tetanus toxin c-fragment (TCF). Following the addition of PNA and ConA, the FE spectra of Ru(bpy-2Gal)(3) and Ru(bpy-2Glc)(3) showed new emission peaks, respectively.

New concept for a toxicity assay based on multiple indexes from the wave shape of damped metabolic oscillation induced in living yeast cells (part I): characterization of the phenomenon.

The damped glycolytic oscillation phenomenon occurring in starved cells of the yeast Saccharomyces cerevisiae (NBRC 0565) was characterization for application to a toxicity bioassay. S. cerevisiae was grown under semi-anaerobic conditions.

New concept for a toxicity assay based on multiple indexes from the wave shape of damped metabolic oscillation induced in living yeast cells (part II): application to analytical toxicology.

An ideal toxicity assay should utilize multiple indexes obtained from transient changes of metabolic activities. Here, we demonstrate the possibility for a novel toxicity bioassay using the damped glycolytic oscillation phenomenon occurring in starved yeast cells. In a previous study, the phenomenon was characterized in detail.