Abstract
BackgroundMicroalgae have recently been attracting attention as a potential platform for the production of biofuels. Euglena gracilis, a unicellular phytoflagellate, has been proposed as an attractive feedstock to produce biodiesel because it can produce large amounts of wax esters, consisting of medium-chain fatty acids and alcohols with 14:0 carbon chains. E. gracilis cells highly accumulate a storage polysaccharide, a β-1,3-glucan known as paramylon, under aerobic conditions. When grown aerobically and then transferred into anaerobic conditions, E. gracilis cells degrade paramylon to actively synthesize and accumulate wax esters. Thus, the enhanced accumulation of paramylon through the genetic engineering of photosynthesis should increase the capacity for wax ester production.ResultsWe herein generated transgenic Euglena (EpFS) cells expressing the cyanobacterial fructose-1,6-/sedoheptulose-1,7-bisphosphatase (FBP/SBPase), which is involved in the Calvin cycle, to enhance its photosynthetic activity. FBP/SBPase was successfully expressed within Euglena chloroplasts. The cell volume of the EpFS4 cell line was significantly larger than that of wild-type cells under normal growth conditions. The photosynthetic activity of EpFS4 cells was significantly higher than that of wild type under high light and high CO2, resulting in enhanced biomass production, and the accumulation of paramylon was increased in transgenic cell lines than in wild-type cells. Furthermore, when EpFS cell lines grown under high light and high CO2 were placed on anaerobiosis, the productivity of wax esters was approximately 13- to 100-fold higher in EpFS cell lines than in wild-type cells.ConclusionOur results obtained here indicate that the efficiency of biomass production in E. gracilis can be improved by genetically modulating photosynthetic capacity, resulting in the enhanced production of wax esters. This is the first step toward the utilization of E. gracilis as a sustainable source for biofuel production under photoautotrophic cultivation.Electronic supplementary materialThe online version of this article (doi:10.1186/s13068-015-0264-5) contains supplementary material, which is available to authorized users.
Highlights
Microalgae have recently been attracting attention as a potential platform for the production of biofuels
Generation of transgenic E. gracilis cells expressing cyanobacterial FBP/SBPase To generate transgenic E. gracilis, the cyanobacterial FBP/SBPase gene with a tomato rubisco small subunit transit peptide driven by the 35S promoter and neomycin phosphotransferase Neomycin phosphotransferase II (II) (NPT II) gene as an antibiotic-resistant marker with the nopaline synthase (NOS) promoter was introduced into E. gracilis wildtype cells by microprojectile bombardment (Fig. 1a)
Genomic DNA was isolated from wild-type and randomly selected six EpFS cell lines, and the transgene was analyzed by genomic PCR with gene-specific primers for the FBP/SBPase sequence
Summary
Microalgae have recently been attracting attention as a potential platform for the production of biofuels. E. gracilis cells highly accumulate a storage polysaccharide, a β-1,3-glucan known as paramylon, under aerobic conditions. The enhanced accumulation of paramylon through the genetic engineering of photosynthesis should increase the capacity for wax ester production. Extensive research has been conducted to identify useful microalgae all over the world and improve biomass production through the metabolic engineering of known microalgae [2]. Several problems such as the low productivity of microalgae and difficulties in consistently producing biomass at a large scale in highly variable outdoor culture conditions need to be overcome before microalgae can be used as an economically viable biofuel feedstock. Genetic transformations of more than 30 different strains of microalgae have been reported [3], further research is still required to fully maximize microalgal biomass production for biofuels
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