Abstract
The microalga Nannochloropsis oceanica is considered a promising platform for the sustainable production of high-value lipids and biofuel feedstocks. However, current lipid yields of N. oceanica are too low for economic feasibility. Gaining fundamental insights into the lipid metabolism of N. oceanica could open up various possibilities for the optimization of this species through genetic engineering. Therefore, the aim of this study was to discover novel genes associated with an elevated neutral lipid content. We constructed an insertional mutagenesis library of N. oceanica, selected high lipid mutants by five rounds of fluorescence-activated cell sorting, and identified disrupted genes using a novel implementation of a rapid genotyping procedure. One particularly promising mutant (HLM23) was disrupted in a putative APETALA2-like transcription factor gene. HLM23 showed a 40%-increased neutral lipid content, increased photosynthetic performance, and no growth impairment. Furthermore, transcriptome analysis revealed an upregulation of genes related to plastidial fatty acid biosynthesis, glycolysis and the Calvin-Benson-Bassham cycle in HLM23. Insights gained in this work can be used in future genetic engineering strategies for increased lipid productivity of Nannochloropsis.
Highlights
Microalgae have recently emerged as a promising platform for sus tainable production of lipids, pigments and other bioactive compounds
N. oceanica based on insertional mutagenesis and fluorescence-activated cell sorting (FACS)
An optimized genotyping meth odology based on the type IIS restriction endonuclease MmeI allowed us to identify the causative mutations in five mutants, which showed 12–59% increased neutral lipids (NLs) contents dry cell weight (DCW)− 1 relative to the wild type
Summary
Microalgae have recently emerged as a promising platform for sus tainable production of lipids, pigments and other bioactive compounds. Nannochloropsis oceanica, in particular, exhibits robust outdoor growth at relatively high growth rates and it can reach neutral lipid contents of up to 50% of the dry cell weight (Ma et al, 2014; Li et al, 2014; Meng et al, 2015) These exceptionally high neutral lipid contents are only reached when N. oceanica is exposed to a stress condition that is unfavorable for cell division, for instance nitrogen (N) deficiency (Rodolfi et al, 2009). About 50% of all predicted genes of the model strain N. oceanica IMET1 lack a functional annotation because the majority of the pre dicted proteins does not display sufficient sequence similarities with known proteins (Vieler et al, 2012; Wang et al, 2014) This poses a serious limitation for targeted modification of the metabolic network of this organism
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