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Dual language exposure and bilingualism are relatively common experiences for children. The present review set out to synthesize the existing research on cognitive development in bilingual children and to identify the gaps and the methodological concerns present in the existing research. A search of major databases for research conducted with typically developing, preschool-age dual language learners between 2000 and 2013 yielded 102 peer-reviewed articles. The existing evidence points to areas of cognitive development in bilingual children where findings are robust or inconclusive, and reveals variables that influence performance. The present review also identifies areas for future research and methodological limitations. 相似文献
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This study examines the uses and gratifications (U&G) of accessing political candidate profiles on social network Web sites. An online survey of visitors to the MySpace profiles of 2008 primary candidates revealed that voters are drawn to this source of political information mainly by the desire for social interaction with other like-minded supporters, followed by information-seeking, and entertainment. While information seeking and entertainment are common U&G of consuming online political content, they were weaker factors compared to the social interaction factor that seems to distinguish MySpace, possibly SNSs in general, from other online sources of political content. 相似文献
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In a rapidly changing world, the mission of education deserves some reflection. Mutual understanding and assessment between trainers and trainees offers a way to promote discussion concerning goals, values, and strategies that should be promoted at schools. This study offers the views of 153 pre-service teachers and their respective trainers during their practicum. We aimed to determine if an association exists between the scores of pre-service teachers and teachers regarding behaviors and attitudes shown by the first. We also want to analyze the extent to which pre-service teachers rate the importance of different educational strategies as well as the extent to which teachers use these strategies in their daily work. We also aim to determine to what extent self-rated behaviors and attitudes of pre-service teachers are associated to their ratings on importance and utilization of different educational strategies. Two questionnaires were utilized to gather the data. Results revealed higher scores on self-evaluation than others’ evaluations; utilization of diverse educational strategies was associated to evaluations on pre-service students’ responsibility, ability to detect and meet students’ needs, and final grade in practicum. Association between pre-service teachers’ self-evaluation and evaluation on the importance of different educational strategies revealed large associations between climate for the expression of ideas, teaching methodology, and the importance given to using language appropriate to the level of the students. Average ratings on importance and utilization of different teaching strategies resulted in high scores, with utilization of teaching methodologies obtaining the lowest scores. Gender resulted in significant differences on importance, with women scoring higher than men. Importance scores were significantly higher than utilization scores. High associations were found between self- and others’ evaluations on values related to compliance with rules, as well as on behaviors associated to maintain order and discipline in the classroom. Differences in views of teaching, importance, and utilization of different teaching strategies should be debated in order to advance our understanding of effectiveness of educational practices. 相似文献
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To collect data on ozone pollution and foliar damage, we established the first Philadelphia Ozone Bioindicator Garden at The Franklin Institute Science Museum. In this paper, we examine the relationship between ozone concentrations in an urban environment and changes in foliar damage in cutleaf coneflower (Rudbeckia laciniata) plants. Higher ozone concentrations were observed during Summer 2015 than during Summer 2014 at our site. Diurnal analysis reveals a nightly diminishment of ozone's rate of dissipation around 4am, which we attributed to intrusions of air from the residual layer (the layer of air above the boundary layer). We saw that visible foliar injury starts relatively slowly and accelerates beginning in late July for stippling and early August for chlorosis and necrosis. We found that injury ratings on ozone-damaged leaves progress faster later in the season despite lower ambient ozone concentrations; we hypothesized that this is evidence of a “latency period” in the cutleaf coneflower's foliar injury response to ozone. Cutleaf coneflowers may serve as engaging tools to alert and inform Philadelphians about air quality issues. Our results suggest that these plants are good candidates for future work in developing bioindicators of ozone where direct monitoring is not feasible. 相似文献
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Raluca Ostafe Radivoje Prodanovic W. Lloyd Ung David A. Weitz Rainer Fischer 《Biomicrofluidics》2014,8(4)
A new ultra-high-throughput screening assay for the detection of cellulase activity was developed based on microfluidic sorting. Cellulase activity is detected using a series of coupled enzymes leading to the formation of a fluorescent product that can be detected on a chip. Using this method, we have achieved up to 300-fold enrichments of the active population of cells and greater than 90% purity after just one sorting round. In addition, we proved that we can sort the cellulase-expressing cells from mixtures containing less than 1% active cells.Cellulases are important enzymes with numerous applications across multiple industries, including biofuel, pulp, paper, textile and laundry, food, feed, brewing, and agriculture.1 Most cellulases have low activity and stability, so improving these properties would have substantial impact on numerous industrial processes.Enzymatic properties can be improved by protein engineering2 but the limiting step is the screening process. Classical screening uses microtiter plates (MTPs), where each well contains cells expressing a single type of mutant enzyme. However, this type of screening is the bottleneck in directed evolution, because a maximum number of 105 clones can be screened over the course of weeks or even months3 and large quantities of reagents and consumables are needed. High-throughput screening methods based on either fluorescence activated cell sorting (FACS)4–7 or microfluidic devices8 increase the number of clones that can be screened and reduce the amount of consumables required. Here, we demonstrate the use of a high-throughput screening system for cellulases by combining lab-on-chip sorting devices with an emulsion-based fluorescent assay previously developed for use in flow cytometry.5Water–in-oil emulsions are needed to maintain the connection between genotype and phenotype by compartmentalizing individual cells expressing a mutant enzyme together with the components of the fluorescence assay corresponding to the enzyme activity.7 For FACS, double emulsions (water-in-oil-in-water) are required because the instrument''s mobile phase is an aqueous solution. Such double emulsions can be produced by stirring or agitation,9,10 but the resulting emulsions are polydisperse and multiple water droplets may be scattered within a single oil droplet. In addition, large droplets tend to produce more fluorescence because there are more substrate molecules available for conversion into the fluorescent product. The emulsions are produced in bulk, so each droplet will be detected at a different time point from the start of the reaction. This means that increased fluorescence may result because an enzyme has worked on the substrate for a longer amount of time, and the fluorescence of the droplet may plateau before sorting as the enzyme consumes all the available substrate. Cell loading is difficult to control because the average number of cells per droplet scales with droplet volume. Also, if several inner droplets, containing cells with different activities, are encapsulated within the same outer droplet, false positives may occur upon sorting. Consequently, it is impossible to differentiate fluorescence changes due to enzyme activity from those due to other effects using polydisperse double emulsions in FACS, but it is possible to achieve plus/minus screening,4 separating cells with activity from those without.Droplet microfluidics overcomes many of the drawbacks of high-throughput enzyme sorting with FACS. Both the size and composition of the droplets can be tuned precisely. Furthermore, once the enzyme is mixed with the substrate, the incubation time can be controlled and all compartments will have the same conditions in terms of concentration and total number of substrate molecules. Although cell loading is still subject to Poisson statistics, the probability for cells to be loaded into a given droplet is the same and can be adjusted by tuning the input cell density. These characteristics make the microfluidic method more sensitive, flexible, and quantitative at detecting changes in enzyme activity than the FACS-based sorting of double emulsions.Here, we report a method in which droplet microfluidics is used to sort libraries containing different percentages of cells expressing cellulase activity and demonstrate enrichment of the cells expressing active cellulases. The entire process is summarized in Figure Figure11.Open in a separate windowFIG. 1.General overview of cellulase screening using droplet microfluidics. In the emulsification device, suspensions of yeast surface displayed libraries are co-flowed with the substrate solution at equal flow rates to a drop-forming junction where they mix. A stream of perfluorinated oil then breaks the aqueous mixture into monodisperse water-in-oil emulsions. Within each droplet, the cellulase reaction starts after compartmentalization and the fluorescent product is formed by a coupled enzymatic cascade in droplets containing cells that express the active enzyme. After a fixed incubation time, the emulsion droplets are re-injected into a microfluidic sorting device, where they are analyzed and sorted based on their fluorescence.To detect cellulase activity, we designed an assay that uses a chain of coupled enzymatic reactions to yield fluorescence corresponding to cellulase activity without needing artificial substrates (which may lead to confounding effects, such as improved binding of the enzyme specifically to the artificial compound but not the natural substrate). In this method, cellulase hydrolyzes cellulose, its natural substrate, into monosaccharides and oligosaccharides that are further detected by the enzymatic cascade5 (Figure (Figure11).Based on previous FACS experiments, no difference in activity can be detected between the positive and the negative droplets before 2 h incubation time.5 Based on these observations, we expected the cells to require more than 2 h of incubation in droplets for the reaction to develop.Emulsions were formed using a co-flow flow-focusing Polydimethylsiloxane device prepared by soft lithography as previously described8 and using fluorocarbon oil containing 1% (v/v) Krytox-PEG-Krytox detergent synthesized as reported in an earlier study.11,14 The solutions, one containing library cells (S. cerevisiae YPH500 cells, Agilent Technologies, Santa Clara, USA) and the other with the substrate,14 were mixed at the same flow rate, giving a one-to-one mixing ratio. The library cells were a defined mixture of cells transformed with cel5A pESC-Trp (positive cells) or empty pESC-Trp (negative cells). The two solutions therefore mixed just prior to encapsulation, minimizing the chance that fluorescent products would enter neighboring droplets. The substrate solution contained carboxymethyl cellulose (CMC), which has a high viscosity. To prevent fluctuations in the flow of substrate during the emulsification process, we optimized the flow rate and the concentration of CMC and found that a CMC concentration of 0.33% (w/v) produced monodisperse emulsions.We discovered that the HOx required for the enzymatic cascade causes droplet coalescence. HOx alone was sufficient to cause the observed change in droplet stability because droplets containing only hexose oxidase in buffer exhibited the same amount of coalescence as those containing the full set of assay components. We hypothesized that the enzyme might be surface active, disturbing the emulsion interface, but emulsions of an inactivated form of the enzyme were stable (Figure 2(a)). One possible explanation is that active HOx may interact with the detergent through the active site. Adding bovine serum albumin (BSA), which is known to have a stabilizing effect,12 to the mixture improved droplet stability (Figure 2(a)). Emulsions of the assay mixture with BSA were stable for more than 1 day at room temperature.Open in a separate windowFIG. 2.(a) Transmission light micrographs of water-in-perfluorinated-oil emulsions produced using the microfluidic emulsification devices after 2 h incubation at room temperature. The emulsions contain 3 U/ml HOx either in its native form (left image), inactivated by heating at 99 °C for 20 min (middle image), or supplemented with 1 mg/ml BSA (right image). (b) Images of the results of the agar plate Congo Red cellulase assay before and after sorting, with the percentage of positive colonies indicated. The cells expressing cellulase activity show clear hallos.The time required for the cellulase reaction to produce detectable quantities of fluorescent product was monitored using the droplet screening instrument. These devices proved to have a higher sensitivity than the FACS system because the optics are designed for the droplet size selected for the assay. We were able to detect cellulase activity just 20 min after the compartmentalization of cells. This shorter incubation time allowed us to couple the emulsification device directly to the droplet sorting device using a short piece of tubing. The rate of emulsion flow and the dimensions of the tube set the droplet incubation time.Using the optimized conditions, we used droplet microfluidics to sort cellulase-expressing cells from a set of reference libraries. The reference libraries were created by mixing different concentrations of positive S. cerevisiae YPH500 cells expressing Cel5A cellulase and negative S. cerevisiae YPH500 cells transformed with the pESC-Trp empty vector. The mixed populations were emulsified together with the assay components in water-in-perfluorinated-oil emulsions and incubated at room temperature for 20 min. The gated population was sorted and the cells were spread on yeast nitrogen base casaminoacids (YNB CAA) Glu agar plates. An aliquot of the reference library was also plated on agar plates prior to sorting. Approximately, 100 cells before and after sorting were transferred to YNB CAA CMC Gal/Raf induction plates, and the Congo red assay13 was used to detect cells expressing cellulase. In this assay, colonies of positive cells developed transparent halos around them.14 The results before and after sorting are presented in Figure 2(b).We enriched cellulase-expressing cells from a pool of negative cells, regardless of the starting concentration of positive cells. We were able to isolate the cellulase-expressing cells even when starting from a low percentage of active cells (0.1%). We obtained high enrichment factors of up to 300 when starting from low concentrations of positive cells, and we were able to sort to a purity of greater than 90%. These results exceed those obtained by comparable experiments using FACS.5In conclusion, we developed a high-throughput screening system for cellulase activity based on droplet microfluidics. We optimized the emulsification conditions to produce highly stable and monodisperse droplets. The low dispersity of the emulsion enables the sensitive, tunable, and quantitative detection of cellulase activity. In addition, we substantially reduced the reaction time needed for the development of a fluorescent product from 2 h to 20 min. As a result, we sorted reference libraries of cellulases with various ratios of positive to negative cells, and regardless of the starting population of positive cells we were always able to enrich the active population to a higher purity than that obtained by FACS. 相似文献
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A total of 104 six-year-old children belonging to 4 groups (English monolinguals, Chinese-English bilinguals, French-English bilinguals, Spanish-English bilinguals) were compared on 3 verbal tasks and 1 nonverbal executive control task to examine the generality of the bilingual effects on development. Bilingual groups differed in degree of similarity between languages, cultural background, and language of schooling. On the executive control task, all bilingual groups performed similarly and exceeded monolinguals; on the language tasks the best performance was achieved by bilingual children whose language of instruction was the same as the language of testing and whose languages had more overlap. Thus, executive control outcomes for bilingual children are general but performance on verbal tasks is specific to factors in the bilingual experience. 相似文献
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