Lipid production in the industrial microalga Nannochloropsis gaditana exceeds that of model algal species and can be maximized by nutrient starvation in batch culture. However, starvation halts growth, thereby decreasing productivity. Efforts to engineer N. gaditana strains that can accumulate biomass and overproduce lipids have previously met with little success. We identified 20 transcription factors as putative negative regulators of lipid production by using RNA-seq analysis of N. gaditana during nitrogen deprivation. Application of a CRISPR–Cas9 reverse-genetics pipeline enabled insertional mutagenesis of 18 of these 20 transcription factors. Knocking out a homolog of fungal Zn(II)₂Cys₆-encoding genes improved partitioning of total carbon to lipids from 20% (wild type) to 40–55% (mutant) in nutrient-replete conditions. Knockout mutants grew poorly, but attenuation of Zn(II)₂Cys₆ expression yielded strains producing twice as much lipid (~5.0 g m⁻² d⁻¹) as that in the wild type (~2.5 g m⁻² d⁻¹) under semicontinuous growth conditions and had little effect on growth.

中文翻译：

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Nannochloropsis gaditana中的脂质产生可通过减少单个转录调节子的表达而增加一倍

工业微藻Nannochloropsis gaditana中的脂质产量超过了模型藻种的脂质产量，并且可以通过分批培养中的营养缺乏来最大程度地提高脂质产量。但是，饥饿会阻止生长，从而降低生产力。以前，工程化能够积累生物质和过量生产脂质的加迪纳猪笼草菌株的努力迄今收效甚微。我们通过使用氮剥夺过程中加迪纳猪笼草的RNA-seq分析，确定了20个转录因子作为脂质产生的负调控因子。CRISPR–Cas9反向遗传学管线的应用使得这20个转录因子中的18个能够插入诱变。敲除真菌Zn（II）₂ Cys ₆的同源物编码基因可以在营养丰富的条件下将总碳到脂质的分配从20％（野生型）提高到40-55％（突变）。敲除突变体生长较差，但是Zn（II）₂ Cys ₆表达的减弱产生的菌株产生的脂质（〜5.0 gm ^-2 d ^-1）是野生型（〜2.5 gm ^-2 d ^-1）的两倍。半连续生长条件，对生长几乎没有影响。