Gene transfer into conchospores of Porphyra haitanensis (Bangiales, Rhodophyta) by glass bead agitation

Genetic transformation by electroporation of protoplasts is a standard procedure for many plants. However, for the genus Porphyra, the method is not effective because of low viability of protoplasts and is a time-consuming and expensive procedure. Based on the life history of Porphyra, a spore-targeted strategy of genetic transformation was developed, that is, using fresh conchospores of Porphyra haitanensis Chang & Zheng transformed by agitation with glass beads. A SV40 promoter-driven lacZ reporter gene was expressed in conchospores 48 h after the agitation. More transformants were obtained by increasing the agitation time from 10 to 25 s. The maximum number of transformants was more than six out of 1 million agitated conchospores. Transfer of a SV40 promoter-driven egfp gene into conchospores resulted in significant green GFP fluorescence. The expression of lacZ and egfp revealed that this strategy of spore-targeted transformation using glass bead agitation is feasible in P. haitanensis and that the SV40 promoter is effective for monitoring expression of foreign genes in this red algal species.

Transformation of Platymonas (Tetraselmis) subcordiformis (Prasinophyceae, Chlorophyta) by agitation with glass beads

A transient transformation system for the unicellular marine green alga, Platymonas subcordiformis, was established in this study. We introduced the pEGFP-N1 vector into P. subcordiformis with a glass bead method. P. subcordiformis was incubated in cell wall lytic enzymes (abalone acetone powder and cellulase solutions) to degrade the cell wall. The applicable conditions for production of viable protoplasts were pH 6.5, 25 degrees C, and 3 h of enzyme treatment. The protoplast yield was 61.2% when P. subcordiformis cells were added to the enzyme solution at a concentration of 10(7) cell ml(-1). The protoplasts were immediately transformed with the pEGFP-N1 vector using glass-bead method. The transformation frequency was about 10(-5), and there was no GFP activity observed in either the negative or the blank controls. This study indicated that GFP was a sensitively transgenic reporter for P. subcordiformis, and the method of cell wall enzymolysis followed by glass bead agitation was applicable for the transformation of P. subcordiformis.

Improvement in growth and toxin production of Alexandrium tamarense by two-step culture method

The growth and toxin content of the dinoflagellate Alexandrium tamarense ATHK was markedly affected by culture methods. In early growth phase at lower cell density static or mild agitation methods were beneficial to growth, but continuous agitation or aeration, to some extent, had an adverse effect on cell growth. Static culture in 2 L Erlenmeyer flasks had the highest growth rate (0.38 d(-1)) but smaller cell size compared with other culture conditions. Cells grown under aerated conditions possessed low nitrogen and phosphorus cell yields, namely high N and P cell-quota. At day 18, cells grown in continuous agitated and 1 h aerated culture entered the late stationary phase and their cellular toxin contents were higher (0.67 and 0.54 pg cell(-1)) compared with cells grown by other culture methods (0.27-0.49 pg cell(-1)). The highest cell density and cellular toxin content were 17190 cells mL(-1) and 1.26 pg cell(-1) respectively in an airlift photobioreactor with two-step culture. The results indicate that A. tamarense could be grown successfully in airlift photobioreactor by a two-step culture method, which involved cultivating the cells statically for 4 days and then aerating the medium. This provides an efficient way to enhance cell and toxin yield of A. tamarense.

Controlled nucleation and growth of surface-confined gold nanoparticles on a (3-aminopropyl)trimethoxysilane-modified class slide: A strategy for SPR substrates

The thickness of the gold film and its morphology, including the surface roughness, are very important for getting a good, reproducible response in the SPR technique. Here, we report a novel alternative approach for preparing SPR-active substrates that is completely solution-based. Our strategy is based on self-assembly of the gold colloid monolayer on a (3-aminopropyl)trimethoxysilane-modified glass slide, followed by electroless gold plating. Using this method, the thickness of films can be easily controlled at the nanometer scale by setting the plating time in the same conditions. Surface roughness and morphology of gold films can be modified by both tuning the size of gold nanoparticles and agitation during the plating. Surface evolution of the Au film was followed in real time by UV-vis spectroscopy and in situ SPRS. To assess the surface roughness and electrochemical stability of the Au films, atomic force microscopy and cyclic voltammetry were used. In addition, the stability of the gold adhesion is demonstrated by three methods. The as-prepared Au films on substrates are reproducible and stable, which allows them to be used as electrodes for electrochemical experiments and as platforms for studying SAMs.

Tank cultivation of the red alga Palmaria palmata: Effects of intermittent light on growth rate, yield and growth kinetics

Tank cultivation of marine macroalgae involves air-agitation of the algal biomass and intermittent light conditions, i.e. periodic, short light exposure of the thalli in the range of 10 s at the water surface followed by plunging to low light or darkness at the tank bottom and recirculation back to the surface in the range of 1-2 min. Open questions relate to effects of surface irradiance on growth rate and yield in such tumble cultures and the possibility of chronic photoinhibition in full sunlight. A specially constructed shallow-depth tank combined with a dark tank allowed fast circulation times of approximately 5 s, at a density of 4.2 kg fresh weight (FW) m(-2) s(-1). Growth rate and yield of the red alga Palmaria palmata increased over a wide range of irradiances, with no signs of chronic photoinhibition, up to a growth-saturating irradiance of approximately 1600 mumol m(-2) s(-1) in yellowish light supplied by a sodium high pressure lamp at 16 h light per day. Maximum growth rate ranged at 12% FW d(-1), and maximum yield at 609 g FW m(-2) d(-1). This shows that high growth rates of individual thalli may be reached in a dense tumble culture, if high surface irradiances and short circulation times are supplied. Another aspect of intermittent light relates to possible changes of basic growth kinetics, as compared to continuous light. For this purpose on-line measurements of growth rate were performed with a daily light reduction by 50% in light-dark cycles of 1, 2 or 3 min duration during the daily light period. Growth rates at 10degreesC and 50 mumol photon m(-2) s- 1 dropped in all three intermittent light regimes during both the main light and dark periods and reached with all three periodicities approximately 50% of the control, with no apparent changes in basic growth kinetics, as compared to continuous light.

Rapid optimization of process conditions for cultivation of transgenic Laminaria japonica gametophyte cells in a stirred-tank bioreactor

Batch cultivation for transgenic kelp gametophyte cells was investigated in an online controlled 5 L stirred-tank photo-bioreactor to rapidly optimize the process conditions by monitoring the rate of increase of pH. The transgenic kelp gametophytes with heterologous gene encoding hepatitis B surface antigen (HBsAg) could rapidly grow in the bioreactor. Optimal temperature and agitation rate for bioreactor cultivation of gametophytes were 15 degrees C and 200 rpm. Optimal incident light intensities depended on the initial cell densities. (c) 2006 Elsevier B.V. All fights reserved.

学习不良少年的自我意识情绪研究

Self-conscious emotions (guilt, shame, embarrassment, pride, etc) are social emotions, and involve complex appraisals of how one’s behavior has been evaluated by the self and other people according to some value standards. Self-conscious emotions play an important role in human life by arousing and regulating human action tendencies, feeling and thoughts, which can promote people to work hard in achievement and task fields, maintain good interpersonal relationship according with social morality and expectation. The present study aimed to examine complex self-conscious emotional understanding capabilities in junior middle school students with and without learning disabilities, how the self-conscious emotions generate, and relationship between self-conscious emotions and self-representation in academic and interpersonal fields. Situational experimental methods were used in this research, and the results would give further supports for learning disabilities intervention. The main results of present research are as follows. 1. The study included 4 parts and 6 experiments. The aim of study 1 was to explore whether juveniles with learning disabilities understood complex self-conscious emotions differently from juveniles without learning disabilities. We surveyed the self-conscious emotions understanding of 37 learning disabilities and 45 non-learning disabilities with the emotional situation stories. The results indicated that the self-conscious emotional recognition in others for learning disabilities was lower than that of non-learning disabilities in different emotional recognition tasks. Moreover, children with learning disabilities were more inclined to recognize emotions in themselves as elemental emotions, however, children without learning disabilities were more inclined to recognize emotions in themselves as self-conscious emotions. 2. The aim of study 2 was to explore the generative mechanism of self-conscious emotions in academic and interpersonal fields with the method of situational experiments, namely to examine whether the self-discrepancy could cause self-conscious emotions for learning disabilities. 84 learning disabilities (in experiment 1) and 80 learning disabilities (in experiment 2) participated in the research, and the results were as follows. (1) Self discrepancy caused participants’ self-conscious emotions effectively in academic and interpersonal fields. One’s own and parents’ perspercive on the actual-ideal self-discrepancy both produced dejection-related emotions (shame、embarrassment) and agitation-related emotions (guilt). （2）In academic fields, children with learning disabilities caused higher level negative self-conscious emotions (embarrassment, shame, and guilt) and lower level positive self-conscious emotion (pride). However, there were no differences of self-conscious emotions for children with and without learning disabilities in non-academic fields. 3. The aim of study 3 was to explore what influence had self-conscious emotions on self-representation for learning disabilities with the method of situational experiments. 57 learning disabilities (in experiment 1) and 67 learning disabilities (in experiment 2) participated in the research, and the results were as follows. （1）The negative self-conscious for learning disabilities could influence their positive or negative academic and positive interpersonal self-representation stability, the ways in which self-evaluation of ability mediate these effects. However, there was no significant effect for the negative self-conscious and self-evaluation of ability predicting negative interpersonal self-representation stability. （2）The stability level of positive academic and interpersonal self-representation for learning disabilities was lower than that of non-learning disabilities. There was no significant difference of the negative interpersonal self-representation stability for children with and without learning disabilities in the positive self-conscious valence condition. However, the stability level of negative interpersonal self-representation for learning disabilities was lower than that of non-learning disabilities in the negative self-conscious valence condition. 4. The aim of study 4 was to explore the intervention effects for self-conscious emotions training course on emotional comprehension cability. 65 learning disabilities (34 in experimental group, and 31 in control group) participated in the research. The results showed that self-conscious emotions course boosted the self-conscious emotions apprehensive level for children with learning disabilities.