Reagents for the selective immobilization of oligonucleotides on solid supports

New reagents (CPGs and phosphoramidites) for automatic solid phase synthesis of modified oligonucleotides were designed. Three oligonucleotides carrying fluorescent label at the 5'-terminus and an anchor group at the 3'-terminus were prepared and their immobilization in orthogonal conditions on solid supports was studied.     

Design of general bidirectional associative memories with improved recall capability

The concept of qth order Hamming ball for general bidirectional associative memories (GBAMs) is presented, which results in an enhanced recall capability. A technique based on the minimum overlap algorithm (MOA) is proposed to deal with the separability problem of the qth order Hamming balls. Simulation results show that the recall capability obtained by using the proposed method is indeed larger than that obtained by using the conventional method.     

Improved activity retention of enzymes deposited on solid supports

Enzymes deposited on solid support usually show good stability when operated in organic solvents. Decreased stability of the enzyme preparations was noticed when low enzyme loadings were used (e.g., with Celite as support; less than 1 mg enzyme/g). It was possible to avoid the activity loss by the addition of an additive which protects the enzyme during the immobilization. Proteins (such as albumin, gelatin, and casein) and poly(ethylene glycol) were effective additives whereas amino acids, monomeric carbohydrates, and polysaccharides had no effect. The amount of additive needed for stabilization was shown to depend on the structure of the support, more additive being required for a support with high porosity. The stabilizing effect was investigated in a series of glyceryl-controlled-pore glass (CPG) with varying specific surface areas (9.5-180 m(2)/g). The minimum addition of albumin, giving full stabilization, on the different supports correlated to a monolayer coverage of the surface, approximately 2-3 mg protein/m(2). The effect of the additive was less pronounced when increasing amounts of enzyme were immobilized (5-40 mg enzyme/g Celite). The effect of the additives was studied using mandelonitrile lyase, but alpha-chymotrypsin and lipase P were also shown to be stabilized. (c) 1993 John Wiley & Sons, Inc.     

Glucocorticoids are necessary for enhancing the acquisition of associative memories after acute stressful experience

Exposure to acute stressful experience can enhance the later ability to acquire new memories about associations between stimuli. This enhanced learning is observed during classical eyeblink conditioning of both hippocampal-dependent and -independent learning. It can be induced within minutes of the stressful event and persists for days. Here we examined the role of the major stress hormones glucocorticoids in the enhancement of learning after stress. In the first two experiments, it was determined that adrenalectomy (ADX), with and without replacement of basal levels of corticosterone, prevented the stress-induced enhancement of trace conditioning, a task that is dependent on the hippocampus for acquisition. In a third experiment, demedullation, which removes the adrenal medulla but leaves the adrenal cortex and corticosterone levels intact, did not affect the enhancement of learning after stress. In a fourth experiment, ADX prevented the stress-induced enhancement of delay conditioning, a hippocampal-independent task. In a final experiment, it was determined that one injection of stress levels of corticosterone enhanced new learning within minutes but not new learning 24 h later. Together these results suggest that endogenous glucocorticoids are necessary and sufficient for transiently enhancing acquisition of new associative memories and necessary but insufficient for persistently enhancing their acquisition after exposure to an acute stressful experience.     

Hippocampal NMDA receptors are important for behavioural inhibition but not for encoding associative spatial memories

The idea that an NMDA receptor (NMDAR)-dependent long-term potentiation-like process in the hippocampus is the neural substrate for associative spatial learning and memory has proved to be extremely popular and influential. However, we recently reported that mice lacking NMDARs in dentate gyrus and CA1 hippocampal subfields (GluN1^ΔDGCA1 mice) acquired the open field, spatial reference memory watermaze task as well as controls, a result that directly challenges this view. Here, we show that GluN1^ΔDGCA1 mice were not impaired during acquisition of a spatial discrimination watermaze task, during which mice had to choose between two visually identical beacons, based on extramaze spatial cues, when all trials started at locations equidistant between the two beacons. They were subsequently impaired on test trials starting from close to the decoy beacon, conducted post-acquisition. GluN1^ΔDGCA1 mice were also impaired during reversal of this spatial discrimination. Thus, contrary to the widely held belief, hippocampal NMDARs are not required for encoding associative, long-term spatial memories. Instead, hippocampal NMDARs, particularly in CA1, act as part of a comparator system to detect and resolve conflicts arising when two competing, behavioural response options are evoked concurrently, through activation of a behavioural inhibition system. These results have important implications for current theories of hippocampal function.     

Optimization of protein-based volumetric optical memories and associative processors by using directed evolution

The potential use of proteins in device applications has advanced in large part due to significant advances in the methods and procedures of protein engineering, most notably, directed evolution. Directed evolution has been used to tailor a broad range of enzymatic proteins for pharmaceutical and industrial applications. Thermal stability, chemical stability, and substrate specificity are among the most common phenotypes targeted for optimization. However, in vivo screening systems for photoactive proteins have been slow in development. A high-throughput screening system for the photokinetic optimization of photoactive proteins would promote the development of protein-based field-effect transistors, artificial retinas, spatial light modulators, photovoltaic fuel cells, three-dimensional volumetric memories, and optical holographic processors. This investigation seeks to optimize the photoactive protein bacteriorhodopsin (BR) for volumetric optical and holographic memories. Semi-random mutagenesis and in vitro screening were used to create and analyze nearly 800 mutants spanning the entire length of the bacterio-opsin (bop) gene. To fully realize the potential of BR in optoelectronic environments, future investigations will utilize global mutagenesis and in vivo screening systems. The architecture for a potential in vivo screening system is explored in this study. We demonstrate the ability to measure the formation and decay of the red-shifted O-state within in vivo colonies of Halobacterium salinarum, and discuss the implications of this screening method to directed evolution. These authors contributed equally to this work.     

Increased resting-state perfusion after repeated encoding is related to later retrieval of declarative associative memories

Electrophysiological studies in animals have shown coordinated reactivation of neuronal ensembles during a restricted time period of behavioral inactivity that immediately followed active encoding. In the present study we directly investigated off-line processing of associative memory formation in the human brain. Subjects' regional cerebral blood flow (rCBF) as a surrogate marker of neural activity during rest was measured by MR-based perfusion imaging in a sample of 14 healthy male subjects prior to (Pre2) and after (Post) extensive learning of 24 face-name associations within a selective reminding task (SR). Results demonstrated significant Post-Pre2 rCBF increases in hippocampal and temporal lobe regions, while in a control comparison of two perfusion scans with no learning task in-between (Pre2-Pre1) no differences in rCBF emerged. Post perfusion scanning was followed by a surprise cued associative recall task from which two types of correctly retrieved names were obtained: older names already correctly retrieved at least once during one of the SR blocks, and recent names acquired during the last SR block immediately prior to the Post scan. In the anterior hippocampus individual perfusion increases were correlated with both correct retrievals of older and recent names. By contrast, older but not recently learned names showed a significant correlation with perfusion increases in the left lateral temporal cortex known to be associated with long-term memory. Recent, but not older names were correlated with dopaminergic midbrain structures reported to contribute to the persistence of memory traces for novel information. Although the direct investigation of off-line memory processing did not permit concomitant experimental control, neither intentional rehearsal, nor substantial variations in subjects' states of alertness appear to contribute to present results. We suggest that the observed rCBF increases might reflect processes that possibly contribute to the long-term persistence of memory traces.     

Correlated receptor and motorneuron changes during retention of associative learning of Hermissenda crassicornis

1. Responses to light of an identified motorneuron (LP1) were recorded simultaneously with those of an identified Hermissenda photoreceptor (the lateral Type B) following three days of training with paired light and rotation. 2. These responses were significantly different when compared to responses of cells from animals trained with unpaired stimuli and from naive animals. 3. The differences of the LP1 responses can be explained as a consequence of the photoreceptor response changes. 4. The same training with paired stimuli has been shown to produce behavioural changes which satisfy criteria for vertebrate associative learning. 5. The observed neural correlates are consistent with previous findings which indicate that membrane changes within the Type B cell bodies play a causal role in associative learning of the nudibranch mollusc, Hermissenda crassicornis.     

Influence of native disulfide bonds of globular proteins on their retention behavior on reverse phase supports

The reduction of the disulfide bonds of globular proteins, for example, those of lysozyme or ribonuclease-A, results in an increase in the hydrodynamic volume of the polypeptide chain. This is reflected in an earlier elution of the reduced protein on gel filtration compared to that of the native disulfide-bonded form. The reduction of the four disulfide bonds of ribonuclease-A increased its retention time on reverse phase support, suggesting an increase in the apparent hydrophobicity of the protein molecule on reduction. Performic acid-oxidized ribonuclease-A eluted ahead of native disulfide-bonded ribonuclease on RP HPLC, suggesting a decrease in the hydrophobicity of the molecule. However, the hydrodynamic volume of performic acid-oxidized ribonuclease-A is similar to that of reduced protein as reflected in its gel filtration behavior. Thus, the increased retention of the reduced protein compared to that of native disulfide-bonded protein is not related to the increased hydrodynamic volume, and is a reflection of the stronger interaction of reduced protein with the reverse phase support. Reoxidation of the reduced ribonuclease-A regenerated the original chromatographic behavior of the protein on the reverse phase support. Similar results were also obtained with hen egg white lysozyme. The results of the present study are interpreted as indicating that the native disulfide bonds of a globular protein restrict the exposure of the hydrophobic amino acid residues of the polypeptide chain with a consequent lower retention on the reverse phase support compared to its reduced form.     

Age and rearing environment interact in the retention of early olfactory memories in honeybees

Due to the changing behavioral contexts at which social insects are exposed during the adult lifespan, they are ideal models to analyze the effect of particular sensory stimuli during young adulthood on later behavior. Specifically, our goal is to understand early influences on later foraging behavior. For that, olfactory memories were established by worker honeybees to different pre-foraging ages using either (1) classical conditioning in the proboscis extension response (PER) paradigm or (2) the offering of scented-sugar solution under different rearing conditions. By testing long-term memories (LTM) through a single PER test in workers of foraging ages (17-25 days), we found that retention of the early olfactory memories in honey bees is age-dependent and not time-dependent. Independently of the environmental conditions in which they were reared (laboratory cages or hives), bees were able to retain food-odor association from 5 days after emergence, but rarely before. In most experiments we observed a bi-modal pattern of response: bees exposed to scented-food at 5-8 and 13-16 days showed better retention than those exposed at 9-12 days. These differences disappeared for bees reared in hives. Retrieval of LTMs depending on the timing and the continuous inputs of appropriate sensory stimuli are discussed.     

Parameter estimation of a monod-type model. Part II: Use of a neural network with associative memories

Summary On the basis of a previous study related with parameter identifiability and sensitivity analysis of a Monod-type model, a parameter estimation method based on Artificial Neural Networks (ANN) with Associative Memories (AMs) is presented. A combination of an iterative procedure and a convergence index given by AMs allows to confirm the nature of relations existing between state variables and parameters which were found in the first part of the study. The convergence criterion is particularly well adapted to showing various influences of state variables on parameter estimation of such a model.     

Molecular optical memories and switches based on photochromic dihydropyridines.

Photochromic behavior of dihydropyridines and the mechanism of the photochromism based on phenyl transfer is described. Formation of polar coloured species, changes in optical absorption and in the tau electron character of the molecular skeleton allow for the fabrication of molecular optical memories and switches.     

Inducible and selective erasure of memories in the mouse brain via chemical-genetic manipulation

Rapid and selective erasures of certain types of memories in the brain would be desirable under certain clinical circumstances. By employing an inducible and reversible chemical-genetic technique, we find that transient alphaCaMKII overexpression at the time of recall impairs the retrieval of both newly formed one-hour object recognition memory and fear memories, as well as 1-month-old fear memories. Systematic analyses suggest that excessive alphaCaMKII activity-induced recall deficits are not caused by disrupting the retrieval access to the stored information but are, rather, due to the active erasure of the stored memories. Further experiments show that the recall-induced erasure of fear memories is highly restricted to the memory being retrieved while leaving other memories intact. Therefore, our study reveals a molecular genetic paradigm through which a given memory, such as new or old fear memory, can be rapidly and specifically erased in a controlled and inducible manner in the brain.     

The future of anaerobic digestion and biogas utilization

One of the common tendencies of animal production activities in Europe and in developed countries in general is to intensify the animal production and to increase the size of the animal production units. High livestock density is always accompanied by production of a surplus of animal manure, representing a considerable pollution threat for the environment in these areas. Avoiding over-fertilization is not only important for environmental protection reasons but also for economical reasons. Intensive animal production areas need therefore suitable manure management, aiming to export and to redistribute the excess of nutrients from manure and to optimize their recycling.     

Behavioral ecology of odometric memories in desert ants: acquisition, retention, and integration

Assuming that the acquisition and retention of memories havecosts, properties of memories should fit the functional requirementsfor the system of memory. Based on a functional analysis ofwhat path integration is meant to do, we predicted that odometricmemories in desert ants should show (1) little improvement withrepeated training: performance should be as good after one trainingtrial as after six training trials, (2) decay of memory after24 h, and (3) performance based solely on the most recent outboundtrip, with no integration over multiple memories. Desert ants(Cataglyphis fortis) traveled in narrow straight plastic channelsto forage for cookie crumbs in a feeder at 6- or 12-m distance.Each ant was tested once by being taken from the feeder andreleased 2 m from the end of a 32-m channel to run home. Thedistance at which the ant first turned back (first turn) constitutedthe data. In acquisition, groups trained one or six times beforebeing tested had unsystematic scatter that did not differ significantly.In retention, ants tested after a 24-h delay showed larger unsystematicscatter than control animals tested after no delay. In integration,ants were trained five times at 6 or 12 m and then tested at12 or 6 m, respectively. No evidence of integration of multipleodometric memories was found. The results show that the propertiesof odometric memories are indeed tailored to what the memorysystem is used for.     

Review: Current status and future prospects of associative nitrogen fixation in rice

Biological sources of nutrients are gaining importance over the chemical and organic sources from the standpoint of environmental safety and quality, and sustainable agriculture. The nutrient input for a growing rice crop can largely be met by promoting the activities of physiologically diverse microorganisms in the aerobic, anaerobic and interface zones in the ecologically important flooded soils. Associative bacteria contribute from 10 to 80 kg N per hectare per cropping season depending upon the ecosystem, cultural practices and rice variety grown. In addition to N contribution, these bacterial associations can improve the nutrient transformations and contribute to plant growth-promoting effects. Current improved agronomic and crop production management systems greatly affect the contributions of biological sources to the overall soil nutrient status. Azospirillum and other associative bacterial systems have been intensively researched using various evaluation techniques to understand the diazotrophic rhizocoenosis. Researches clearly indicate that these associations are governed by several soil, water, nutrient, agrochemical, plant genotype and other biological factors. Considerable efforts have been made so far in selecting efficient bacterial strains as inoculants and identifying host genotypes which support maximum nitrogenase activity in addition to other beneficial traits of effective associative relationships. Knowledge gained so far on how the N2-fixing system in rice functions suggests the need for providing optimum management practices to ensure greater contribution from the plant-microbe associations. Holistic approaches integrating technological developments and achievements in biological sciences could lead to crop improvement. Research on extending nitrogen-fixing symbiosis to rice using molecular and genetic approaches is underway, albeit at a slow pace. The need for further fine-tuning and developing management practices, innovative approaches to improve rice-bacterial systems and the strategies to sustain the benefits from associative diazotrophy are discussed.     

Phosphoramidite reagents and solid-phase supports based on hydroxyprolinol for the synthesis of modified oligonucleotides

The synthesis of phosphoramidite reagents and solid-phase supports based on hydroxyprolinol for the introduction of the residues of biotin, lipoic acid, amino groups, and terminal acetylene groups at different positions of the oligonucleotide chain has been described. The efficiency of the reagents and supports has been confirmed by the synthesis of the corresponding modified oligonucleotides.     

Inclined sedimentation for selective retention of viable hybridomas in a continuous suspension bioreactor.

The continuous separation of nonviable hybridoma cells from viable hybridoma cells by using a narrow rectangular channel that is inclined from the vertical has been investigated experimentally. The effectiveness of the settler in selectively retaining viable hybridomas in the bioreactor while permitting the removal of nonviable hybridomas has been shown to depend on the flow rate through the settler. Intermediate flow rates through the settler have been found to provide the highest removal of nonviable hybridomas relative to viable hybridoma retention. At high dilution rates through the chemostat, over 95% of the viable cells could be partitioned to the bottom of the settler while over 50% of the nonviable cells are removed through the top of the settler. This successful separation is due to the significantly larger size of the viable hybridomas than the nonviable ones. A continuous perfusion experiment was performed in which an external inclined settler was used to retain virtually all of the viable hybridomas in the culture, while selectively removing from the culture approximately 20% of the nonviable cells that entered the settler. A stable viable cell concentration of 1.0 x 10(7) cells/mL was achieved, as was an antibody productivity of over 50 micrograms/(mL.day). These represent 3- and 6-fold increases, respectively, over the values obtained from a chemostat culture without cell retention.     

A new MapReduce associative classifier based on a new storage format for large-scale imbalanced data

The process of knowledge discovery from big and high dimensional datasets has become a popular research topic. The classification problem is a key task in bioinformatics, business intelligence, decision science, astronomy, physics, etc. Building associative classifiers has been a notable research interest in recent years because of their superior accuracy. In associative classifiers, using under-sampling or over-sampling methods for imbalanced big datasets reduces accuracy or increases running time, respectively. Hence, there is a significant need to create efficient associative classifiers for imbalanced big data problems. These classifiers should be able to handle challenges such as memory usage, running time and efficiently exploring the search space. To this end, efficient calculation of measures is a primary objective for associative classifiers. In this paper, we propose a new efficient associative classifier for big imbalanced datasets. The proposed method is based on Rare-PEARs (a multi-objective evolutionary algorithm that efficiently discovers rare and reliable association rules) and is able to evaluate rules in a distributed manner by using a new storing data format. This format simplifies measures calculation and is fully compatible with the MapReduce programming model. We have applied the proposed method (RPII) on a well-known big dataset (ECBDL’14) and have compared our results with seven other learning methods. The experimental results show that RPII outperform other methods in sensitivity and final score measures (the values of sensitivity and final score measures were approximately 0.74 and 0.54 respectively). The results demonstrate that the proposed method is a good candidate for large-scale classification problems; furthermore, it achieves reasonable execution time when the target platform is a typical computer clusters.     

基于蓝牙4.0的便携式睡眠监测仪的研制

目的:实时监测睡眠状况,从而帮助人们特别是老人找到影响睡眠的原因。方法:设计了一个低功耗便携式睡眠监测仪,它是通过加速度传感器采集腕动信号、蓝牙4.0低功耗无线传输、Micro-SD卡存储、上位机显示等实现对睡眠状态的检测。为了验证睡眠监测仪的准确性,本文采用了视频分析方法,并且对不同人群进行监测。结果:研制的睡眠监测仪具有便携低功耗等特点,能够准确监测睡眠状态。结论:睡眠监测仪的研制对使用者特别是老人帮助很大,能够帮助使用者方便适时了解自己的睡眠状况,找到影响睡眠原因和改善睡眠质量方法。