眼点拟微绿球藻对抗生素的敏感性及其无菌藻株的培养

研究了5种抗生素在不同浓度下对眼点拟微绿球藻(Nannochloropsis ocutala)生长的影响。结果表明:浓度为50μg·mL^-1时,氯霉素对眼点拟微绿球藻生长的抑制作用最明显,抑制率高达85.30%,2d和4d的Ic₅₀分别为293.64μg·mL^-1和67.69μg·mL^-1。其次为青霉素、硫酸庆大霉素和硫酸卡那霉素,4d的抑制率分别为8.95%、7.62%和3.84%,其中硫酸庆大霉素2d的Ic₅₀为316.40μg·mL^-1。链霉素对藻细胞生长的抑制作用最轻,抑制率仅为3%。按照抗生素对眼点拟微绿球藻生长抑制作用,由强到弱依次加入50μg·mL^-1青霉素,50μg·mL^-1硫酸庆大霉素,100μg·mL^-1硫酸卡那霉素和50μg·mL^-1链霉素,获得了无菌藻株。     

Enhanced lipid production in Nannochloropsis sp. using fluorescence‐activated cell sorting

To advance the utilization of microalgae as a viable feedstock for biodiesel production, the intracellular lipid content of three strains of the marine microalgae Nannochloropsis sp. was enhanced using flow cytometry (FC) coupled with cell sorting. Total lipid content was doubled to 55% (biomass dry weight) in the sorted, daughter cells of Nannochloropsis (strain 47) after consecutive three rounds of cell sorting, and this trait was maintained for approximately 100 subsequent cell generations. In addition, daughter cells had a fatty acid profile similar to that of the parent, wild‐type strain. The study demonstrates that FC coupled with cell sorting is a powerful tool for the enhancement of intracellular lipid content in microalgae exploited for biodiesel feedstock.     

High-efficiency nuclear transformation of the oleaginous marine Nannochloropsis species using PCR product

Nannochloropsis are model species for investigating biofuel production by algae. To develop them into an integrated photons-to-fuel production platform, high efficiency transformation methods are necessary. Here, we obtained the β-tubulin promoter regions of all recognized species of genus Nannochloropsis, and successfully transformed all five marine species by electroporation. In addition, the PCR amplified double stranded DNA fragments (PCR fragments) based transformation system was established in these Nannochloropsis species, which showed much higher transformation efficiency (10.7–61.2 × 10^?6, 1.5–13-fold) than that of linearized plasmid based transformation. The cotransformation of N. salina using a circular plasmid containing a non-selectable GUS gene and a PCR fragment containing only a selection marker cassette was also achieved and found to be very efficient (over 50%). This simple and highly efficient transformation protocol reported in our study provided a useful tool for gene functional analysis and genetic engineering of the oleaginous Nannochloropsis species.     

Process optimization and modeling for the cultivation of Nannochloropsis sp. and Tetraselmis striata via response surface methodology

The aim of this study was to determine the optimal physical process conditions for the cultivation of locally isolated strains of Nannochloropsis sp. and Tetraselmis striata to achieve maximum growth rate. It was essential to evaluate biomass production at different agitation rates, light intensities, and temperature levels. Central composite design and response surface methodology were applied to design the experiments and optimize the cultivation process for Nannochloropsis sp. and T. striata. The specific growth rate of 0.250 d^?1 was obtained for Nannochloropsis sp. cells under the light intensity of 54 μmol photons · m^?2 · s^?1, at the agitation rate of 151 rpm in 24.5°C. The optimal physical process conditions for T. striata were obtained under the light intensity of 56 μmol photons · m^?2 · s^?1 in 25.5°C at the agitation rate of 151 rpm in 25.5°C, resulting in a specific growth rate of 0.226 d^?1. The predicted values were justified by the verification tests. Good agreement between the predicted values and the experimental values confirmed the validity of the models for the cultivation of microalgal strains. In this article, the noteworthy result was that temperature was a dominant factor in obtaining high chl‐a content for Nannochloropsis sp., whereas the growth of T. striata strongly depended on light exposure.     

响应面法对微拟球藻产微藻蛋白的发酵条件优化

以稳定期微藻蛋白浓度为评价指标,利用响应面设计对微拟球藻(Nannochloropsis gaditana)的分批发酵条件进行优化。在单因素试验的基础上,选取温度、p H、搅拌速度及通气量为影响因子,采用四因素三水平的Box-Benhnken中心组合法设计试验。结果表明:微拟球藻的最佳发酵条件为温度30℃、p H 6.9、搅拌速度340 r/min以及通气量0.65 vvm,在此优化条件下得到微藻蛋白浓度为6.18 g/L,与模型预测值基本相符,较优化前提高了9.18%。     

Enriching Rotifers with “Premium” Microalgae. Nannochloropsis gaditana

The nutritive quality of Nannochloropsis gaditana cultured semicontinuously with different daily renewal rates was tested as a diet for short-term enrichment of the rotifer Brachionus plicatilis. After 24 h, dramatic differences in the survival, dry weight, and biochemical composition of the rotifers depending on the renewal rate of microalgal cultures were observed. Survival after the feeding period increased with increasing renewal rates. Rotifers fed microalgae from low renewal rate, nutrient-deficient cultures showed low dry weight and organic contents very similar to those of the initial rotifers that were starved for 12 h before the start of the feeding period. On the contrary, rotifers fed nutrient-sufficient microalgal cells underwent up to twofold increases of dry weight and protein, lipid, and carbohydrate contents with regard to rotifers fed nutrient-depleted N. gaditana. Consequently, feed conversion rate decreased in these conditions, indicating a better assimilation of the microalgal biomass obtained at high renewal rates. No single microalgal biochemical parameter among those studied can explain the response of the filter feeder. Similarly to gross composition, EPA and n-3 contents in rotifers fed microalgae from nutrient-sufficient cultures were double than the contents found in rotifers fed nutrient-limited microalgae. In addition, very high positive correlations between the contents of EPA and n-3 in N. gaditana and B. plicatilis were observed. These results demonstrate that selecting the appropriate conditions of semicontinuous culture can strongly enhance the nutritional value of microalgae that is reflected in the growth and biochemical composition of the filter-feeder even in short exposure periods.     

A screening model to predict microalgae biomass growth in photobioreactors and raceway ponds

A microalgae biomass growth model was developed for screening novel strains for their potential to exhibit high biomass productivities under nutrient‐replete conditions in photobioreactors or outdoor ponds. Growth is modeled by first estimating the light attenuation by biomass according to Beer‐Lambert's Law, and then calculating the specific growth rate in discretized culture volume slices that receive declining light intensities due to attenuation. The model uses only two physical and two species‐specific biological input parameters, all of which are relatively easy to determine: incident light intensity, culture depth, as well as the biomass light absorption coefficient and the specific growth rate as a function of light intensity. Roux bottle culture experiments were performed with Nannochloropsis salina at constant temperature (23°C) at six different incident light intensities (10, 25, 50, 100, 250, and 850 µmol/m² s) to determine both the specific growth rate under non‐shading conditions and the biomass light absorption coefficient as a function of light intensity. The model was successful in predicting the biomass growth rate in these Roux bottle batch cultures during the light‐limited linear phase at different incident light intensities. Model predictions were moderately sensitive to minor variations in the values of input parameters. The model was also successful in predicting the growth performance of Chlorella sp. cultured in LED‐lighted 800 L raceway ponds operated in batch mode at constant temperature (30°C) and constant light intensity (1,650 µmol/m² s). Measurements of oxygen concentrations as a function of time demonstrated that following exposure to darkness, it takes at least 5 s for cells to initiate dark respiration. As a result, biomass loss due to dark respiration in the aphotic zone of a culture is unlikely to occur in highly mixed small‐scale photobioreactors where cells move rapidly in and out of the light. By contrast, as supported also by the growth model, biomass loss due to dark respiration occurs in the dark zones of the relatively less well‐mixed pond cultures. In addition to screening novel microalgae strains for high biomass productivities, the model can also be used for optimizing the pond design and operation. Additional research is needed to validate the biomass growth model for other microalgae species and for the more realistic case of fluctuating temperatures and light intensities observed in outdoor pond cultures. Biotechnol. Bioeng. 2013; 110: 1583–1594. © 2012 Wiley Periodicals, Inc.     

CO2激光辐照拟微绿球藻的当代生物学效应研究

采用CO₂激光（波长10.6μm,功率10 w,光束长74 cm）辐照拟微绿球藻（Nannochloropsis sp.YW0980）,辐照时间为30 s、60 s、90 s,通过测定藻色素、多糖、蛋白质及油脂含量,研究CO₂激光对藻的生物学效应。结果表明:30 s、60 s、90 s辐照条件下,对拟微绿球藻细胞生长及代谢产物均有一定的促进作用,其中CO₂激光60 s处理组有利于拟微绿球藻的生长及色素的积累,但30 s剂量下更有利于多糖、蛋白质及油脂含量的积累,分别比对照组提高了51.05%（胞外多糖）,289.45%（总多糖）、37.05%、172.16%。