Improved lipid productivity in Nannochloropsis gaditana in nitrogen-replete conditions by selection of pale green mutants

Michela Cecchin,Silvia Berteotti,Ivano Vigliante,Barbara Giovannone,Barbara Iadarola,Massimo E. Maffei,Stefania Paltrinieri,Matteo Ballottari,Massimo Delledonne

doi:10.1186/s13068-020-01718-8

Michela Cecchin, Silvia Berteotti + Show 7 more

Open Access

https://doi.org/10.1186/s13068-020-01718-8

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Abstract

BackgroundNannochloropsis gaditana is a photosynthetic unicellular microalgae considered one of the most interesting marine algae to produce biofuels and food additive due to its rapid growth rate and high lipid accumulation. Although microalgae are attractive platforms for solar energy bioconversion, the overall efficiency of photosynthesis is reduced due to the steep light gradient in photobioreactors. Moreover, accumulation of lipids in microalgae for biofuels production is usually induced in a two-phase cultivation process by nutrient starvation, with additional time and costs associated. In this work, a biotechnological approach was directed for the isolation of strains with improved light penetration in photobioreactor combined with increased lipids productivity.ResultsMutants of Nannochloropsis gaditana were obtained by chemical mutagenesis and screened for having both a reduced chlorophyll content per cell and increased affinity for Nile red, a fluorescent dye which binds to cellular lipid fraction. Accordingly, one mutant, called e8, was selected and characterized for having a 30% reduction of chlorophyll content per cell and an almost 80% increase of lipid productivity compared to WT in nutrient-replete conditions, with C16:0 and C18:0 fatty acids being more than doubled in the mutant. Whole-genome sequencing revealed mutations in 234 genes in e8 mutant among which there is a non-conservative mutation in the dgd1 synthase gene. This gene encodes for an enzyme involved in the biosynthesis of DGDG, one of the major lipids found in the thylakoid membrane and it is thus involved in chloroplast biogenesis. Lipid biosynthesis is strongly influenced by light availability in several microalgae species, including Nannochloropsis gaditana: reduced chlorophyll content per cell and more homogenous irradiance in photobioreactor is at the base for the increased lipid productivity observed in the e8 mutant.ConclusionsThe results herein obtained presents a promising strategy to produce algal biomass enriched in lipid fraction to be used for biofuel and biodiesel production in a single cultivation process, without the additional complexity of the nutrient starvation phase. Genome sequencing and identification of the mutations introduced in e8 mutant suggest possible genes responsible for the observed phenotypes, identifying putative target for future complementation and biotechnological application.

Highlights

Nannochloropsis gaditana is a photosynthetic unicellular microalgae considered one of the most interesting marine algae to produce biofuels and food additive due to its rapid growth rate and high lipid accumulation
We report a biotechnological approach by chemical mutagenesis to isolate N. gaditana strains with increased lipid productivity in absence of nutrient starvation
Mutagenesis and selection of mutant strains N. gaditana mutants were obtained by chemical mutagenesis, using the alkylating agent ethyl methane sulfonate (EMS) that inserts random single-point mutations (SNP) in the genome

Summary

Introduction

Nannochloropsis gaditana is a photosynthetic unicellular microalgae considered one of the most interesting marine algae to produce biofuels and food additive due to its rapid growth rate and high lipid accumulation. Species belonging to the genus Nannochloropsis are marine unicellular microalgae [9] considered among the most promising strains for cultivation in large scale systems, as open ponds or closed photobioreactors, for biodiesel production due to their fast growth rate, lipid accumulation (up to 65–70% of total dry weight) and ability to adapt to different irradiation conditions [5, 10, 11]. 30% of fatty acids accumulated in Nannochloropsis are polyunsaturated fatty acids among which eicosapentaenoic acid (EPA, 20:5ω3), one of the major omega-3 fatty acid reported to have positive effect in human health [12] This yellow green alga belongs to the class the pico-plankton Eustigmatophyceae, composed by species mainly living on the coasts. The availability of a genome sequence and transformation methods allow genetic engineering strategies to further improve this naturally productive species [18, 19]

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