No abstract

To obtain a detailed picture of sulfur deprivation-induced H 2 production in microalgae, metabolome analyses were performed during key time points of the anaerobic H 2 production process of Chlamydomonas reinhardtii. Analyses were performed using gas chromatography coupled to mass spectrometry (GC/MS), two-dimensional gas chromatography combined with time-of-flight mass spectrometry (GCxGC-TOFMS), lipid and starch analysis, and enzymatic determination of fermentative products. The studies were designed to provide a detailed metabolite profile of the solar Bio-H 2 production process. This work reports on the differential analysis of metabolic profiles of the high H 2 -producing strain Stm6Glc4 and the wild-type cc406 (WT) before and during the H 2 production phase. Using GCxGC-TOFMS analysis the number of detected peaks increased from 128 peaks, previously detected by GC/MS techniques, to ϳ1168. More detailed analysis of the anaerobic H 2 production phase revealed remarkable differences between wild-type and mutant cells in a number of metabolic pathways. Under these physiological conditions the WT produced up to 2.6 times more fatty acids, 2.2 times more neutral lipids, and up to 4 times more fermentation products compared with Stm6Glc4. Based on these results, specific metabolic pathways involving the synthesis of fatty acids, neutral lipids, and fermentation products during anaerobiosis in C. reinhardtii have been identified as potential targets for metabolic engineering to further enhance substrate supply for the hydrogenase(s) in the chloroplast.Renewable, CO 2 -free energy is increasingly important due to concerns over fuel security and the increase in atmospheric CO 2 concentration. Plants and cyanobacteria use oxygenic photosynthesis to convert sunlight and water into oxygen and chemical energy. A specific group of green microalgae and cyanobacteria, including the microalga Chlamydomonas reinhardtii, have evolved the additional ability to use sunlight for the production of molecular H 2 (1-5). The process of H 2 production has been described in detail for C. reinhardtii. Under anaerobic conditions two oxygen-sensitive FeFe-hydrogenases (HydA1 and HydA2) are induced (6) catalyzing the reduction of protons to molecular H 2 . There are two possible sources of electron supply: light energy is needed to generate protons and electrons from the water-splitting reaction (7) and, in parallel, a Photosystem II (PSII) 3 -independent process uses electrons originating from the breakdown of starch (8, 9). In both cases the reduced ferredoxin serves as electron donor for the FeFehydrogenases. Anaerobic culture conditions, required for H 2 production, can be achieved by bubbling inert gases through the culture or by sulfur depletion of the culture medium. During sulfur depletion the oxygen in the culture is consumed, and H 2 production can be observed (8).Recently, several molecular approaches have been applied to gain more detailed insights into the H 2 production metabolism (10 -16), to guide molecular genetics for fu...

No abstract

BackgroundChlamydomonas reinhardtii is widely accepted as a model organism regarding photosynthesis, circadian rhythm, cell mobility, phototaxis, and biotechnology. The complete annotation of the genome allows transcriptomic studies, however a new microarray platform was needed. Based on the completed annotation of Chlamydomonas reinhardtii a new microarray on an Agilent platform was designed using an extended JGI 3.1 genome data set which included 15000 transcript models.ResultsIn total 44000 probes were determined (3 independent probes per transcript model) covering 93% of the transcriptome. Alignment studies with the recently published AUGUSTUS 10.2 annotation confirmed 11000 transcript models resulting in a very good coverage of 70% of the transcriptome (17000). Following the estimation of 10000 predicted genes in Chlamydomonas reinhardtii our new microarray, nevertheless, covers the expected genome by 90-95%.ConclusionsTo demonstrate the capabilities of the new microarray, we analyzed transcript levels for cultures grown under nitrogen as well as sulfate limitation, and compared the results with recently published microarray and RNA-seq data. We could thereby confirm previous results derived from data on nutrient-starvation induced gene expression of a group of genes related to protein transport and adaptation of the metabolism as well as genes related to efficient light harvesting, light energy distribution and photosynthetic electron transport.

The microalga Rhodomonas salina is a widely used species for rearing live feed organisms in the aquaculture feed market. A species specific media is an essential step towards enhancing productivity and decreasing production costs for microalgae cultivation. However, relevant aspects of media composition such as nitrogen source and elemental ratio have not yet been characterized for this alga. This study aimed to optimize the following three aspects of culture media: 1) optimal ratio between nitrogen and phosphorus (N:P ratio); 2) preferred source of nitrogen; and 3) tolerance of R. salina towards free ammonia. To investigate this, we conducted a series of controlled laboratory experiments in shake flasks. Our experiments revealed a 45% increase in growth rate when an N:P ratio of 15:1 was used compared to the standard ratio of 25:1. Ammonium and nitrate were equally well accepted as a nitrogen source, however, a mix of ammonium and nitrate resulted in significant growth reduction. Free ammonia did not affect growth of the alga at the tested concentrations of up to 5 mg ammonia-nitrogen L− 1. We conclude that for optimal R. salina cultivation, an N:P ratio of 15:1 is strongly preferred, as it leads to a significant increase in growth rate. Further, media with a single source of nitrogen promote faster growth over media with mixed sources, and ammonium may safely be used as a nitrogen source, since R. salina tolerates certain levels of free ammonia. Overall, this work provides insights into the optimal cultivation conditions for R. salina, allowing for more efficient and reliable production of this relevant species.

The microalga Rhodomonas salina is a widely used species for rearing live feed organisms in the aquaculture feed market. A species-specific medium is an essential step towards enhancing productivity and decreasing production costs for microalgae cultivation. However, relevant aspects of medium composition such as nitrogen source and elemental ratio have not yet been characterized for this alga. This study aimed to optimize the following three aspects of culture media: 1) optimal ratio between nitrogen and phosphorus (N:P ratio); 2) preferred source of nitrogen; and 3) tolerance of R. salina towards free ammonia. To investigate this, we conducted a series of controlled laboratory experiments in shake flasks. Our experiments revealed a 45% increase in growth rate when an N:P ratio of 15:1 was used compared to the standard ratio of 25:1. Ammonium and nitrate were equally well accepted as a nitrogen source, however, a mix of ammonium and nitrate resulted in significant growth reduction. Free ammonia did not affect growth of the alga at the tested concentrations of up to 5 mg ammonia–nitrogen L−1. We conclude that for optimal R. salina cultivation, an N:P ratio of 15:1 is strongly preferred, as it leads to a significant increase in growth rate. Further, media with a single source of nitrogen promote faster growth over media with mixed sources, and ammonium may safely be used as a nitrogen source, since R. salina tolerates certain levels of free ammonia. Overall, this work provides insights into the optimal cultivation conditions for R. salina, allowing for more efficient and reliable production of this relevant species.

No abstract