Characterization and evaluation of substratum material selection for microalgal biofilm cultivation.

Biofilm cultivation is considered a promising method to achieve higher microalgae biomass productivity with less water consumption and easier harvest compared to conventional suspended cultivation. However, studies focusing on the selection of substratum material and optimization of the growth of certain microalgae species on specific substratum are limited. This study investigated the selection of membranous and fabric fiber substrata for the attachment of unicellular microalgae Scenedesmus dimorphus and filamentous microalgae Tribonema minus in biofilm cultivation. The results indicated that both algal species preferred hydrophilic membranous substrata and nitrate cellulose/cellulose acetate membrane (CN-CA) was selected as a suitable candidate on which the obtained biomass yields were up to 10.24 and 7.81 g m-2 day-1 for S. dimorphus and T. minus, respectively. Furthermore, high-thread cotton fiber (HCF) and low-thread polyester fiber (LPEF) were verified as the potential fabric fiber substrata for S. dimorphus (5.42 g m-2 day-1) and T. minus (5.49 g m-2 day-1) attachment, respectively. The regrowth of microalgae biofilm cultivation strategy was applied to optimize the algae growth on the fabric fiber substrata, with higher biomass density and shear resistibility achieved for both algal species. The present data highlight the importance to establish the standards for selection the suitable substratum materials in ensuring the high efficiency and sustainability of the attached microalgal biomass production. KEY POINTS: • CN-CA was suitable membranous substratum candidate for algal biofilm cultivation. • HCF and LPEF were potential fabric fiber substrata for S. dimorphus and T. minus. • Regrowth biofilm cultivation was effective in improving algal biomass and attachment.

Identification and Functional Analysis of Acyl-Acyl Carrier Protein Δ9 Desaturase from Nannochloropsis oceanica.

The oleaginous marine microalga Nannochloropsis oceanica strain IMET1 has attracted increasing attention as a promising photosynthetic cell factory due to its unique excellent capacity to accumulate large amounts of triacylglycerols and eicosapentaenoic acid. To complete the genomic annotation for genes in the fatty acid biosynthesis pathway of N. oceanica, we conducted the present study to identify a novel candidate gene encoding the archetypical chloroplast stromal acyl-acyl carrier protein Δ9 desaturase. The full-length cDNA was generated using rapid-amplification of cDNA ends, and the structure of the coding region interrupted by four introns was determined. The RT-qPCR results demonstrated the upregulated transcriptional abundance of this gene under nitrogen starvation condition. Fluorescence localization studies using EGFP-fused protein revealed that the translated protein was localized in chloroplast stroma. The catalytic activity of the translated protein was characterized by inducible expression in Escherichia coli and a mutant yeast strain BY4389, indicating its potential desaturated capacity for palmitoyl-ACP (C16:0-ACP) and stearoyl-ACP (C18:0-ACP). Further functional complementation assay using BY4839 on plate demonstrated that the expressed enzyme restored the biosynthesis of oleic acid. These results support the desaturated activity of the expressed protein in chloroplast stroma to fulfill the biosynthesis and accumulation of monounsaturated fatty acids in N. oceanica strain IMET1.

Pretreatment and enzymatic hydrolysis optimization of lignocellulosic biomass for ethanol, xylitol, and phenylacetylcarbinol co-production using Candida magnoliae.

Cellulosic bioethanol production generally has a higher operating cost due to relatively expensive pretreatment strategies and low efficiency of enzymatic hydrolysis. The production of other high-value chemicals such as xylitol and phenylacetylcarbinol (PAC) is, thus, necessary to offset the cost and promote economic viability. The optimal conditions of diluted sulfuric acid pretreatment under boiling water at 95°C and subsequent enzymatic hydrolysis steps for sugarcane bagasse (SCB), rice straw (RS), and corn cob (CC) were optimized using the response surface methodology via a central composite design to simplify the process on the large-scale production. The optimal pretreatment conditions (diluted sulfuric acid concentration (% w/v), treatment time (min)) for SCB (3.36, 113), RS (3.77, 109), and CC (3.89, 112) and the optimal enzymatic hydrolysis conditions (pretreated solid concentration (% w/v), hydrolysis time (h)) for SCB (12.1, 93), RS (10.9, 61), and CC (12.0, 90) were achieved. CC xylose-rich and CC glucose-rich hydrolysates obtained from the respective optimal condition of pretreatment and enzymatic hydrolysis steps were used for xylitol and ethanol production. The statistically significant highest (p ≤ 0.05) xylitol and ethanol yields were 65% ± 1% and 86% ± 2% using Candida magnoliae TISTR 5664. C. magnoliae could statistically significantly degrade (p ≤ 0.05) the inhibitors previously formed during the pretreatment step, including up to 97% w/w hydroxymethylfurfural, 76% w/w furfural, and completely degraded acetic acid during the xylitol production. This study was the first report using the mixed whole cells harvested from xylitol and ethanol production as a biocatalyst in PAC biotransformation under a two-phase emulsion system (vegetable oil/1 M phosphate (Pi) buffer). PAC concentration could be improved by 2-fold compared to a single-phase emulsion system using only 1 M Pi buffer.

Producing high value unsaturated fatty acid by whole-cell catalysis using microalga: A case study with Tribonema minus.

High value unsaturated fatty acids can be produced by de novo synthesis in microalgal cells, especially via heterotrophic cultivation. Unfortunately, the lipid accumulation of heterotrophic microalgae cannot be improved efficiently in conventional ways. Here we reported heterotrophic Tribonema minus, a promising resource for the production of palmitoleic acid which has increasing demands in health service for patients with metabolic syndrome, as whole-cell biocatalyst to develop a novel way of shifting low value exogenous saturated fatty acids to high value ones. Results showed that myristic acid is the best precursor for whole-cell catalysis; it elevated the lipid content of T. minus to 42.2%, the highest among the tried precursors. The influences of cultivation condition on the utilization of extrinsic myristic acid and lipid accumulation were also determined. Under the optimized condition, the lipid content reached as high as 48.9%. In addition, our findings showed that ~13.0% of C16:1 in T. minus is derived from extrinsic myristic acid, and 30.1% of metabolized precursor is converted into heterologous fatty acids. Thus, a feasible approach for both increasing the value of low value saturated fatty acid by bioconversion and enhancing the lipid accumulation in microalgae is proposed by supplementing extrinsic myristic acid.

Modeling and improving arrayed microalgal biofilm attached culture system.

A microalgal biofilm-attached-system is an alternative cultivation method, that offers potential advantages of improved biomass productivity, efficient harvesting, and water saving. These biofilm systems have been widely tested and utilized for microalgal biomass production and wastewater treatment. This research a microalgal growth model for the biofilm attached culture system has been developed and experimentally validated, both, in single and arrayed biofilm systems. It has been shown that the model has the capability to accurately describe microalgae growth. Moreover, via the model simulation, it was observed that system structural parameters, light dilution rate, and light intensity significantly affected the culture performance. The limitations, and improvement aspects of the model, are also discussed in this study. To our knowledge, this is the first time that a mathematical model for an arrayed-biofilm-attached-system has been developed and validated. This model will certainly be helpful in the design, improvement, optimization, and evaluation of the biofilm-attached-systems.

Impacts of surface wettability and roughness of styrene-acrylic resin films on adhesion behavior of microalgae Chlorella sp.

Biofilm attached cultivation is a promising method for efficient production of microalgae. Determining the surface property index to select an appropriate substrate benefiting the algae adhesion and biofilm formation is very important for the cultivation method. This work focused on elucidating and quantifying the influence of surface wettability and roughness of substrate on Chlorella vulgaris adhesion. Firstly, surface modified styrene-acrylic (SA) resin films by adding different dosage of perfluoroalkyl ethyl acrylate (FM) were prepared. Property characterization shows that the surface contact angle in water, formamide and diiodomethane of FM modified SA films is significantly associated with the FM dosage, while the other surface properties including zeta potential, surface potential and surface roughness have insignificant difference. The calculated surface free energy parameters show that the SA films belong to the non-polar substrata. A well quantitative correlation that the adhesion capacity of C. vulgaris linearly declines with the increase of water contact angle was obtained. And a near linear relationship between the adhesion capacity and the surface free energy (γ), or the cohesion free energy (ΔGcoh) was also observed. Secondly, the surface roughness solely changed SA films were prepared by replicating the morphology of stainless steel sieves through the PDMS template method. The patterned SA films have alternately arranged rectangular "valleys" and "ridges". A well linear correlation between the microalgae adhesion capacity and the surface roughness was also obtained.

Chloroplast Genetic Engineering of a Unicellular Green Alga Haematococcus pluvialis with Expression of an Antimicrobial Peptide.

The purpose of this study was to express an antimicrobial peptide in the chloroplast to further develop the plastid engineering of H. pluvialis. Homologous targeting of the 16S-trnI/trnA-23S region and four endogenous regulatory elements, including the psbA promoter, rbcL promoter, rbcL terminator, and psbA terminator in H. pluvialis, were performed to construct a chloroplast transformation vector for H. pluvialis. The expression of codon-optimized antimicrobial peptide piscidin-4 gene (ant1) and selection marker gene (bar, biolaphos resistance gene) in the chloroplast of H. pluvialis was controlled by the rbcL promoter and psbA promoter, respectively. Upon biolistic transformation and selection with phosphinothricin, integration and expression of ant1 in the chloroplast genome were detected using polymerase chain reaction (PCR), southern blotting, and western blotting. Using this method, we successfully expressed antimicrobial peptide piscidin-4 in H. pluvialis. Hence, our results showed H. pluvialis promises as a platform for expressing recombinant proteins for biotechnological applications, which will further contribute to promoting genetic engineering improvement of this strain.

Genetic Transformation of Tribonema minus, a Eukaryotic Filamentous Oleaginous Yellow-Green Alga.

Eukaryotic filamentous yellow-green algae from the Tribonema genus are considered to be excellent candidates for biofuels and value-added products, owing to their ability to grow under autotrophic, mixotrophic, and heterotrophic conditions and synthesize large amounts of fatty acids, especially unsaturated fatty acids. To elucidate the molecular mechanism of fatty acids and/or establish the organism as a model strain, the development of genetic methods is important. Towards this goal, here, we constructed a genetic transformation method to introduce exogenous genes for the first time into the eukaryotic filamentous alga Tribonema minus via particle bombardment. In this study, we constructed pSimple-tub-eGFP and pEASY-tub-nptⅡ plasmids in which the green fluorescence protein (eGFP) gene and the neomycin phosphotransferase Ⅱ-encoding G418-resistant gene (nptⅡ) were flanked by the T. minus-derived tubulin gene (tub) promoter and terminator, respectively. The two plasmids were introduced into T. minus cells through particle-gun bombardment under various test conditions. By combining agar and liquid selecting methods to exclude the pseudotransformants under long-term antibiotic treatment, plasmids pSimple-tub-eGFP and pEASY-tub- nptⅡ were successfully transformed into the genome of T. minus, which was verified using green fluorescence detection and the polymerase chain reaction, respectively. These results suggest new possibilities for efficient genetic engineering of T. minus for future genetic improvement.

Comparison of Lipid and Palmitoleic Acid Induction of Tribonema minus under Heterotrophic and Phototrophic Regimes by Using High-Density Fermented Seeds.

Palmitoleic acid, one scarce omega-7 monounsaturated fatty acid, has important applications in the fields of medicine and health products. Tribonema has been considered as a promising candidate for the production of palmitoleic acid due to its high lipid and palmitoleic acid content and remarkable heterotrophic ability. The high-density heterotrophic cultivation of Tribonema minus was conducted in this work, and the highest biomass of 42.9 g L-1 and a relatively low lipid content of 28.7% were observed. To further enhance the lipid and palmitoleic acid accumulation, induction strategies under two regimes of phototrophy and heterotrophy with different conditions were investigated and compared. Results demonstrated encouraging promotions both by heterotrophic and phototrophic ways, and the final lipid contents reached 41.9% and 49.0%, respectively. In consideration of the time cost, however, the induction under heterotrophic conditions was much more advantageous, by which the highest lipid and palmitoleic acid productivities of 1.77 g L-1 d-1 and 924 mg L-1 d-1 were obtained respectively, with the lipid yield on glucose of 0.26 g g-1.

Biolistic Transformation of Haematococcus pluvialis With Constructs Based on the Flanking Sequences of Its Endogenous Alpha Tubulin Gene.

Haematococcus pluvialis has high commercial value, yet it displays low development of genetic transformation systems. In this research, the endogenous 5' and 3' flanking sequences of the constitutive alpha tubulin (tub) gene were cloned along with its encoding region in H. pluvialis, in which some putative promoter elements and polyadenylation signals were identified, respectively. Three selection markers of tub/aadA, tub/hyr and tub/ble with three different antibiotic-resistance genes fused between the endogenous tub promoter (Ptub) and terminator (Ttub) were constructed and utilized for biolistic transformation of H. pluvialis. Stable resistant colonies with introduced aadA genes were obtained after bombardments of either H. pluvialis NIES144 or SCCAP K0084 with the tub/aadA cassette, the efficiency of which could reach up to 3 × 10-5 per µg DNA through an established manipulation flow. Two key details, including the utilization of culture with motile flagellates dominant and controlled incubation of them on membrane filters during bombardments, were disclosed firstly. In obtained transformants, efficient integration and transcription of the foreign tub/aadA fragments could be identified through genome PCR examination and qPCR analysis, nonetheless with random style instead of homologous crossover in the H. pluvialis genome. The presented selection marker and optimized transforming procedures in this report would strengthen the platform for genetic manipulation and modification of H. pluvialis.

Field study on attached cultivation of Arthrospira (Spirulina) with carbon dioxide as carbon source.

Microalga is considered as a promising candidate for CO2 bio-sequestration. Biofilm attached cultivation is a newly developed technology with many advantages over conventional aqua-suspended methods. In this research, the field performance of this technology was investigated with a 10 m2 pilot system under greenhouse condition by cultivating Arthrospira (Spirulina) platensis with CO2 as carbon source. The system run continuously for two months without contamination bloom. Averaged biomass productivity was 38.3 g m-2 d-1 with protein content over 60% and overall CO2 usage efficiency of 75.1%. Construction cost for the pilot system was over US$200 per m2 which was much higher than that of open pond. However, there was a great reduction space in future large-scale application if the most expensive materials were substituted with cheaper ones. These results indicated the attached cultivation was a promising technology for industrialized application of microalga in CCUS (carbon capture, utilization and storage).

Low pH rather than high CO2 concentration itself inhibits growth of Arthrospira.

Microalga is a promising candidate for bio-mitigation of CO2. It has been longtime recognized that high CO2 concentration would impose stresses on microalga to suppress the growth. However, this concept was challenged in this research by investigating the growth, photosynthesis and anti-oxidant characteristics of Arthrospira platensis under independent effects of CO2 concentrations and pH. Results showed the growth of A. platensis was only inhibited when broth was in acidic pH. Microalgal cells could deal with high CO2 concentration readily if medium pH was maintained in favorite level. Photosynthesis was inhibited swiftly and significantly under acidified condition. The singlet oxygen was produced in low level for alkalic pH treatment, however it burst quickly after low pH stress was imposed. Accordingly, it was proposed that the phenomena of high CO2 intolerance was caused by CO2 induced pH decline rather than high CO2 concentration itself. This finding has significance on large scale application of microalga based CO2 mitigation and flue gas treatment since it proved concentrated CO2 could be directly assimilated without dilution.

Partial Characterization, the Immune Modulation and Anticancer Activities of Sulfated Polysaccharides from Filamentous Microalgae Tribonema sp.

Recently, Tribonema sp., a kind of filamentous microalgae, has been studied for biofuel production due to its accumulation of triacylglycerols. However, the polysaccharides of Tribonema sp. and their biological activities have rarely been reported. In this paper, we extracted sulfated polysaccharides from Tribonema sp. (TSP), characterized their chemical composition and structure, and determined their immunostimulation and anticancer activities on RAW264.7 macrophage cells and HepG2 cells. The results showed that TSP is a sulfated polysaccharide with a Mw of 197 kDa. TSP is a heteropolysaccharide that is composed mainly of galactose. It showed significant immune-modulatory activity by stimulating macrophage cells, such as upregulating interleukin 6 (IL-6), interleukin 10 (IL-10), and tumor necrosis factor α (TNF-α). In addition, TSP also showed significant dose-dependent anticancer activity (with an inhibition rate of up to 66.8% at 250 µg/mL) on HepG2 cells as determined by the 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. The cycle analysis indicated that the anticancer activity of TSP is mainly the result of induced cell apoptosis rather than affecting the cell cycle and mitosis of HepG2 cells. These findings suggest that TSP might have potential as an anticancer resource, but further research is needed, especially in vivo experiments, to explore the anticancer mechanism of TSP.

Mechanism and enhancement of lipid accumulation in filamentous oleaginous microalgae Tribonema minus under heterotrophic condition.

BACKGROUND: The filamentous microalgae Tribonema minus accumulates large amounts of lipids under photoautotrophic condition, while under heterotrophic condition, the lipid content decreased dramatically. Determination of the differences in metabolic pathways between photoautotrophic and heterotrophic growth will provide targets and strategies for improvement of lipid accumulation in heterotrophic cells. METHODS: The metabolic differences between photoautotrophically and heterotrophically cultivated T. minus cells were studied by comparing the growth, biochemical compositions and transcriptomic and metabolomic profiles of the cells. Based on comparative transcriptomic and metabolomic studies, we generated a global model of the changes in central carbon metabolism and lipid biosynthetic pathways that occur under photoautotrophic and heterotrophic growth conditions. Moreover, the specific effects of supplementation with exogenous key metabolic intermediates on the lipid accumulation in heterotrophic culture were analyzed. RESULTS: Compared to photoautotrophic cultures, heterotrophic cultures exhibited enhanced biomass levels and carbohydrate content, but decreased lipid accumulation. These effects were accompanied by low expression levels of genes involved in glycolysis, de novo fatty acids biosynthesis and lipid biosynthesis, and high levels of genes involved in gluconeogenesis. In addition, the levels of key metabolites involved in glycolysis/gluconeogenesis were elevated in abundance, whereas those of certain fatty acids and citric acid were decreased in heterotrophic cultures. Upon supplementation with exogenous potassium palmitate, the lipid content increased dramatically in heterotrophically cultivated T. minus. CONCLUSION: An insufficient supply of carbon precursors caused the low levels of lipid accumulation during heterotrophic cultivation. Appropriate carbon metabolite supplementation based on the metabolomic data was shown to promote lipid accumulation. Moreover, gene regulatory metabolic targets were also identified via omics analysis.

Comparative transcriptome analyses of oleaginous Botryococcus braunii race A reveal significant differences in gene expression upon cobalt enrichment.

BACKGROUND: Botryococcus braunii is known for its high hydrocarbon content, thus making it a strong candidate feedstock for biofuel production. Previous study has revealed that a high cobalt concentration can promote hydrocarbon synthesis and it has little effect on growth of B. braunii cells. However, mechanisms beyond the cobalt enrichment remain unknown. This study seeks to explore the physiological and transcriptional response and the metabolic pathways involved in cobalt-induced hydrocarbon synthesis in algae cells. RESULTS: Growth curves were similar at either normal or high cobalt concentration (4.5 mg/L), suggesting the absence of obvious deleterious effects on growth introduced by cobalt. Photosynthesis indicators (decline in Fv/Fm ratio and chlorophyll content) and reactive oxygen species parameters revealed an increase in physiological stress in the high cobalt concentration. Moreover, cobalt enrichment treatment resulted in higher crude hydrocarbon content (51.3% on day 8) compared with the control (43.4% on day 8) throughout the experiment (with 18.2% improvement finally). Through the de novo assembly and functional annotation of the B. braunii race A SAG 807-1 transcriptome, we retrieved 196,276 non-redundant unigenes with an average length of 1086 bp. Of the assembled unigenes, 89,654 (45.7%), 42,209 (21.5%), and 32,318 (16.5%) were found to be associated with at least one KOG, GO, or KEGG ortholog function. In the early treatment (day 2), the most strongly upregulated genes were those involved in the fatty acid biosynthesis and metabolism and oxidative phosphorylation, whereas the most downregulated genes were those involved in carbohydrate metabolism and photosynthesis. Genes that produce terpenoid liquid hydrocarbons were also well identified and annotated, and 21 (or 29.2%) were differentially expressed along the cobalt treatment. CONCLUSIONS: Botryococcus braunii SAG 807-1 can tolerate high cobalt concentration and benefit from hydrocarbon accumulation. The time-course expression profiles for fatty acid biosynthesis, metabolism, and TAG assembly were obtained through different approaches but had equally satisfactory results with the redirection of free long-chain fatty acid and VLCFA away from TAG assembly and oxidation. These molecules served as precursors and backbone supply for the fatty acid-derived hydrocarbon accumulation. These findings provide a foundation for exploiting the regulation mechanisms in B. braunii race A for improved photosynthetic production of hydrocarbons.

Combined production of fucoxanthin and EPA from two diatom strains Phaeodactylum tricornutum and Cylindrotheca fusiformis cultures.

Fucoxanthin and eicosapentaenoic acid (EPA) provide significant health benefits for human population. Diatom is a potential natural livestock for the combined production of EPA and fucoxanthin. In this study, first, the effects of three important parameters including light intensity, nitrogen concentration and salinity were evaluated for the production of EPA and fucoxanthin in two diatom strains Phaeodactylum tricornutum and Cylindrotheca fusiformis. And then, two steps method based on light intensity were applied to produce EPA and fucoxanthin in large scale. Higher light intensity was first adopted for the high growth rate and lipid content of diatom, and after a period of time, light intensity was lowered to enhance the accumulation of fucoxanthin and EPA. In final, the highest EPA yields were 62.55 and 27.32 mg L-1 for P. tricornutum and C. fusiformis, and the fucoxanthin yield reached 8.32 and 6.05 mg L-1, respectively.

Development of nitrogen supply strategy for Scenedesmus rubescens attached cultivation toward growth and lipid accumulation.

In this study, the microalgae Scenedesmus rubescens were cultivated under the following nitrogen sources, nitrogen concentrations, and nitrogen feeding times (NFTs). This was to help assess biomass and lipid productivity. Scenedesmus rubescens can grow well by adhering to the cellulose acetate membrane in five kinds of nitrogen medium: KNO3, urea, NaNO3, (NH4)2CO3, and NH4NO3. Under the criteria of bio-productivity and lipid productivity, urea was the optimal nitrogen source. Among different urea concentrates, biomass productivity and lipid content of S. rubescens cultivated in 0.27 g/L urea medium were optimized at 8.8 g/(m2 day) and 31.1%, respectively. With attached cultivation, the highest biomass of 9.4 g/m2 was obtained at NFTs of 4 days. These results showed that culturing S. rubescens using urea as sole nitrogen source by improving nitrogen uptake with attached cultivation is more efficient.

Biofilm Attached Cultivation of Chlorella pyrenoidosa Is a Developed System for Swine Wastewater Treatment and Lipid Production.

This study showed the new potential of using soluble contents and heavy metals in swine wastewater as nutrient supplements for the algae Chlorella pyrenoidosa with biofilm attached method. Algae with biofilm attached cultivation grew well in unpasteurized wastewater reaching a biomass productivity of 5.03 g m-2 d-1, lipid content of 35.9% and lipid productivity of 1.80 g m-2 d-1. Chlorella grew in BG11 medium delivered lower values for each of the aforementioned parameters. The FAMEs compositions in the algae paste were mainly consisted of C16:0, C18:2, and C18:3. Algae removed NH4+-N, total phosphorus (TP), and COD by 75.9, 68.4, and 74.8%, respectively. Notably, Zn2+, Cu+, and Fe2+ were removed from wastewater with a ratio of 65.71, 53.64, and 58.89%, respectively. Biofilm attached cultivation of C. pyrenoidosa in swine wastewater containing heavy metals could accumulate considerable biomass and lipid, and the removal ratio of NH4+-N, TP, COD, and as well as heavy metal were high. Treatment of wastewater with biofilm attached cultivation showed an increasingly popular for the concentration of microalgae and environmental sustainability.

Lipid accumulation and metabolic analysis based on transcriptome sequencing of filamentous oleaginous microalgae Tribonema minus at different growth phases.

Filamentous oleaginous microalgae specie Tribonema minus is a promising feedstock for biodiesel production. However, the metabolic mechanism of lipid production in this filamentous microalgal specie remains unclear. Here, we compared the lipid accumulation of T. minus at different growth phases, and described the de novo transcriptome sequencing and assembly and identified important pathways and genes involved in TAG production. Total lipid increased by 2.5-fold and its TAG level in total lipid reached 81.1% at stationary phase. Using the genes involved in the lipid metabolism, the TAG biosynthesis pathways were generated. Moreover, results also demonstrated that, in addition to the observed overexpression of the fatty acid synthesis pathway, TAG production at stationary growth phase was bolstered by repression of the ß-oxidation pathway, up-regulation of genes that funnels acetyl-CoA to lipid biosynthesis, especially gene encoding for phospholipid:diacylglycerol acyltransferase (PDAT) which funnels DAG to TAG biosynthesis.

Biofilm based attached cultivation technology for microalgal biorefineries-A review.

The attached cultivation for microalga has many superiorities over the conventional aqua-suspend methods, which make it a promising pathway to supply feedstock for microalgae based bio-refinery attempts. In this review, the current reports on bioreactor, application, modeling, substratum material and engineering aspects were summarized and the future research and developments should be focused on the following aspects: 1) Build principles and guidelines for rational structure design by studying the relationship of physiological properties with typical structures and light regimes; 2) Set up theory foundation of substratum material selection by studying the physic-chemical properties of algal cells and substratum materials; 3) Further understanding the mass transfer behaviors of both CO2 and nutrients in biofilm for enhanced growth rate and products accumulation; 4) New equipment and machines for inoculation, harvesting and moisture keeping should be developed and integrated with bioreactor structure.