Real-time quantitative PCR analysis of Nannochloropsis gaditana
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PROVIDER: PRJNA864746 | ENA |
REPOSITORIES: ENA
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Project description:Nannochloropsis gaditana cells were cultivated under continuous light as a control condition versus 3 different flashing light conditions (FL5, FL50 and FL500). This study is complementary to Lima et al. (2020) where physiology and photosynthetic capacity were assesed. Here we analyzed the mRNA expression level of key genes related to photoshynthesis, lipid and starch synthesis, and nitrogen assimilation. Total cell RNA was extracted from cultures of N. gaditana at a concentration of 1x106 cells/mL using the E.Z.N.A® total DNA/RNA isolation kit (Omega Bio-tek, Inc., USA). Reverse transcription reactions were performed using SuperScript™ IV VILO™ Master Mix with the enzyme ezDNase (Invitrogen™, ThermoFisher scientific, USA) according to the manufacturer’s protocol. The qPCR reactions were performed by mixing 5L of FastStart Universal SYBR Green Master mix (Rox) (Roche Molecular Systems, Inc., USA) with 1 L of primer mix (consisting of forward and the reverse primers) at a concentration of 300 nM and 4 L of the cDNA. The conditions of thermocycling were: 95 C for 600 s (preincubation), followed by 40 cycles of denaturation at 95 C for 10 s, annealing/extension at 60 C for 30 s. qPCR were performed in the LightCycler 96 Roche Life Science instrument.
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Project description:Nannochloropsis gaditana is a microalgae of the phylum eustigmatophyceae that has been reported from fresh and brackish waters. This species has been widely cultured, mainly directed to biofuel production, due to its capacity to produce and accumulates high amounts of lipids under different culturing conditions. Furthermore, nowadays N. gaditana is being used as fish and mollusc food in aquaculture facilities. Besides, microalgae are recognized protein producers, thus, are posited as an alternative protein source that could be very valuable in areas such as agri-food or biomedicine. In this sense, seas and oceans had showed their great potential for innovation, being the main focus of the initiative Blue Growth from EU. Thereby, marine biotechnology will provide of new pharmaceuticals or industrial products from marine biomass. In order to study the whole proteome of Nannochloropsis gaditana, a proteomic approach was initiated using fresh and atomized microalgae samples, as the main commercial forms. Around 7500 peptides were detected, getting 1950 protein identification hits. Qualitative and quantitative differences were analysed. The identified proteins were categorized according to gene ontology classification. In this study, it has been developed and described the first proteomic analysis of the microalgae Nannochloropsis gaditana, containing an important number of identified proteins that may have a relevant role in different agri-food and biomedical processes.
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